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  • 1.
    Alberti, Esteban
    et al.
    Department of Neurobiology, International Center of Neurological Restoration, CIREN, Havana, Cuba..
    Los, Marek Jan
    Interfaculty Institute for Biochemistry, University of Tübingen, Germany; BioApplications Enterprises, Winnipeg, MB, Canada.
    Garcia, Rocio
    Department of Neurobiology, International Center of Neurological Restoration, CIREN, Havana, Cuba..
    Fraga, JL
    Department of Neurobiology, International Center of Neurological Restoration, CIREN, Havana, Cuba..
    Serrano, T.
    Department of Neurobiology, International Center of Neurological Restoration, CIREN, Havana, Cuba..
    Hernandez, E.
    Department of Neurobiology, International Center of Neurological Restoration, CIREN, Havana, Cuba..
    Klonisch, Thomas
    Department of Human Anatomy and Cell Sciences, and Manitoba Institute of Child Health, Winnipeg, Canada.
    Macías, R.
    Department of Neurobiology, International Center of Neurological Restoration, CIREN, Havana, Cuba..
    Martinez, L.
    Department of Neurobiology, International Center of Neurological Restoration, CIREN, Havana, Cuba..
    Castillo, L.
    Department of Neurobiology, International Center of Neurological Restoration, CIREN, Havana, Cuba..
    de la Cuétara, K.
    Department of Neurobiology, International Center of Neurological Restoration, CIREN, Havana, Cuba.
    Prolonged Survival and expression of neural markers by bone marrow-derived stem cells transplanted into brain lesions2009Ingår i: Medical Science Monitor, ISSN 1234-1010, E-ISSN 1643-3750, Vol. 15, nr 2, s. BR47-BR54Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    BACKGROUND: Bone marrow-derived stem cell transplantation is a potentially viable therapeutic option for the treatment of neurodegenerative disease. MATERIAL/METHODS: We have isolated bone marrow stem cells by standard method. We then evaluated the survival of rats' bone marrow mononuclear cells implanted in rats' brain. The cells were extracted from rats' femurs, and marked for monitoring purposes by adenoviral transduction with Green Fluorescent Protein (GFP). Labeled cells were implanted within the area of rats' striatum lesions that were induced a month earlier employing quinolinic acid-based method. The implants were phenotyped by monitoring CD34; CD38; CD45 and CD90 expression. Bone marrow stromal cells were extracted from rats' femurs and cultivated until monolayer bone marrow stromal cells were obtained. The ability of bone marrow stromal cells to express NGF and GDNF was evaluated by RT-PCR. RESULTS: Implanted cells survived for at least one month after transplantation and dispersed from the area of injection towards corpus callosum and brain cortex. Interestingly, passaged rat bone marrow stromal cells expressed NGF and GDNF mRNA. CONCLUSIONS: The bone marrow cells could be successfully transplanted to the brain either for the purpose of trans-differentiation, or for the expression of desired growth factors.

  • 2.
    Alizadeh, Javad
    et al.
    University of Manitoba, Canada.
    Zeki, Amir A.
    Centre Comparat Resp Biol and Med, CA USA.
    Mirzaei, Nima
    University of Manitoba, Canada.
    Tewary, Sandipan
    University of Manitoba, Canada.
    Rezaei Moghadam, Adel
    University of Manitoba, Canada; University of Manitoba, Canada.
    Glogowska, Aleksandra
    University of Manitoba, Canada.
    Nagakannan, Pandian
    University of Manitoba, Canada.
    Eftekharpour, Eftekhar
    University of Manitoba, Canada.
    Wiechec, Emilia
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för Logopedi, Audiologi och Otorhinolaryngologi. Linköpings universitet, Medicinska fakulteten.
    Gordon, Joseph W.
    University of Manitoba, Canada; University of Manitoba, Canada; University of Manitoba, Canada.
    Xu, Fred. Y.
    University of Manitoba, Canada; University of Manitoba, Canada.
    Field, Jared T.
    University of Manitoba, Canada.
    Yoneda, Ken Y.
    Centre Comparat Resp Biol and Med, CA USA.
    Kenyon, Nicholas J.
    Centre Comparat Resp Biol and Med, CA USA.
    Hashemi, Mohammad
    Zehedan University of Medical Science, Iran.
    Hatch, Grant M.
    University of Manitoba, Canada; University of Manitoba, Canada.
    Hombach-Klonisch, Sabine
    University of Manitoba, Canada.
    Klonisch, Thomas
    University of Manitoba, Canada.
    Ghavami, Saeid
    University of Manitoba, Canada; University of Manitoba, Canada; Shiraz University of Medical Science, Iran.
    Mevalonate Cascade Inhibition by Simvastatin Induces the Intrinsic Apoptosis Pathway via Depletion of Isoprenoids in Tumor Cells2017Ingår i: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 7, artikel-id 44841Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The mevalonate (MEV) cascade is responsible for cholesterol biosynthesis and the formation of the intermediate metabolites geranylgeranylpyrophosphate (GGPP) and farnesylpyrophosphate (FPP) used in the prenylation of proteins. Here we show that the MEV cascade inhibitor simvastatin induced significant cell death in a wide range of human tumor cell lines, including glioblastoma, astrocytoma, neuroblastoma, lung adenocarcinoma, and breast cancer. Simvastatin induced apoptotic cell death via the intrinsic apoptotic pathway. In all cancer cell types tested, simvastatin-induced cell death was not rescued by cholesterol, but was dependent on GGPP-and FPP-depletion. We confirmed that simvastatin caused the translocation of the small Rho GTPases RhoA, Cdc42, and Rac1/2/3 from cell membranes to the cytosol in U251 (glioblastoma), A549 (lung adenocarcinoma) and MDA-MB231( breast cancer). Simvastatin-induced Rho-GTP loading significantly increased in U251 cells which were reversed with MEV, FPP, GGPP. In contrast, simvastatin did not change Rho-GTP loading in A549 and MDA-MB-231. Inhibition of geranylgeranyltransferase I by GGTi-298, but not farnesyltransferase by FTi-277, induced significant cell death in U251, A549, and MDA-MB-231. These results indicate that MEV cascade inhibition by simvastatin induced the intrinsic apoptosis pathway via inhibition of Rho family prenylation and depletion of GGPP, in a variety of different human cancer cell lines.

  • 3.
    Alvarez-Rodriguez, Manuel
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Vicente-Carrillo, Alejandro
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Exogenous Individual Lecithin-Phospholipids (Phosphatidylcholine and Phosphatidylglycerol) Cannot Prevent the Oxidative Stress Imposed by Cryopreservation of Boar Sperm.2017Ingår i: Journal of veterinary medicine and surgery, ISSN 2574-2868, Vol. 1, nr 1Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Objective: Despite the use of high proportions of the chemically undefined lipoprotein/phospholipid-rich egg-yolk in extenders, boar sperm are highly sensitive to cooling, which induces ROS generation and disrupts the plasma membrane.

    Here, we studied whether replacement of hen egg-yolk by commercially defined lecithin phospholipids, derived from egg (LPGE: phosphatidyl glycerol, LPCE: phosphatidyl choline) or soybean (LPCS: phosphatidyl choline), could individually ameliorate such oxidative effects during cryopreservation of ejaculated (sperm rich fraction, SRF) or of cauda-epididymal sperm, retrieved post-mortem from the same males.

    Methods: A conventional extender (lactose buffer, with 20% egg-yolk, 0.5% OEP and 3% glycerol) was used as control. Cryodamage was assessed as loss of sperm motility, membrane and acrosome intactness, early membrane destabilization changes, mitochondrial potential, superoxide and ROS production, to finally determine lipid peroxidation (LPO) using specific probes.

    Results and conclusion: In general, the exogenous phospholipids assayed were unable of maintaining neither sperm motility nor viability post-thaw compared to controls, owing to increased ROS production and lipid peroxidation. In our study, mitochondrial superoxide production resulted in very high levels for all groups, whereas both ROS production and lipid peroxidation were reduced in the control group, containing emulsified hen egg yolk. Further studies using various dosage and combination of LPCS should be followed for their eventual protective effect.

    Keywords: Cryodamage; Sperm; Boar; Mitochondrial activation; Mitochondrial superoxide; ROS production; Lipid peroxidation

  • 4.
    Amirhosseini, Mehdi
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Medicinska fakulteten.
    Aseptic Loosening of Orthopedic Implants: Osteoclastogenesis Regulation and Potential Therapeutics2019Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Aseptic loosening is the main cause of failure of orthopedic prostheses. With no pharmaceuticals to prevent or mitigate periprosthetic bone degradation, a surgery to replace the loose implant with a new one is the only choice to restore patients’ function. Most studies on mechanisms for aseptic loosening investigate wear debris particle-induced osteolysis. However, pathological loading conditions around unstable implants can also trigger osteoclast differentiation and bone loss.

    In the first study, global gene expression changes induced by mechanical instability of implants, and by titanium particles were compared in a validated rat model for aseptic loosening. Microarray analysis showed that similar signaling pathways and gene expression patterns are involved in particle- and instability-induced periprosthetic osteolysis with an early onset innate immune response as a hallmark of osteolysis induced by mechanical instability.

    Further, effects of potential therapeutics on restriction of excessive osteoclast differentiation were evaluated. Wnt signaling pathway is known to regulate bone remodeling. In the second study, effects of inactivation of glycogen synthase kinase 3 beta (GSK-3β), a negative regulator of canonical Wnt signaling, on instability-induced periprosthetic osteolysis were examined using our rat model for aseptic loosening. Inhibition of GSK-3β led to a decrease in osteoclast numbers in the periprosthetic bone tissue exposed to mechanical instability while osteoblast perimeter showed an increase. This was accompanied by higher bone volume fraction (BV/TV) in animals treated with the GSK-3β inhibitor.

    In the third study, potential beneficial effects of two selective inhibitors of cyclindependent kinase 8/19 (CDK8/19) on bone tissue were evaluated. CDK8/19 is a Mediator complex-associated transcriptional regulator involved in several signaling pathways. CDK8/19 inhibitors, mainly under investigation as treatments for tumors, are reported to enhance osteoblast differentiation and bone formation. We show in this study, for the first time, that inhibition of CDK8/19 led to marked suppression of osteoclast differentiation from bone marrow macrophages in vitro through disruption of the RANK signaling. In mouse primary osteoblasts downregulation of osteopontin mRNA, a negative regulator of mineralization, together with increased alkaline phosphatase activity and calcium deposition indicated that osteoblast mineralization was promoted by CDK8/19 inhibition. Moreover, local administration of a CDK8/19 inhibitor promoted cancellous bone regeneration in a rat model for bone healing.

    These studies contribute to better understanding of mechanisms behind mechanical instability-induced periprosthetic osteolysis and propose potential therapeutics to restrict bone loss with effects on both osteoclasts and osteoblasts.

    Delarbeten
    1. Mechanical instability and titanium particles induce similar transcriptomic changes in a rat model for periprosthetic osteolysis and aseptic loosening
    Öppna denna publikation i ny flik eller fönster >>Mechanical instability and titanium particles induce similar transcriptomic changes in a rat model for periprosthetic osteolysis and aseptic loosening
    2017 (Engelska)Ingår i: Bone Reports, ISSN 2352-1872, Vol. 7, s. 17-25Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Wear debris particles released from prosthetic bearing surfaces and mechanical instability of implants are two main causes of periprosthetic osteolysis. While particle-induced loosening has been studied extensively, mechanisms through which mechanical factors lead to implant loosening have been less investigated. This study compares the transcriptional profiles associated with osteolysis in a rat model for aseptic loosening, induced by either mechanical instability or titanium particles. Rats were exposed to mechanical instability or titanium particles. After 15 min, 3, 48 or 120 h from start of the stimulation, gene expression changes in periprosthetic bone tissue was determined by microarray analysis. Microarray data were analyzed by PANTHER Gene List Analysis tool and Ingenuity Pathway Analysis (IPA). Both types of osteolytic stimulation led to gene regulation in comparison to unstimulated controls after 3, 48 or 120 h. However, when mechanical instability was compared to titanium particles, no gene showed a statistically significant difference (fold change = ± 1.5 and adjusted p-value = 0.05) at any time point. There was a remarkable similarity in numbers and functional classification of regulated genes. Pathway analysis showed several inflammatory pathways activated by both stimuli, including Acute Phase Response signaling, IL-6 signaling and Oncostatin M signaling. Quantitative PCR confirmed the changes in expression of key genes involved in osteolysis observed by global transcriptomics. Inflammatory mediators including interleukin (IL)-6, IL-1ß, chemokine (C-C motif) ligand (CCL)2, prostaglandin-endoperoxide synthase (Ptgs)2 and leukemia inhibitory factor (LIF) showed strong upregulation, as assessed by both microarray and qPCR. By investigating genome-wide expression changes we show that, despite the different nature of mechanical implant instability and titanium particles, osteolysis seems to be induced through similar biological and signaling pathways in this rat model for aseptic loosening. Pathways associated to the innate inflammatory response appear to be a major driver for osteolysis. Our findings implicate early restriction of inflammation to be critical to prevent or mitigate osteolysis and aseptic loosening of orthopedic implants.

    Ort, förlag, år, upplaga, sidor
    Elsevier, 2017
    Nyckelord
    Aseptic loosening; Implant; Instability; Microarray; Wear debris
    Nationell ämneskategori
    Cell- och molekylärbiologi Ortopedi
    Identifikatorer
    urn:nbn:se:liu:diva-146297 (URN)10.1016/j.bonr.2017.07.003 (DOI)28795083 (PubMedID)
    Tillgänglig från: 2018-04-07 Skapad: 2018-04-07 Senast uppdaterad: 2019-03-08
    2. GSK-3 beta inhibition suppresses instability-induced osteolysis by a dual action on osteoblast and osteoclast differentiation
    Öppna denna publikation i ny flik eller fönster >>GSK-3 beta inhibition suppresses instability-induced osteolysis by a dual action on osteoblast and osteoclast differentiation
    Visa övriga...
    2018 (Engelska)Ingår i: Journal of Cellular Physiology, ISSN 0021-9541, E-ISSN 1097-4652, Vol. 233, nr 3, s. 2398-2408Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Currently, there are no medications available to treat aseptic loosening of orthopedic implants. Using osteoprotegerin fusion protein (OPG-Fc), we previously blocked instability-induced osteoclast differentiation and peri-prosthetic osteolysis. Wnt/beta-catenin signaling, which regulates OPG secretion from osteoblasts, also modulates the bone tissue response to mechanical loading. We hypothesized that activating Wnt/beta-catenin signaling by inhibiting glycogen synthase kinase-3 beta (GSK-3 beta) would reduce instability-induced bone loss through regulation of both osteoblast and osteoclast differentiation. We examined effects of GSK-3 beta inhibition on regulation of RANKL and OPG in a rat model of mechanical instability-induced peri-implant osteolysis. The rats were treated daily with a GSK-3 beta inhibitor, AR28 (20 mg/kg bw), for up to 5 days. Bone tissue and blood serum were assessed by qRT-PCR, immunohistochemistry, and ELISA on days 3 and 5, and by micro-CT on day 5. After 3 days of treatment with AR28, mRNA levels of beta-catenin, Runx2, Osterix, Col1 alpha 1, and ALP were increased leading to higher osteoblast numbers compared to vehicle-treated animals. BMP-2 and Wnt16 mRNA levels were downregulated by mechanical instability and this was rescued by GSK-3 beta inhibition. Osteoclast numbers were decreased significantly after 3 days of GSK-3 beta inhibition, which correlated with enhanced OPG mRNA expression. This was accompanied by decreased serum levels of TRAP5b on days 3 and 5. Treatment with AR28 upregulated osteoblast differentiation, while osteoclastogenesis was blunted, leading to increased bone mass by day 5. These data suggest that GSK-3 beta inactivation suppresses osteolysis through regulating both osteoblast and osteoclast differentiation in a rat model of instability-induced osteolysis.

    Ort, förlag, år, upplaga, sidor
    WILEY, 2018
    Nyckelord
    bone implant; GSK-3 beta; mechanical instability; osteolysis; Wnt signaling
    Nationell ämneskategori
    Farmakologi och toxikologi
    Identifikatorer
    urn:nbn:se:liu:diva-148660 (URN)10.1002/jcp.26111 (DOI)000433519300056 ()28731198 (PubMedID)
    Anmärkning

    Funding Agencies|VINNOVA [2012-04409]; National Institutes of Health [AR056802]; Vetenskapsradet [K2014-7X-22506-01-3]; Swedish Research Council; Swedish Governmental Agency for Innovation Systems

    Tillgänglig från: 2018-06-18 Skapad: 2018-06-18 Senast uppdaterad: 2019-04-08
    3. Cyclin-dependent kinase 8/19 inhibition suppresses osteoclastogenesis by downregulating RANK and promotes osteoblast mineralization and cancellous bone healing.
    Öppna denna publikation i ny flik eller fönster >>Cyclin-dependent kinase 8/19 inhibition suppresses osteoclastogenesis by downregulating RANK and promotes osteoblast mineralization and cancellous bone healing.
    Visa övriga...
    2019 (Engelska)Ingår i: Journal of Cellular Physiology, ISSN 0021-9541, E-ISSN 1097-4652, Vol. 234, nr 9, s. 16503-16516Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Cyclin-dependent kinase 8 (CDK8) is a mediator complex-associated transcriptional regulator that acts depending on context and cell type. While primarily under investigation as potential cancer therapeutics, some inhibitors of CDK8-and its paralog CDK19-have been reported to affect the osteoblast lineage and bone formation. This study investigated the effects of two selective CDK8/19 inhibitors on osteoclastogenesis and osteoblasts in vitro, and further evaluated how local treatment with a CDK8/19 inhibitor affects cancellous bone healing in rats. CDK8/19 inhibitors did not alter the proliferation of neither mouse bone marrow-derived macrophages (BMMs) nor primary mouse osteoblasts. Receptor activator of nuclear factor κΒ (NF-κB) ligand (RANKL)-induced osteoclastogenesis from mouse BMMs was suppressed markedly by inhibition of CDK8/19, concomitant with reduced tartrate-resistant acid phosphatase (TRAP) activity and C-terminal telopeptide of type I collagen levels. This was accompanied by downregulation of PU.1, RANK, NF-κB, nuclear factor of activated T-cells 1 (NFATc1), dendritic cell-specific transmembrane protein (DC-STAMP), TRAP, and cathepsin K in RANKL-stimulated BMMs. Downregulating RANK and its downstream signaling in osteoclast precursors enforce CDK8/19 inhibitors as anticatabolic agents to impede excessive osteoclastogenesis. In mouse primary osteoblasts, CDK8/19 inhibition did not affect differentiation but enhanced osteoblast mineralization by promoting alkaline phosphatase activity and downregulating osteopontin, a negative regulator of mineralization. In rat tibiae, a CDK8/19 inhibitor administered locally promoted cancellous bone regeneration. Our data indicate that inhibitors of CDK8/19 have the potential to develop into therapeutics to restrict osteolysis and enhance bone regeneration.

    Nyckelord
    CDK8, RANK, osteoblasts, osteoclasts
    Nationell ämneskategori
    Cell- och molekylärbiologi Läkemedelskemi
    Identifikatorer
    urn:nbn:se:liu:diva-154927 (URN)10.1002/jcp.28321 (DOI)000470174200186 ()30793301 (PubMedID)
    Anmärkning

    Funding agencies: Vetenskapsradet [521-2013-2593, 2016-06097, K2015-99x-10363-23-4, 2016-01822]; Swedish Research Council

    Tillgänglig från: 2019-03-05 Skapad: 2019-03-05 Senast uppdaterad: 2019-07-03
  • 5.
    Backteman, Karin
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för inflammationsmedicin. Linköpings universitet, Hälsouniversitetet. Östergötlands Läns Landsting, Diagnostikcentrum, Klinisk immunologi och transfusionsmedicin.
    T Cells and NK Cells in Coronary Artery Disease: Longitudinal and methodological studies in humans2014Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Coronary artery disease (CAD) is the leading cause of death worldwide and most often due to atherosclerosis. Atherosclerosis is a chronic inflammatory process that involves the arteries, inclouding those that supply blood to the heart muscle. Although inflammation is an important contributing factor to atherosclerosis, the mechanisms are not fully understood. One mechanism contributing to atherogenesis may involve some infectious microorganisms such as cytomegalovirus (CMV). In atherosclerosis, the arterial wall becomes infiltrated with lipids followed by different types of leukocytes and inflammatory mediators (atherogenesis). Leukocytes recirculate continuously between the blood and lymphoid organs, such as lymph nodes, where the adaptive immune response is started and regulated.

    The general aim of this thesis was to increase the understanding of associations between lymphocyte populations and different conditions of CAD (unstable and stable). To assess changes over time, a longitudinal follow up design was mostly used. Therefore, also perspectives of longitudinal variation were included in the thesis.

    Paper I showed that flow cytometric evaluation of lymphocyte populations is a robust technique that can be used in longitudinal studies, both in clinical and research settings. It was also shown that the time of sampling over the year did not have a major impact on the findings.

    In paper II, thoracic lymph nodes were investigated to assess whether CAD-associated changes were more prominent in comparison with blood. As expected, there were several major differences in lymphocyte composition between lymph nodes and blood. However, the analysis of thoracic lymph nodes did not reveal any further changes that were not detected in blood. Thus, blood is still the most reliable compartment for studies of lymphocyte populations in CAD since it is not possible to examine the local findings in the artery wall.

    Natural killer (NK) cells are innate lymphocytes with both regulatory and effector functions. In paper II and III we confirmed previous findings that CAD patients have lower proportions of NK cells in blood. However, the NK subtype and cytokine profile (paper III, measured by subtype markers and intra-cellular cytokine staining) did not differ between patients and controls. During a 12-month follow-up, the proportions of NK cells increased, although not in all patients. Failure to reconstitute NK cell levels was associated with several components of the metabolic syndrome and with a persistent low-grade inflammation as measured by plasma IL-6 levels. The findings support the notion of a protective role for NK cells in inflammation.

    CD4+ but not CD8+ T cells were significantly increased in patients with both unstable and stable conditions compared with healthy individuals (paper IV). Subpopulations of CD4+ T cells (CD4+CD28null) have previously been associated with CAD. However, we show that CD28null and CD28null57+ cells within the CD4+ and CD8+ T cell populations were similar in CAD patients and healthy controls. Instead, CMV seropositivity was the major determinant of expanded CD28null and CD57+ T cell fractions in both patients and healthy individuals. During the 1 year follow up the proportion of CD4+CD28null and CD8+CD28null cells increased in patients, which may reflect an accelerated immunological ageing occurring after the cardiac event.

    Delarbeten
    1. Biological and methodological variation of lymphocyte subsets in blood of human adults
    Öppna denna publikation i ny flik eller fönster >>Biological and methodological variation of lymphocyte subsets in blood of human adults
    2007 (Engelska)Ingår i: JIM - Journal of Immunological Methods, ISSN 0022-1759, E-ISSN 1872-7905, Vol. 322, nr 1-2, s. 20-27Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Although lymphocyte populations are often monitored over time, information about the biological variation over time is limited. Three-colour-flow cytometry was used to investigate the biological and methodological variation of lymphocyte populations in blood. Fifteen healthy individuals (11 females and 4 males) were longitudinally monitored for 2-8 years. Blood samples were drawn monthly when possible. In total, 493 observations were included. Absolute counts and proportions were determined for T-cells (CD3+), T-helper cells (CD3+ CD4+), cytolytic T-cells (CD3+ CD8+), B-cells (CD3- CD19+) and NK-cells (CD3- CD16+/56+). As to variation over the year, ANOVA testing showed only a minor monthly variation for absolute counts of the CD8+ population (p < 0.05) for October compared with June and July, whereas no significant differences were found for the other populations or in the proportions of lymphocyte subsets. Although lower than the longitudinal variation, the methodological variation, expressed as coefficient of variation (CV %), was in a similar range as the variation over time, indicating that the normal biological variation should not be overestimated, while the methodological inter-assay should be taken into consideration in longitudinal studies or monitoring of patients. © 2007 Elsevier B.V. All rights reserved.

    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-41169 (URN)10.1016/j.jim.2007.01.021 (DOI)55291 (Lokalt ID)55291 (Arkivnummer)55291 (OAI)
    Tillgänglig från: 2009-10-10 Skapad: 2009-10-10 Senast uppdaterad: 2017-12-13
    2. Lymphocyte Subpopulations in Lymph Nodes and Peripheral Blood: A Comparison between Patients with Stable Angina and Acute Coronary Syndrome
    Öppna denna publikation i ny flik eller fönster >>Lymphocyte Subpopulations in Lymph Nodes and Peripheral Blood: A Comparison between Patients with Stable Angina and Acute Coronary Syndrome
    2012 (Engelska)Ingår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 7, nr 3Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Objective: Atherosclerosis is characterized by a chronic inflammatory response involving activated T cells and impairment of natural killer (NK) cells. An increased T cell activity has been associated with plaque instability and risk of acute cardiac events. Lymphocyte analyses in blood are widely used to evaluate the immune status. However, peripheral blood contains only a minor proportion of lymphocytes. In this study, we hypothesized that thoracic lymph nodes from patients with stable angina (SA) and acute coronary syndrome (ACS) might add information to peripheral blood analyses. less thanbrgreater than less thanbrgreater thanMethods: Peripheral blood and lymph nodes were collected during coronary by-pass surgery in 13 patients with SA and 13 patients with ACS. Lymphocyte subpopulations were assessed by flow cytometry using antibodies against CD3, CD4, CD8, CD19, CD16/56, CD25, Foxp3, CD69, HLA-DR, IL-18 receptor (R) and CCR4. less thanbrgreater than less thanbrgreater thanResults: Lymph nodes revealed a lymphocyte subpopulation profile substantially differing from that in blood including a higher proportion of B cells, lower proportions of CD8(+) T cells and NK cells and a 2-fold higher CD4/CD8 ratio. CD4(+)CD69(+) cells as well as Foxp3(+) regulatory T cells were markedly enriched in lymph nodes (p andlt; 0.001) while T helper 1-like (CD4(+)IL-18R(+)) cells were more frequent in blood (p andlt; 0.001). The only significant differences between ACS and SA patients involved NK cells that were reduced in the ACS group. However, despite being reduced, the NK cell fraction in ACS patients contained a significantly higher proportion of IL-18R(+) cells compared with SA patients (p andlt; 0.05). less thanbrgreater than less thanbrgreater thanConclusion: There were several differences in lymphocyte subpopulations between blood and lymph nodes. However, the lymphocyte perturbations in peripheral blood of ACS patients compared with SA patients were not mirrored in lymph nodes. The findings indicate that lymph node analyses in multivessel coronary artery disease may not reveal any major changes in the immune response that are not detectable in blood.

    Ort, förlag, år, upplaga, sidor
    Public Library of Science, 2012
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-77542 (URN)10.1371/journal.pone.0032691 (DOI)000303005000033 ()
    Anmärkning
    Funding Agencies|Swedish Heart-Lung Foundation|20090489|Swedish Research Council|2008-2282|Tillgänglig från: 2012-05-25 Skapad: 2012-05-22 Senast uppdaterad: 2017-12-07
    3. Natural killer (NK) cell deficit in coronary artery disease: no aberrations in phenotype but sustained reduction of NK cells is associated with low-grade inflammation
    Öppna denna publikation i ny flik eller fönster >>Natural killer (NK) cell deficit in coronary artery disease: no aberrations in phenotype but sustained reduction of NK cells is associated with low-grade inflammation
    2014 (Engelska)Ingår i: Clinical and Experimental Immunology, ISSN 0009-9104, E-ISSN 1365-2249, Vol. 175, nr 1, s. 104-112Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Although reduced natural killer (NK) cell levels have been reported consistently in patients with coronary artery disease (CAD), the clinical significance and persistence of this immune perturbation is not clarified. In this study we characterized the NK cell deficit further by determining (i) differentiation surface markers and cytokine profile of NK cell subsets and (ii) ability to reconstitute NK cell levels over time. Flow cytometry was used to analyse NK cell subsets and the intracellular cytokine profile in 31 patients with non-ST elevation myocardial infarction (non-STEMI), 34 patients with stable angina (SA) and 37 healthy controls. In blood collected prior to coronary angiography, the proportions of NK cells were reduced significantly in non-STEMI and SA patients compared with controls, whereas NK cell subset analyses or cytokine profile measurements did not reveal any differences across groups. During a 12-month follow-up, the proportions of NK cells increased, although not in all patients. Failure to reconstitute NK cell levels was associated with several components of metabolic syndrome. Moreover, interleukin (IL)-6 levels remained high in patients with sustained NK cell deficit, whereas a decline in IL-6 (P < 0·001) was seen in patients with a pronounced increase in NK cells. In conclusion, we found no evidence that reduction of NK cells in CAD patients was associated with aberrations in NK cell phenotype at any clinical stage of the disease. Conversely, failure to reconstitute NK cell levels was associated with a persistent low-grade inflammation, suggesting a protective role of NK cells in CAD.

    Ort, förlag, år, upplaga, sidor
    Wiley-Blackwell, 2014
    Nyckelord
    coronary artery disease; cytokines; inflammation; leukocytes; natural killer cell
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-103363 (URN)10.1111/cei.12210 (DOI)000329165400012 ()24298947 (PubMedID)
    Tillgänglig från: 2014-01-17 Skapad: 2014-01-17 Senast uppdaterad: 2017-12-06Bibliografiskt granskad
    4. Cytomegalovirus seropositivity is a major determinant of CD28null T cell expansion in patients with coronary artery disease
    Öppna denna publikation i ny flik eller fönster >>Cytomegalovirus seropositivity is a major determinant of CD28null T cell expansion in patients with coronary artery disease
    2014 (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    Objective: Accumulation of CD4+28null cells, with a proinflammatory and senescent phenotype, has been associated with unstable conditions of coronary artery disease (CAD). Human cytomegalovirus (HCMV) is known to exert profound effects on T cells, including loss of CD28. Here, we longitudinally assessed the proportions of CD28null and CD28nullCD57+ cells in CD4+ and CD8+ T cell populations of patients with CAD and related the findings to HCMV seropositivity.

    Methods: HCMV antibody levels and expression of CD28 and CD57 on CD4+ and CD8+ T cells were analysed in 31 patients with acute coronary syndrome (ACS), 34 patients with stable angina (SA) and 37 healthy controls. Samples were taken prior to 34 coronary angiography and after 3 and 12 months. In a subsample, HCMV-specific IFN-γ and  TNF production was assessed ex vivo.

    Results: Increased proportions of CD4+CD28null, but not CD8+CD28null cells, were significantly associated with presence of CAD. Significant increases in CD28null 37 and CD28nullCD57+ cells occurred within CD4+ and CD8+ T cell compartments in both ACS and SA patients during 12-month follow-up. HCMV was the major determinant of CD28null and CD28nullCD57+ T cell levels in both patients and controls (p <0.001). There were no obvious signs of CMV reactivation in patients.

    Conclusion: HCMV was a major determinant of the presence of CD28null and CD28nullCD57+ T cells in patients with CAD, independent of clinical stage. Findings also indicate that HCMV might have a large impact on the T cell aging process that occurred in patients after a cardiac event.

    Nyckelord
    Coronary artery disease, acute coronary syndrome, CD28null T cells, CD57+ 49 T cells, Human cytomegalovirus
    Nationell ämneskategori
    Klinisk medicin Immunologi
    Identifikatorer
    urn:nbn:se:liu:diva-111049 (URN)
    Tillgänglig från: 2014-10-06 Skapad: 2014-10-06 Senast uppdaterad: 2015-03-25Bibliografiskt granskad
  • 6.
    Bahrampour, Shahrzad
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för mikrobiologi och molekylär medicin. Linköpings universitet, Medicinska fakulteten.
    Genetic mechanisms regulating proliferation and cell specification in the Drosophila embryonic CNS2017Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The central nervous system (CNS) consists of an enormous number of cells, and large cellular variance, integrated into an elaborate network. The CNS is the most complex animal organ, and therefore its establishment must be controlled by many different genetic programs. Considering the high level of complexity in the human CNS, addressing issues related to human neurodevelopment represents a major challenge. Since comparative studies have revealed that neurodevelopmental programs are well conserved through evolution, on both the genetic and functional levels, studies on invertebrate neurodevelopmental programs are often translatable to vertebrates. Indeed, the basis of our current knowledge about vertebrate CNS development has been greatly aided by studies on invertebrates, and in particular on the Drosophila melanogaster (fruit fly) model system.

    This thesis attempted to identify novel genes regulating neural cell specification and proliferation in the CNS, using the Drosophila model system. Moreover, I aimed to address how those genes govern neural progenitor cells (neuroblasts; NBs) to obtain/maintain their stemness identity and proliferation capacity, and how they drive NBs through temporal windows and series of programmed asymmetric division, which gradually reduces their stemness identity in favor of neural differentiation, resulting in appropriate lineage progression. In the first project, we conducted a forward genetic screen in Drosophila embryos, aimed at isolating genes involved in regulation of neural proliferation and specification, at the single cell resolution. By taking advantage of the restricted expression of the neuropeptide FMRFa in the last-born cell of the NB lineage 5-6T, the Ap4 neuron, we could monitor the entire lineage progression. This screen succeeded in identifying 43 novel genes controlling different aspects of CNS development. One of the genes isolated, Ctr9, displayed extra Ap4/FMRFa neurons. Ctr9 encodes a component of the RNA polymerase II complex Paf1, which is involved in a number of transcriptional processes. The Paf1C, including Ctr9, is highly conserved from yeast to human, and in the past couple of years, its importance for transcription has become increasingly appreciated. However, studies in the Drosophila system have been limited. In the screen, we isolated the first mutant of Drosophila Ctr9 and conducted the first detailed phenotypic study on its function in the Drosophila embryonic CNS. Loss of function of Ctr9 leads to extra NB numbers, higher proliferation ratio and lower expression of neuropeptides. Gene expression analysis identified several other genes regulated by Ctr9, which may explain the Ctr9 mutant phenotypes. In summary, we identified Ctr9 as an essential gene for proper CNS development in Drosophila, and this provides a platform for future study on the Drosophila Paf1C. Another interesting gene isolated in the screen was worniou (wor), a member of the Snail family of transcription factors. In contrast to Ctr9, whichdisplayed additional Ap4/FMRFa neurons, wor mutants displayed a loss of these neurons. Previous studies in our group have identified many genes acting to stop NB lineage progression, but how NBs are pushed to proliferate and generate their lineages was not well known. Since wor may constitute a “driver” of proliferation, we decided to study it further. Also, we identified five other transcription factors acting together with Wor as pro-proliferative in both NBs and their daughter cells. These “drivers” are gradually replaced by the previously identified late-acting “stoppers.” Early and late factors regulate each other and the cell cycle, and thereby orchestrate proper neural lineage progression.

    Delarbeten
    1. Novel Genes Involved in Controlling Specification of Drosophila FMRFamide Neuropeptide Cells
    Öppna denna publikation i ny flik eller fönster >>Novel Genes Involved in Controlling Specification of Drosophila FMRFamide Neuropeptide Cells
    Visa övriga...
    2015 (Engelska)Ingår i: Genetics, ISSN 0016-6731, E-ISSN 1943-2631, Vol. 200, nr 4, s. 1229-1244Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The expression of neuropeptides is often extremely restricted in the nervous system, making them powerful markers for addressing cell specification . In the developing Drosophila ventral nerve cord, only six cells, the Ap4 neurons, of some 10,000 neurons, express the neuropeptide FMRFamide (FMRFa). Each Ap4/FMRFa neuron is the last-born cell generated by an identifiable and well-studied progenitor cell, neuroblast 5-6 (NB5-6T). The restricted expression of FMRFa and the wealth of information regarding its gene regulation and Ap4 neuron specification makes FMRFa a valuable readout for addressing many aspects of neural development, i.e., spatial and temporal patterning cues, cell cycle control, cell specification, axon transport, and retrograde signaling. To this end, we have conducted a forward genetic screen utilizing an Ap4-specific FMRFa-eGFP transgenic reporter as our readout. A total of 9781 EMS-mutated chromosomes were screened for perturbations in FMRFa-eGFP expression, and 611 mutants were identified. Seventy-nine of the strongest mutants were mapped down to the affected gene by deficiency mapping or whole-genome sequencing. We isolated novel alleles for previously known FMRFa regulators, confirming the validity of the screen. In addition, we identified novel essential genes, including several with previously undefined functions in neural development. Our identification of genes affecting most major steps required for successful terminal differentiation of Ap4 neurons provides a comprehensive view of the genetic flow controlling the generation of highly unique neuronal cell types in the developing nervous system.

    Ort, förlag, år, upplaga, sidor
    Genetics Society of America, 2015
    Nyckelord
    Drosophila; CNS development; neural cell fate specification; forward genetic screening; FMRFamide
    Nationell ämneskategori
    Klinisk medicin
    Identifikatorer
    urn:nbn:se:liu:diva-121318 (URN)10.1534/genetics.115.178483 (DOI)000359917000020 ()26092715 (PubMedID)
    Tillgänglig från: 2015-09-16 Skapad: 2015-09-14 Senast uppdaterad: 2019-03-13Bibliografiskt granskad
    2. Ctr9, a Key Component of the Paf1 Complex, Affects Proliferation and Terminal Differentiation in the Developing Drosophila Nervous System
    Öppna denna publikation i ny flik eller fönster >>Ctr9, a Key Component of the Paf1 Complex, Affects Proliferation and Terminal Differentiation in the Developing Drosophila Nervous System
    2016 (Engelska)Ingår i: G3: Genes, Genomes, Genetics, ISSN 2160-1836, E-ISSN 2160-1836, Vol. 6, nr 10, s. 3229-3239Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The Paf1 protein complex (Paf1C) is increasingly recognized as a highly conserved and broadly utilized regulator of a variety of transcriptional processes. These include the promotion of H3K4 and H3K36 trimethylation, H2BK123 ubiquitination, RNA Pol II transcriptional termination, and also RNA-mediated gene silencing. Paf1C contains five canonical protein components, including Paf1 and Ctr9, which are critical for overall complex integrity, as well as Rtf1, Leo1, and Cdc73/Parafibromin(Hrpt2)/Hyrax. In spite of a growing appreciation for the importance of Paf1C from yeast and mammalian studies, there has only been limited work in Drosophila. Here, we provide the first detailed phenotypic study of Ctr9 function in Drosophila. We found that Ctr9 mutants die at late embryogenesis or early larval life, but can be partly rescued by nervous system reexpression of Ctr9. We observed a number of phenotypes in Ctr9 mutants, including increased neuroblast numbers, increased nervous system proliferation, as well as downregulation of many neuropeptide genes. Analysis of cell cycle and regulatory gene expression revealed upregulation of the E2f1 cell cycle factor, as well as changes in Antennapedia and Grainy head expression. We also found reduction of H3K4me3 modification in the embryonic nervous system. Genome-wide transcriptome analysis points to additional downstream genes that may underlie these Ctr9 phenotypes, revealing gene expression changes in Notch pathway target genes, cell cycle genes, and neuropeptide genes. In addition, we find significant effects on the gene expression of metabolic genes. These findings reveal that Ctr9 is an essential gene that is necessary at multiple stages of nervous system development, and provides a starting point for future studies of the Paf1C in Drosophila.

    Ort, förlag, år, upplaga, sidor
    Genetics Society of America, 2016
    Nyckelord
    neuroblast, lineage tree, cell cycle, epigenetics, terminal differentiation, FlyBook
    Nationell ämneskategori
    Genetik
    Identifikatorer
    urn:nbn:se:liu:diva-132856 (URN)10.1534/g3.116.034231 (DOI)000386581200018 ()27520958 (PubMedID)
    Anmärkning

    Funding Agencies|Swedish Research Council [621-2013-5258]; Knut and Alice Wallenberg Foundation [KAW2011.0165]; Swedish Cancer Foundation [120531]; Swedish Royal Academy of Sciences

    Tillgänglig från: 2016-12-06 Skapad: 2016-11-30 Senast uppdaterad: 2017-11-29
    3. Neural Lineage Progression Controlled by a Temporal Proliferation Program.
    Öppna denna publikation i ny flik eller fönster >>Neural Lineage Progression Controlled by a Temporal Proliferation Program.
    Visa övriga...
    2017 (Engelska)Ingår i: Developmental Cell, ISSN 1534-5807, E-ISSN 1878-1551, Vol. 43, nr 3, s. 332-348Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Great progress has been made in identifying transcriptional programs that establish stem cell identity. In contrast, we have limited insight into how these programs are down-graded in a timely manner to halt proliferation and allow for cellular differentiation. Drosophila embryonic neuroblasts undergo such a temporal progression, initially dividing to bud off daughters that divide once (type I), then switching to generating non-dividing daughters (type 0), and finally exiting the cell cycle. We identify six early transcription factors that drive neuroblast and type I daughter proliferation. Early factors are gradually replaced by three late factors, acting to trigger the type I→0 daughter proliferation switch and eventually to stop neuroblasts. Early and late factors regulate each other and four key cell-cycle genes, providing a logical genetic pathway for these transitions. The identification of this extensive driver-stopper temporal program controlling neuroblast lineage progression may have implications for studies in many other systems.less thanbr /greater than (Copyright © 2017 Elsevier Inc. All rights reserved.)

    Ort, förlag, år, upplaga, sidor
    Cell Press, 2017
    Nationell ämneskategori
    Utvecklingsbiologi
    Identifikatorer
    urn:nbn:se:liu:diva-143117 (URN)10.1016/j.devcel.2017.10.004 (DOI)000414584300011 ()29112852 (PubMedID)
    Anmärkning

    Funding agencies: Swedish Research Council [621-2013-5258]; Knut and Alice Wallenberg Foundation [KAW2011.0165, KAW2012.0101]; Swedish Cancer Foundation [140780, 150633]

    Tillgänglig från: 2017-11-20 Skapad: 2017-11-20 Senast uppdaterad: 2017-11-20Bibliografiskt granskad
  • 7.
    Barczyk, K.
    et al.
    Department of Immunology, Faculty of Biotechnology, Jagiellonian University, Krakow, Poland; Institute of Experimental Dermatology, University of Münster, Münster, Germany.
    Kreuter, M.
    Department of Medicine/Hematology and Oncology, University of Münster, Münster, Germany.
    Pryjma, J.
    Department of Immunology, Faculty of Biotechnology, Jagiellonian University, Krakow, Poland.
    Booy, Evan P.
    Manitoba Institute of Cell Biology, and Department of Biochemistry and Medical Genetics, Univ. Manitoba, Winnipeg, Canada.
    Maddika, Subbareddy
    Manitoba Institute of Cell Biology, Cancer Care Manitoba; Department of Biochemistry and Medical Genetics,University of Manitoba, Winnipeg, Canada .
    Ghavami, Saeid
    Department of Biochemistry and Medical Genetics, Manitoba Institute of Cell Biology, Cancer Care Manitoba, Winnipeg, Manitoba, Canada.
    Berdel, W. E.
    Department of Medicine/Hematology and Oncology, University of Münster, Münster, Germany.
    Roth, J.
    Institute of Experimental Dermatology, University of Münster, Münster, Germany.
    Los, Marek Jan
    Institute of Experimental Dermatology, University of Münster, Münster, Germany Manitoba Institute of Cell Biology, Cancer Care Manitoba; Manitoba Institute of Child Health; Department of Biochemistry and Medical Genetics; Department of Human Anatomy and Cell Science, University Manitoba, Winnipeg, Canada, .
    Serum cytochrome c indicates in vivo apoptosis and can serve as a prognostic marker during cancer therapy2005Ingår i: International Journal of Cancer, ISSN 0020-7136, E-ISSN 1097-0215, Vol. 116, nr 2, s. 167-173Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Despite significant progress in cancer therapy, the outcome of the treatment is often unfavorable. Better treatment monitoring would not only allow an individual more effective, patient-adjusted therapy, but also it would eliminate some of the side effects. Using a cytochrome c ELISA that was modified to increase sensitivity, we demonstrate that serum cytochrome c is a sensitive apoptotic marker in vivo reflecting therapy-induced cell death burden. Furthermore, increased serum cytochrome c level is a negative prognostic marker. Cancer patients whose serum cytochrome c level was normal 3 years ago have a twice as high probability to be still alive, as judged from sera samples collected for years, analyzed recently and matched with survival data. Moreover, we show that serum cytochrome c and serum LDH-activity reflect different stages and different forms of cell death. Cellular cytochrome c release is specific for apoptosis, whereas increased LDH activity is an indicator of (secondary) necrosis. Whereas serum LDH activity reflects the "global" degree of cell death over a period of time, the sensitive cytochrome c-based method allows confirmation of the individual cancer therapy-induced and spontaneous cell death events. The combination of cytochrome c with tissue-specific markers may provide the foundation for precise monitoring of apoptosis in vivo, by "lab-on-the-chip" technology. (c) 2005 Wiley-Liss, Inc.

  • 8.
    Baumgartner, Johanna
    et al.
    Linköpings universitet, Institutionen för fysik, kemi och biologi. Linköpings universitet, Tekniska fakulteten.
    Jönsson, Jan-Ingvar
    Linköpings universitet, Medicinska fakulteten. Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för hematopoes och utvecklingsbiologi.
    Jager, Edwin W. H.
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Sensor- och aktuatorsystem. Linköpings universitet, Tekniska fakulteten.
    Switchable presentation of cytokines on electroactive polypyrrole surfaces for hematopoietic stem and progenitor cells2018Ingår i: Journal of Materials Chemistry B, ISSN 2050-750X, Vol. 6, s. 4665-4675Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Hematopoietic stem cells are used in transplantations for patients with hematologic malignancies. Scarce sources require efficient strategies of expansion, including polymeric biomaterials mimicking architectures of bone marrow tissue. Tissue microenvironment and mode of cytokine presentation strongly influence cell fate. Although several cytokines with different functions as soluble or membrane-bound mediators have already been identified, their precise roles have not yet been clarified. A need exists for in vitro systems that mimic the in vivo situation to enable such studies. One way is to establish surfaces mimicking physiological presentation using protein-immobilization onto polymer films. However these films merely provide a static presentation of the immobilized proteins. It would be advantageous to also dynamically change protein presentation and functionality to better reflect the in vivo conditions. The electroactive polymer polypyrrole shows excellent biocompatibility and electrochemically alters its surface properties, becoming an interesting choice for such setups. Here, we present an in vitro system for switchable presentation of membrane-bound cytokines. We use interleukin IL-3, known to affect hematopoiesis, and show that when immobilized on polypyrrole films, IL-3 is bioavailable for the bone marrow-derived FDC-P1 progenitor cell line. Moreover, IL-3 presentation can be successfully altered by changing the redox state of the film, in turn influencing FDC-P1 cell viability. This novel in vitro system provides a valuable tool for stimuli-responsive switchable protein presentation allowing the dissection of relevant mediators in stem and progenitor cell behavior.

  • 9.
    Belogurov, G A
    et al.
    A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia.
    Fabrichniy, I P
    A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia.
    Pohjanjoki, P
    Department of Biochemistry, University of Turku, Turku, Finland.
    Kasho, V N
    Center for Ulcer Research and Education, Department of Medicine, University of California, Los Angeles, California, USA.
    Lehtihuhta, E
    Department of Biochemistry, University of Turku, Turku, Finland.
    Turkina, Maria V
    A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia.
    Cooperman, B S
    Department of Chemistry, University of Pennsylvania, Pennsylvania, USA.
    Goldman, A
    Institute of Biotechnology, University of Helsinki, Helsinki, Finland.
    Baykov, A A
    A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia.
    Lahti, R
    Department of Biochemistry, University of Turku, Turku, Finland.
    Catalytically important ionizations along the reaction pathway of yeast pyrophosphatase2000Ingår i: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 39, nr 45, s. 13931-13938Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Five catalytic functions of yeast inorganic pyrophosphatase were measured over wide pH ranges: steady-state PP(i) hydrolysis (pH 4. 8-10) and synthesis (6.3-9.3), phosphate-water oxygen exchange (pH 4. 8-9.3), equilibrium formation of enzyme-bound PP(i) (pH 4.8-9.3), and Mg(2+) binding (pH 5.5-9.3). These data confirmed that enzyme-PP(i) intermediate undergoes isomerization in the reaction cycle and allowed estimation of the microscopic rate constant for chemical bond breakage and the macroscopic rate constant for PP(i) release. The isomerization was found to decrease the pK(a) of the essential group in the enzyme-PP(i) intermediate, presumably nucleophilic water, from >7 to 5.85. Protonation of the isomerized enzyme-PP(i) intermediate decelerates PP(i) hydrolysis but accelerates PP(i) release by affecting the back isomerization. The binding of two Mg(2+) ions to free enzyme requires about five basic groups with a mean pK(a) of 6.3. An acidic group with a pK(a) approximately 9 is modulatory in PP(i) hydrolysis and metal ion binding, suggesting that this group maintains overall enzyme structure rather than being directly involved in catalysis.

  • 10.
    Belogurov, Georgiy A
    et al.
    Department of Biochemistry and Food Chemistry, University of Turku, Turku, Finland; A. N. Belozersky Institute of Physico-Chemical Biology and School of Chemistry, Moscow State University, Moscow, Russia.
    Malinen, Anssi M
    Department of Biochemistry and Food Chemistry, University of Turku, Turku, Finland.
    Turkina, Maria V
    A. N. Belozersky Institute of Physico-Chemical Biology and School of Chemistry, Moscow State University, Moscow, Russia.
    Jalonen, Ulla
    Department of Biochemistry and Food Chemistry, University of Turku, Turku, Finland.
    Rytkönen, Kalle
    Department of Biochemistry and Food Chemistry, University of Turku, Turku, Finland.
    Baykov, Alexander A
    A. N. Belozersky Institute of Physico-Chemical Biology and School of Chemistry, Moscow State University, Moscow, Russia.
    Lahti, Reijo
    Department of Biochemistry and Food Chemistry, University of Turku, Turku, Finland.
    Membrane-bound pyrophosphatase of Thermotoga maritima requires sodium for activity2005Ingår i: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 44, nr 6, s. 2088-2096Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Membrane-bound pyrophosphatase of the hyperthermophilic bacterium Thermotoga maritima(Tm-PPase), a homologue of H(+)-translocating pyrophosphatase, was expressed in Escherichia coli and isolated as inner membrane vesicles. In contrast to all previously studied H(+)-PPases, both native and recombinant Tm-PPases exhibited an absolute requirement for Na(+) but displayed the highest activity in the presence of millimolar levels of both Na(+) and K(+). Detergent-solubilized recombinant Tm-PPase was thermostable and retained the monovalent cation requirements of the membrane-embedded enzyme. Steady-state kinetic analysis of pyrophosphate hydrolysis by the wild-type enzyme suggested that two Na(+) binding sites and one K(+) binding site are involved in enzyme activation. The affinity of the site that binds Na(+) first is increased with increasing K(+) concentration. In contrast, only one Na(+) binding site (K(+)-dependent) and one K(+) binding site were involved in activation of the Asp(703) --> Asn variant. Thus, Asp(703) may form part of the K(+)-independent Na(+) binding site. Unlike all other membrane and soluble PPases, Tm-PPase did not catalyze oxygen exchange between phosphate and water. However, solubilized Tm-PPase exhibited low but measurable PP(i)-synthesizing activity, which also required Na(+) but was inhibited by K(+). These results demonstrate that T. maritima PPase belongs to a previously unknown subfamily of Na(+)-dependent H(+)-PPase homologues and may be an analogue of Na(+),K(+)-ATPase.

  • 11.
    Belogurov, Georgiy A
    et al.
    Department of Biochemistry and Food Chemistry, University of Turku, Turku, Finland; A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia.
    Turkina, Maria V
    A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia.
    Penttinen, Anni
    Department of Biochemistry and Food Chemistry, University of Turku, Turku, Finland.
    Huopalahti, Saila
    Department of Biochemistry and Food Chemistry, University of Turku, Turku, Finland.
    Baykov, Alexander A
    A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia.
    Lahti, Reijo
    Department of Biochemistry and Food Chemistry, University of Turku, Turku, Finland.
    H+-pyrophosphatase of Rhodospirillum rubrum. High yield expression in Escherichia coli and identification of the Cys residues responsible for inactivation my mersalyl2002Ingår i: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 277, nr 25Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    H(+)-translocating pyrophosphatase (H(+)-PPase) of the photosynthetic bacterium Rhodospirillum rubrum was expressed in Escherichia coli C43(DE3) cells. Recombinant H(+)-PPase was observed in inner membrane vesicles, where it catalyzed both PP(i) hydrolysis coupled with H(+) transport into the vesicles and PP(i) synthesis. The hydrolytic activity of H(+)-PPase in E. coli vesicles was eight times greater than that in R. rubrum chromatophores but exhibited similar sensitivity to the H(+)-PPase inhibitor, aminomethylenediphosphonate, and insensitivity to the soluble PPase inhibitor, fluoride. Using this expression system, we showed that substitution of Cys(185), Cys(222), or Cys(573) with aliphatic residues had no effect on the activity of H(+)-PPase but decreased its sensitivity to the sulfhydryl modifying reagent, mersalyl. H(+)-PPase lacking all three Cys residues was completely resistant to the effects of mersalyl. Mg(2+) and MgPP(i) protected Cys(185) and Cys(573) from modification by this agent but not Cys(222). Phylogenetic analyses of 23 nonredundant H(+)-PPase sequences led to classification into two subfamilies. One subfamily invariably contains Cys(222) and includes all known K(+)-independent H(+)-PPases, whereas the other incorporates a conserved Cys(573) but lacks Cys(222) and includes all known K(+)-dependent H(+)-PPases. These data suggest a specific link between the incidence of Cys at positions 222 and 573 and the K(+) dependence of H(+)-PPase.

  • 12.
    Bettiga, Arianna
    et al.
    IRCCS Osped San Raffaele, Italy.
    Aureli, Massimo
    University of Milan, Italy.
    Colciago, Giorgia
    IRCCS Osped San Raffaele, Italy.
    Murdica, Valentina
    University of Milan, Italy.
    Moschini, Marco
    IRCCS Osped San Raffaele, Italy.
    Luciano, Roberta
    IRCCS Osped San Raffaele, Italy.
    Canals, Daniel
    SUNY Stony Brook, NY 11794 USA.
    Hannun, Yusuf
    SUNY Stony Brook, NY 11794 USA.
    Hedlund, Petter
    Linköpings universitet, Institutionen för medicin och hälsa, Avdelningen för läkemedelsforskning. Linköpings universitet, Medicinska fakulteten. Region Östergötland, Diagnostikcentrum, Klinisk farmakologi. IRCCS Osped San Raffaele, Italy.
    Lavorgna, Giovanni
    IRCCS Osped San Raffaele, Italy.
    Colombo, Renzo
    IRCCS Osped San Raffaele, Italy.
    Bassi, Rosaria
    University of Milan, Italy.
    Samarani, Maura
    University of Milan, Italy.
    Montorsi, Francesco
    IRCCS Osped San Raffaele, Italy; University of Vita Salute San Raffaele, Italy.
    Salonia, Andrea
    University of Vita Salute San Raffaele, Italy.
    Benigni, Fabio
    IRCCS Osped San Raffaele, Italy.
    Bladder cancer cell growth and motility implicate cannabinoid 2 receptor-mediated modifications of sphingolipids metabolism2017Ingår i: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 7, artikel-id 42157Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The inhibitory effects demonstrated by activation of cannabinoid receptors (CB) on cancer proliferation and migration may also play critical roles in controlling bladder cancer (BC). CB expression on human normal and BC specimens was tested by immunohistochemistry. Human BC cells RT4 and RT112 were challenged with CB agonists and assessed for proliferation, apoptosis, and motility. Cellular sphingolipids (SL) constitution and metabolism were evaluated after metabolic labelling. CB1-2 were detected in BC specimens, but only CB2 was more expressed in the tumour. Both cell lines expressed similar CB2. Exposure to CB2 agonists inhibited BC growth, down-modulated Akt, induced caspase 3-activation and modified SL metabolism. Baseline SL analysis in cell lines showed differences linked to unique migratory behaviours and cytoskeletal re-arrangements. CB2 activation changed the SL composition of more aggressive RT112 cells by reducing (p amp;lt; 0.01) Gb3 ganglioside (-50 +/- 3%) and sphingosine 1-phosphate (S1P, -40 +/- 4%), which ended up to reduction in cell motility (-46 +/- 5%) with inhibition of p-SRC. CB2-selective antagonists, gene silencing and an inhibitor of SL biosynthesis partially prevented CB2 agonist-induced effects on cell viability and motility. CB2 activation led to ceramide-mediated BC cell apoptosis independently of SL constitutive composition, which instead was modulated by CB2 agonists to reduce cell motility.

  • 13.
    Bivik Stadler, Caroline
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för mikrobiologi och molekylär medicin. Linköpings universitet, Medicinska fakulteten.
    Genetic pathways controlling CNS development: The role of Notch signaling in regulating daughter cell proliferation in Drosophila2016Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The human central nervous system (CNS) displays the greatest cellular diversity of any organ system, consisting of billions of neurons, of numerous cell sub-types, interconnected in a vast network. Given this enormous complexity, decoding the genetic programs controlling the multistep process of CNS development remains a major challenge. While great progress has been made with respect to understanding sub-type specification, considerably less is known regarding how the generation of the precise number of each sub-type is controlled.

    The aim of this thesis was to gain deeper knowledge into the regulatory programs controlling cell specification and proliferation. To address these questions I have studied the Drosophila embryonic CNS as a model system, to thereby be able to investigate the genetic mechanisms at high resolution. Despite the different size and morphology between the Drosophila and the mammalian CNS, the lineages of their progenitors share similarity. Importantly for this thesis, both species progenitors show elaborate variations in their proliferation modes, either giving rise to daughters that can directly differentiate into neurons or glia (type 0), divide once (type I), or multiple times (type II).

    The studies launched off with a comprehensive chemical forward genetic screen, for the very last born cell in the well-studied lineage of progenitor NB5-6T: the Ap4 neuron, which expresses the neuropeptide FMRFa. NB5-6T is a powerful model to use, because it undergoes a programmed type I>0 daughter cell proliferation switch. An FMRF-eGFP transgenic reporter was utilized as readout for successful terminal differentiation of Ap4/FMRFa and thereby proper lineage progression of the ∼20 cells generated. The strongest mutants were mapped to genes with both known and novel essential functions e.g., spatial and temporal patterning, cell cycle control, cell specification and chromatin modification. Subsequently, we focused on some of the genes that showed a loss of function phenotype with an excess of lineage cells. We found that Notch is critical for the type I>0 daughter cell proliferation switch in the NB5-6T lineage and globally as well. When addressing the broader relevance of these findings, and to further decipher the Notch pathway, we discovered that selective groups of E(spl) genes is controlling the switch in a close interplay with four key cell cycle factors: Cyclin E, String, E2F and Dacapo, in most if not all embryonic progenitors. The Notch mediation of the switch is likely to be by direct transcriptional regulation. Furthermore, another gene identified in the screen, sequoia, was investigated. The analysis revealed that sequoia is also controlling the daughter cell switch in the CNS, and this partly through context dependent interactions with the Notch pathway.

    Taken together, the findings presented in this thesis demonstrate that daughter cell proliferation switches in Drosophila neural lineages are genetically programmed, and that Notch contributes to the triggering of these events. Given that early embryonic processes is frequently shown to be evolutionary conserved, you can speculate that changeable daughter proliferation programs could be applied to mammals, and contribute to a broader understanding of proliferation processes in humans as well.

     

    Delarbeten
    1. Novel Genes Involved in Controlling Specification of Drosophila FMRFamide Neuropeptide Cells
    Öppna denna publikation i ny flik eller fönster >>Novel Genes Involved in Controlling Specification of Drosophila FMRFamide Neuropeptide Cells
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    2015 (Engelska)Ingår i: Genetics, ISSN 0016-6731, E-ISSN 1943-2631, Vol. 200, nr 4, s. 1229-1244Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The expression of neuropeptides is often extremely restricted in the nervous system, making them powerful markers for addressing cell specification . In the developing Drosophila ventral nerve cord, only six cells, the Ap4 neurons, of some 10,000 neurons, express the neuropeptide FMRFamide (FMRFa). Each Ap4/FMRFa neuron is the last-born cell generated by an identifiable and well-studied progenitor cell, neuroblast 5-6 (NB5-6T). The restricted expression of FMRFa and the wealth of information regarding its gene regulation and Ap4 neuron specification makes FMRFa a valuable readout for addressing many aspects of neural development, i.e., spatial and temporal patterning cues, cell cycle control, cell specification, axon transport, and retrograde signaling. To this end, we have conducted a forward genetic screen utilizing an Ap4-specific FMRFa-eGFP transgenic reporter as our readout. A total of 9781 EMS-mutated chromosomes were screened for perturbations in FMRFa-eGFP expression, and 611 mutants were identified. Seventy-nine of the strongest mutants were mapped down to the affected gene by deficiency mapping or whole-genome sequencing. We isolated novel alleles for previously known FMRFa regulators, confirming the validity of the screen. In addition, we identified novel essential genes, including several with previously undefined functions in neural development. Our identification of genes affecting most major steps required for successful terminal differentiation of Ap4 neurons provides a comprehensive view of the genetic flow controlling the generation of highly unique neuronal cell types in the developing nervous system.

    Ort, förlag, år, upplaga, sidor
    Genetics Society of America, 2015
    Nyckelord
    Drosophila; CNS development; neural cell fate specification; forward genetic screening; FMRFamide
    Nationell ämneskategori
    Klinisk medicin
    Identifikatorer
    urn:nbn:se:liu:diva-121318 (URN)10.1534/genetics.115.178483 (DOI)000359917000020 ()26092715 (PubMedID)
    Tillgänglig från: 2015-09-16 Skapad: 2015-09-14 Senast uppdaterad: 2019-03-13Bibliografiskt granskad
    2. Control of neuronal cell fate and number by integration of distinct daughter cell proliferation modes with temporal progression
    Öppna denna publikation i ny flik eller fönster >>Control of neuronal cell fate and number by integration of distinct daughter cell proliferation modes with temporal progression
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    2012 (Engelska)Ingår i: Development, ISSN 0950-1991, E-ISSN 1477-9129, Vol. 139, nr 4, s. 678-689Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    During neural lineage progression, differences in daughter cell proliferation can generate different lineage topologies. This is apparent in the Drosophila neuroblast 5-6 lineage (NB5-6T), which undergoes a daughter cell proliferation switch from generating daughter cells that divide once to generating neurons directly. Simultaneously, neural lineages, e.g. NB5-6T, undergo temporal changes in competence, as evidenced by the generation of different neural subtypes at distinct time points. When daughter proliferation is altered against a backdrop of temporal competence changes, it may create an integrative mechanism for simultaneously controlling cell fate and number. Here, we identify two independent pathways, Prospero and Notch, which act in concert to control the different daughter cell proliferation modes in NB5-6T. Altering daughter cell proliferation and temporal progression, individually and simultaneously, results in predictable changes in cell fate and number. This demonstrates that different daughter cell proliferation modes can be integrated with temporal competence changes, and suggests a novel mechanism for coordinately controlling neuronal subtype numbers.

    Ort, förlag, år, upplaga, sidor
    Company of Biologists, 2012
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-74790 (URN)10.1242/dev.074500 (DOI)000300259800005 ()
    Anmärkning

    funding agencies|Swedish Research Council||Knut and Alice Wallenberg foundation||Swedish Cancer Foundation||

    Tillgänglig från: 2012-02-08 Skapad: 2012-02-08 Senast uppdaterad: 2019-03-13
    3. Control of Neural Daughter Cell Proliferation by Multi-level Notch/Su(H)/E(spl)-HLH Signaling
    Öppna denna publikation i ny flik eller fönster >>Control of Neural Daughter Cell Proliferation by Multi-level Notch/Su(H)/E(spl)-HLH Signaling
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    2016 (Engelska)Ingår i: PLoS Genetics, ISSN 1553-7390, E-ISSN 1553-7404, Vol. 12, nr 4, artikel-id e1005984Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The Notch pathway controls proliferation during development and in adulthood, and is frequently affected in many disorders. However, the genetic sensitivity and multi-layered transcriptional properties of the Notch pathway has made its molecular decoding challenging. Here, we address the complexity of Notch signaling with respect to proliferation, using the developing Drosophila CNS as model. We find that a Notch/Su(H)/E(spl)-HLH cascade specifically controls daughter, but not progenitor proliferation. Additionally, we find that different E(spl)-HLH genes are required in different neuroblast lineages. The Notch/Su(H)/E(spl)-HLH cascade alters daughter proliferation by regulating four key cell cycle factors: Cyclin E, String/Cdc25, E2f and Dacapo (mammalian p21(CIP1)/p27(KIP1)/p57(Kip2)). ChIP and DamID analysis of Su(H) and E(spl)-HLH indicates direct transcriptional regulation of the cell cycle genes, and of the Notch pathway itself. These results point to a multi-level signaling model and may help shed light on the dichotomous proliferative role of Notch signaling in many other systems.

    Ort, förlag, år, upplaga, sidor
    PUBLIC LIBRARY SCIENCE, 2016
    Nationell ämneskategori
    Klinisk medicin
    Identifikatorer
    urn:nbn:se:liu:diva-128759 (URN)10.1371/journal.pgen.1005984 (DOI)000375231900032 ()27070787 (PubMedID)
    Anmärkning

    Funding Agencies|Knut and Alice Wallenberg Foundation [KAW2012.0101]; Swedish Research Council [621-2010-5214]; Swedish Cancer Foundation [120531]

    Tillgänglig från: 2016-05-31 Skapad: 2016-05-30 Senast uppdaterad: 2019-03-13
    4. sequoia controls the type I>0 daughter proliferation switch in the developing Drosophila nervous system
    Öppna denna publikation i ny flik eller fönster >>sequoia controls the type I>0 daughter proliferation switch in the developing Drosophila nervous system
    2016 (Engelska)Ingår i: Development, ISSN 0950-1991, E-ISSN 1477-9129, Vol. 143, nr 20, s. 3774-3784Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Neural progenitors typically divide asymmetrically to renew themselves, while producing daughters with more limited potential. In the Drosophila embryonic ventral nerve cord, neuroblasts initially produce daughters that divide once to generate two neurons/glia (type I proliferation mode). Subsequently, many neuroblasts switch to generating daughters that differentiate directly (type 0). This programmed type I&gt;0 switch is controlled by Notch signaling, triggered at a distinct point of lineage progression in each neuroblast. However, how Notch signaling onset is gated was unclear. We recently identified Sequoia (Seq), a C2H2 zinc-finger transcription factor with homology to Drosophila Tramtrack (Ttk) and the positive regulatory domain (PRDM) family, as important for lineage progression. Here, we find that seq mutants fail to execute the type I&gt;0 daughter proliferation switch and also display increased neuroblast proliferation. Genetic interaction studies reveal that seq interacts with the Notch pathway, and seq furthermore affects expression of a Notch pathway reporter. These findings suggest that seq may act as a context-dependent regulator of Notch signaling, and underscore the growing connection between Seq, Ttk, the PRDM family and Notch signaling.

    Ort, förlag, år, upplaga, sidor
    The Company of Biologists Ltd, 2016
    Nyckelord
    Lineage tree, Cell cycle, Asymmetric division, Combinatorial control, Notch
    Nationell ämneskategori
    Cell- och molekylärbiologi Biokemi och molekylärbiologi Cellbiologi Medicinsk bioteknologi
    Identifikatorer
    urn:nbn:se:liu:diva-132739 (URN)10.1242/dev.139998 (DOI)000393452500013 ()27578794 (PubMedID)
    Anmärkning

    Funding agencies: Swedish Research Council (Vetenskapsradet); Knut and Alice Wallenberg Foundation (Knut och Alice Wallenbergs Stiftelse); Swedish Cancer Foundation (Cancerfonden)

    Tillgänglig från: 2016-11-22 Skapad: 2016-11-22 Senast uppdaterad: 2019-03-13Bibliografiskt granskad
  • 14.
    Bivik Stadler, Caroline
    et al.
    Linköpings universitet, Institutionen för medicin och hälsa, Avdelningen för radiologiska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Arefin, Md Badrul
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för hematopoes och utvecklingsbiologi. Linköpings universitet, Medicinska fakulteten.
    Ekman, Helen
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för mikrobiologi, infektion och inflammation. Linköpings universitet, Medicinska fakulteten.
    Thor, Stefan
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för hematopoes och utvecklingsbiologi. Linköpings universitet, Medicinska fakulteten. Univ Queensland, Australia.
    PIP degron-stabilized Dacapo/p21(Cip)(1) and mutations in ago act in an anti- versus pro-proliferative manner, yet both trigger an increase in Cyclin E levels2019Ingår i: Development, ISSN 0950-1991, E-ISSN 1477-9129, Vol. 146, nr 13, artikel-id UNSP dev175927Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    During cell cycle progression, the activity of the CycE-Cdk2 complex gates S-phase entry. CycE-Cdk2 is inhibited by CDK inhibitors (CKIs) of the Cip/Kip family, which include the human p21(Cip)(1) and Drosophila Dacapo (Dap) proteins. Both the CycE and Cip/Kip family proteins are under elaborate control via protein degradation, mediated by the Cullin-RING ligase (CRL) family of ubiquitin ligase complexes. The CRL complex SCFFoxw7/Ago targets phosphorylated CycE, whereas p21(Cip)(1) and Dap are targeted by the CRLCdf2 complex, binding to the PIP degron. The role of CRL-mediated degradation of CycE and Cip/Kip proteins during CNS development is not well understood. Here, we analyse the role of ago (Fbxw7)-mediated CycE degradation, and of Dap and p21(Cip)(1) degradation during Drosophila CNS development. We find that ago mutants display over-proliferation, accompanied by elevated CycE expression levels. By contrast, expression of PIP degron mutant Dap and p21(Cip)(1) transgenes inhibit proliferation. However, surprisingly, this is also accompanied by elevated CycE levels. Hence, ago mutation and PIP degron Cip/Kip transgenic expression trigger opposite effects on proliferation, but similar effects on CycE levels.

  • 15.
    Björk Wilhelms, Daniel
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Fever: Role of brain endothelial prostaglandins2014Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Fever and loss of appetite are two of the most fundamental manifestations of disease. These disease symptoms, which lead to deviations from normal body temperature and food intake patterns, are seen in a vast array of infectious and inflammatory conditions. It is known that peripheral signals from the immune system are essential triggers for these responses, which are orchestrated by neuronal circuits in the brain. Due to the blood‐brain barrier, peripheral inflammatory signals require a specific mode of transmission into the brain. Such mechanisms have been proposed, but interventional studies of these mechanisms have never rendered conclusive results. In this thesis, we present the first functional evidence of cyclooxygenase 2 (COX‐2) and microsomal prostaglandin E synthase type 1 (mPGES‐1) mediated prostaglandin E2 synthesis in the blood‐brain barrier endothelial cells as a signaling mechanism in the initiation of inflammatory fever. We also show that one of the world’s most widely used antipyretics, paracetamol, acts by inhibition of COX‐2. Combined with the finding that COX‐2 and mPGES‐1 in brain endothelial cells play a key role in inflammatory fever, this finding suggests that paracetamol inhibits fever by specifically blocking prostaglandin E2 synthesis in blood‐brain barrier endothelium. In another symptom of inflammation, anorexia, the cellular origin of peripheral signals triggering acute anorexia are largely unknown. We show that the expression of myeloid differentiation primary response gene 88 (Myd88) in myeloid cells is important for the initiation of acute inflammatory anorexia and the maintenance of cancer anorexia‐cachexia.

    Taken together, these findings provide a significant advancement of our understanding of the mechanisms triggering acute inflammatory fever and anorexia and also explain the antipyretic effect of paracetamol.

    Delarbeten
    1. Inflammation- and tumor-induced anorexia and weight loss require MyD88 in hematopoietic/myeloid cells but not in brain endothelial or neural cells
    Öppna denna publikation i ny flik eller fönster >>Inflammation- and tumor-induced anorexia and weight loss require MyD88 in hematopoietic/myeloid cells but not in brain endothelial or neural cells
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    2013 (Engelska)Ingår i: The FASEB Journal, ISSN 0892-6638, E-ISSN 1530-6860, Vol. 27, nr 5, s. 1973-1980Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Loss of appetite is a hallmark of inflammatory diseases. The underlying mechanisms remain undefined, but it is known that myeloid differentiation primary response gene 88 (MyD88), an adaptor protein critical for Toll-like and IL-1 receptor family signaling, is involved. Here we addressed the question of determining in which cells the MyD88 signaling that results in anorexia development occurs by using chimeric mice and animals with cell-specific deletions. We found that MyD88-knockout mice, which are resistant to bacterial lipopolysaccharide (LPS)-induced anorexia, displayed anorexia when transplanted with wild-type bone marrow cells. Furthermore, mice with a targeted deletion of MyD88 in hematopoietic or myeloid cells were largely protected against LPS-induced anorexia and displayed attenuated weight loss, whereas mice with MyD88 deletion in hepatocytes or in neural cells or the cerebrovascular endothelium developed anorexia and weight loss of similar magnitude as wild-type mice. Furthermore, in a model for cancer-induced anorexia-cachexia, deletion of MyD88 in hematopoietic cells attenuated the anorexia and protected against body weight loss. These findings demonstrate that MyD88-dependent signaling within the brain is not required for eliciting inflammation-induced anorexia. Instead, we identify MyD88 signaling in hematopoietic/myeloid cells as a critical component for acute inflammatory-driven anorexia, as well as for chronic anorexia and weight loss associated with malignant disease.

    Ort, förlag, år, upplaga, sidor
    Federation of American Society of Experimental Biology (FASEB), 2013
    Nyckelord
    lipopolysaccharide; methylcholanthrene-induced sarcoma; food intake; chimeric mice; Cre-LoxP; inducible cell-specific deletion
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-96147 (URN)10.1096/fj.12-225433 (DOI)000318226100017 ()
    Tillgänglig från: 2013-08-14 Skapad: 2013-08-14 Senast uppdaterad: 2017-12-06
    2. Acetaminophen reduces lipopolysaccharide-induced fever by inhibiting cyclooxygenase-2
    Öppna denna publikation i ny flik eller fönster >>Acetaminophen reduces lipopolysaccharide-induced fever by inhibiting cyclooxygenase-2
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    2013 (Engelska)Ingår i: Neuropharmacology, ISSN 0028-3908, E-ISSN 1873-7064, Vol. 71, s. 124-129Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Acetaminophen is one of the world's most commonly used drugs to treat fever and pain, yet its mechanism of action has remained unclear. Here we tested the hypothesis that acetaminophen blocks fever through inhibition of cyclooxygenase-2 (Cox-2), by monitoring lipopolysaccharide induced fever in mice with genetic manipulations of enzymes in the prostaglandin cascade. We exploited the fact that lowered levels of a specific enzyme make the system more sensitive to any further inhibition of the same enzyme. Mice were immune challenged by an intraperitoneal injection of bacterial wall lipopolysaccharide and their body temperature recorded by telemetry. We found that mice heterozygous for Cox-2, but not for microsomal prostaglandin E synthase-1 (mPGES-1), displayed attenuated fever, indicating a rate limiting role of Cox-2. We then titrated a dose of acetaminophen that did not inhibit the lipopolysaccharide-induced fever in wild-type mice. However, when the same dose of acetaminophen was given to Cox-2 heterozygous mice, the febrile response to lipopolysaccharide was strongly attenuated, resulting in an almost normalized temperature curve, whereas no difference was seen between wild-type and heterozygous mPGES-1 mice. Furthermore, the fever to intracerebrally injected prostaglandin E2 was unaffected by acetaminophen treatment. These findings reveal that acetaminophen, similar to aspirin and other non-steroidal anti-inflammatory drugs, is antipyretic by inhibiting cyclooxygenase-2, and not by inhibiting mPGES-1 or signaling cascades downstream of prostaglandin E2.

    Ort, förlag, år, upplaga, sidor
    Elsevier, 2013
    Nyckelord
    Fever; Cyclooxygenase-2; Cyclooxygenase-1; Microsomal prostaglandin E synthase-1; Gene dosage; Hypothalamus
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-96170 (URN)10.1016/j.neuropharm.2013.03.012 (DOI)000320424200012 ()
    Tillgänglig från: 2013-08-14 Skapad: 2013-08-14 Senast uppdaterad: 2017-12-06
    3. Deletion of Prostaglandin E-2 Synthesizing Enzymes in Brain Endothelial Cells Attenuates Inflammatory Fever
    Öppna denna publikation i ny flik eller fönster >>Deletion of Prostaglandin E-2 Synthesizing Enzymes in Brain Endothelial Cells Attenuates Inflammatory Fever
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    2014 (Engelska)Ingår i: Journal of Neuroscience, ISSN 0270-6474, E-ISSN 1529-2401, Vol. 34, nr 35, s. 11684-11690Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Fever is a hallmark of inflammatory and infectious diseases. The febrile response is triggered by prostaglandin E-2 synthesis mediated by induced expression of the enzymes cyclooxygenase-2 (COX-2) and microsomal prostaglandin E synthase 1 (mPGES-1). The cellular source for pyrogenic PGE(2) remains a subject of debate; several hypotheses have been forwarded, including immune cells in the periphery and in the brain, as well as the brain endothelium. Here we generated mice with selective deletion of COX-2 and mPGES1 in brain endothelial cells. These mice displayed strongly attenuated febrile responses to peripheral immune challenge. In contrast, inflammation-induced hypoactivity was unaffected, demonstrating the physiological selectivity of the response to the targeted gene deletions. These findings demonstrate that PGE(2) synthesis in brain endothelial cells is critical for inflammation-induced fever.

    Ort, förlag, år, upplaga, sidor
    Society for Neuroscience, 2014
    Nyckelord
    COX-2; endothelium; fever; mPGES-1; PGE(2); prostaglandin
    Nationell ämneskategori
    Cell- och molekylärbiologi Neurovetenskaper
    Identifikatorer
    urn:nbn:se:liu:diva-111281 (URN)10.1523/JNEUROSCI.1838-14.2014 (DOI)000341314900017 ()25164664 (PubMedID)
    Anmärkning

    Funding Agencies|Swedish Medical Research Council; Swedish Cancer Foundation; European Research Council; Knut and Alice Wallenberg Foundation; Swedish Brain foundation; County Council of stergotland; Wenner-Gren Fellowship

    Tillgänglig från: 2014-10-14 Skapad: 2014-10-14 Senast uppdaterad: 2018-01-11
    4. Cyclooxygenase isoform exchange blocks inflammatory symptoms
    Öppna denna publikation i ny flik eller fönster >>Cyclooxygenase isoform exchange blocks inflammatory symptoms
    2014 (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    Cyclooxygenase‐2 (COX‐2) is the main source of inducible prostaglandin E2 production and mediates inflammatory symptoms including fever, loss of appetite and hyperalgesia. In contrast, COX‐1 is dispensable for most inflammatory symptoms. Global deletion of COX‐2 leads to a blockade of inflammation‐induced fever and appetite loss but also to high rates of fetal mortality. The latter is unfortunate since mice without COX‐2 are powerful tools in the study of inflammation and cardiovascular medicine. The differential functionality of the COX isoforms could be due to differences in regulatory regions of the genes, leading to different expression patterns, or to differences in the coding sequence, leading to distinct functional properties of the proteins. To study this in the context of inflammatory symptoms, we used mice in which the coding sequence of COX‐2 was replaced by the corresponding sequence of COX‐1. In these mice, COX‐1 mRNA was induced by inflammation but COX‐1 protein expression did not fully mimic inflammation‐induced COX‐2 expression. Just like mice globally lacking COX‐2, these mice showed a complete lack of fever and inflammation‐induced anorexia. However, as previously reported, they displayed close to normal survival rates. This shows that the COX activity generated from the hybrid gene was strong enough to allow survival but not strong enough to mediate inflammatory symptoms, making the line an interesting alternative to COX‐2 knockouts for the study of inflammation. Our results also show that the functional differences between COX‐1 and COX‐2 in the context of inflammatory symptoms is not only dependent on the features of the promoter regions. Instead they indicate that there are fundamental differences between the isoforms at translational or posttranslational levels, which make hybrid genes less functional.

    Nationell ämneskategori
    Cellbiologi Reumatologi och inflammation
    Identifikatorer
    urn:nbn:se:liu:diva-111725 (URN)
    Tillgänglig från: 2014-10-29 Skapad: 2014-10-29 Senast uppdaterad: 2015-11-06Bibliografiskt granskad
  • 16.
    Björk Wilhelms, Daniel
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Mirrasekhian, Elahe
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Engblom, David
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Cyclooxygenase isoform exchange blocks inflammatory symptoms2014Manuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    Cyclooxygenase‐2 (COX‐2) is the main source of inducible prostaglandin E2 production and mediates inflammatory symptoms including fever, loss of appetite and hyperalgesia. In contrast, COX‐1 is dispensable for most inflammatory symptoms. Global deletion of COX‐2 leads to a blockade of inflammation‐induced fever and appetite loss but also to high rates of fetal mortality. The latter is unfortunate since mice without COX‐2 are powerful tools in the study of inflammation and cardiovascular medicine. The differential functionality of the COX isoforms could be due to differences in regulatory regions of the genes, leading to different expression patterns, or to differences in the coding sequence, leading to distinct functional properties of the proteins. To study this in the context of inflammatory symptoms, we used mice in which the coding sequence of COX‐2 was replaced by the corresponding sequence of COX‐1. In these mice, COX‐1 mRNA was induced by inflammation but COX‐1 protein expression did not fully mimic inflammation‐induced COX‐2 expression. Just like mice globally lacking COX‐2, these mice showed a complete lack of fever and inflammation‐induced anorexia. However, as previously reported, they displayed close to normal survival rates. This shows that the COX activity generated from the hybrid gene was strong enough to allow survival but not strong enough to mediate inflammatory symptoms, making the line an interesting alternative to COX‐2 knockouts for the study of inflammation. Our results also show that the functional differences between COX‐1 and COX‐2 in the context of inflammatory symptoms is not only dependent on the features of the promoter regions. Instead they indicate that there are fundamental differences between the isoforms at translational or posttranslational levels, which make hybrid genes less functional.

  • 17.
    Bolshakova, A.V.
    et al.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Petukhova, O.A.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Pinaev, G.P.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Magnusson, Karl-Erik
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Medicinsk mikrobiologi. Linköpings universitet, Hälsouniversitetet.
    The comparative analysis of subcellular fractionation methods for revealing of a-actinin 1 and a-actinin 4 in A431 cells2009Ingår i: Tsitologiya, ISSN 0041-3771, Vol. 51, nr 2, s. 122-129Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    a-Actinin 1 and a-actinin 4 belong to a family of actin-binding proteins with shared structural function and regulation of several processes in a cell. Based on previous data on different distribution of these proteins in the nucleus and cytoplasm, we have explored in detail the distribution of a-actinin 1 and a-actinin 4 in subcellular fractions in A431 cells spread on fibronectin. Several methods of subcellular fractionation were used. Complex approach allowed resuming that revealing of a-actinin isoforms in fractions depended on the composition of lysis buffer and preliminary low-temperature freezing of the cells. We have drawn a conclusion that a-actinin 4 can be found in all cytoplasmic and nuclear subfractions, while a-actinin 1 is characterized by cytoplasmic and membrane localization with specificity of its distribution tightly to the nuclear membrane.

  • 18.
    Bosagna, Carlos Guerrero
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biologi. Linköpings universitet, Tekniska fakulteten.
    Developmental and Epigenetic Origins of Male Reproductive Pathologies2015Ingår i: The Epigenome and Developmental Origins of Health and Disease / [ed] Cheryl Rosenfeld, Elsevier, 2015, 1, s. 171-189Kapitel i bok, del av antologi (Refereegranskat)
    Abstract [en]

    Transgenerational epigenetic inheritance has gained increased attention due to the possibility that exposure to environmental toxicants or other stressors can induce long-lasting changes in lineages of organisms. The mechanism involves exposure of pregnant females and induction of germline epigenetic alterations in their developing embryos. This early developmental exposure generates phenotypic alterations in the adults. The germline epigenomic changes produced are then transmitted to future generations and associate with disease phenotypes in the unexposed individuals of subsequent generations. Exposures to environmental toxicants such as fungicides, pesticides, or plastic compounds have been shown in rodents to produce abnormal reproductive or metabolic phenotypes that are transgenerationally transmitted. These include transgenerational increases in the incidence of obesity, polycystic ovary syndrome (PCOS)-like symptoms, pregnancy defects, or germ cell apoptosis. Importantly, the increased incidence of these transgenerationally transmitted diseases in response to environmental exposures in animal models is sometimes drastic. The current evidence on transgenerational epigenetic inheritance observed in animal models allows predicting that environmental exposures of today's inhabitants of the world may affect the incidence of noninfectious diseases in future generations, which would be correlated with long-lasting alterations in the epigenome. The present chapter summarizes the evidence to date for transgenerational epigenetic inheritance, in both humans and animal models.

  • 19.
    Brodin Patcha, Veronika
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Medicinsk mikrobiologi. Linköpings universitet, Hälsouniversitetet.
    Pro- and anti-inflammatory regulation of β2 integrin signalling in human neutrophils2007Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The body is under constant attack from pathogens trying to slip by our immune defence. If the barrier is breached, invading pathogens enter the tissues and cause inflammation. During this process neutrophils, constituting the first line of defence, leave the bloodstream and seek out and kill the invading pathogens. The mechanisms leading to activation of receptors on neutrophils must be closely orchestrated. Pro- and anti-inflammatory substances can influence the outcome of the inflammation process by affecting the involved players. If not well balanced, inflammatory diseases, such as atherosclerosis and rheumatoid arthritis, can be the outcome.

    The aim of this thesis was to elucidate the effect of pro- (fMLP, Leukotriene B4, and Interleukin-8) and anti- (lipoxins, aspirin and statins) inflammatory substances on the β2 integrins, mediating adhesion of neutrophils both under “normal” conditions and during coronary artery disease. More specifically, the effect of these substances on the β2 integrins were studied in regard to: i) the activity (i.e. affinity and avidity) of β2 integrins, ii) the signalling capacity of β2 integrins (i.e. detected as release of arachidonic acid, and the production of reactive oxygen species, and iii) the signal transduction mediated by the β2 integrins (i.e. phosphorylation of Pyk2).

    The pro-inflammatory substances belong to the family of chemoattractants that induces transmigration and chemotaxis. A hierarchy exists between the different family members; the end-target chemoattractants (e.g. fMLP) being more potent than intermediary chemoattractants (e.g. IL-8 and LTB4). It was found that intermediary chemoattractants regulate β2 integrins by mainly affecting the avidity of β2 integrins. End-target chemoattractants on the other hand, affected the β2 integrins by increasing the avidity and the affinity, as well as their signalling capacity.

    The anti-inflammatory substances used in this study were the exogenous aspirin and statins, and the endogenous lipoxins. In the presence of aspirin, stable analogues of lipoxin (i.e. epi-lipoxins) are formed in a trans-cellular process. Lipoxin inhibited the signalling capacity of β2 integrins mediated by intermediary chemoattractants, as well as the signal transduction induced by end-target chemoattractants. Moreover, the signalling capacity of β2 integrins in neutrophils from patients suffering from coronary artery disease (CAD) was impaired. Arachidonic acid, the precursor for both pro- and anti-inflammatory eicosanoid, induced an increase in the β2 integrin activity (both affinity and avidity), but had no effect on the signal transduction.

    In conclusion, different “roles” were observed for end-target and intermediary chemoattractants in the regulation of β2 integrins. The inhibitory effects of the anti-inflammatory lipoxins support earlier studies suggesting that these agents function as “stop signals” in inflammation. This is also confirmed by our findings in CAD patients, who have elevated levels of epi-lipoxins due to aspirin treatment. Moreover, Pyk2 was identified as a possible target for the inhibitory effect of anti-inflammatory drugs.

    Delarbeten
    1. Differential inside-out activation of β2 integrins by leukotriene B4 and fMLP in human neutrophils
    Öppna denna publikation i ny flik eller fönster >>Differential inside-out activation of β2 integrins by leukotriene B4 and fMLP in human neutrophils
    Visa övriga...
    2004 (Engelska)Ingår i: Experimental Cell Research, ISSN 0014-4827, E-ISSN 1090-2422, Vol. 300, nr 2, s. 308-319Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    We have investigated how LTB4, an endogenous chemoattractant encountered early in the inflammatory process, and fMLP, a bacteria-derived chemotactic peptide emanating from the site of infection, mediate inside-out regulation of the β2-integrin. The role of the two chemoattractants on β2-integrin avidity was investigated by measuring their effect on β2-integrin clustering and surface mobility, whereas their effect on β2-integrin affinity was measured by the expression of a high affinity epitope, a ligand-binding domain on β2-integrins, and by integrin binding to s-ICAM. We find that the two chemoattractants modulate the β2-integrin differently. LTB4 induces an increase in integrin clustering and surface mobility, but only a modest increase in integrin affinity. fMLP evokes a large increase in β2-integrin affinity as well as in clustering and mobility. Lipoxin, which acts as a stop signal for the functions mediated by pro-inflammatory agents, was used as a tool for further examining the inside-out mechanisms. While LTB4-induced integrin clustering and mobility were inhibited by lipoxin, only a minor inhibition of fMLP-induced β2-integrin avidity and no inhibition of integrin affinity were detected. The different modes of the inside-out regulation of β2-integrins suggest that distinct mechanisms are involved in the β2-integrin modulation induced by various chemoattractants.

    Nyckelord
    β2-Integrins, Cell adhesion, Chemotactic factors, Eicosanoids, Inflammation, Leukotriene B4, Lipoxins, Human Neutrophils, Signal transduction
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-14629 (URN)10.1016/j.yexcr.2004.07.015 (DOI)
    Tillgänglig från: 2007-09-13 Skapad: 2007-09-13 Senast uppdaterad: 2017-12-13Bibliografiskt granskad
    2. LIpoxin A4 inhibits the fMet-Leu-Phe-induced, but not the β2 integrin-induced activation of the non-receptor tyrosine kinase Pyk2 in Human Leukemia 60 cells
    Öppna denna publikation i ny flik eller fönster >>LIpoxin A4 inhibits the fMet-Leu-Phe-induced, but not the β2 integrin-induced activation of the non-receptor tyrosine kinase Pyk2 in Human Leukemia 60 cells
    Manuskript (Övrigt vetenskapligt)
    Identifikatorer
    urn:nbn:se:liu:diva-14630 (URN)
    Tillgänglig från: 2007-09-13 Skapad: 2007-09-13 Senast uppdaterad: 2010-01-13
    3. Inside-out regulated β2-integrin-induced release of arachidonic acid in Human Leukemia 60 cells
    Öppna denna publikation i ny flik eller fönster >>Inside-out regulated β2-integrin-induced release of arachidonic acid in Human Leukemia 60 cells
    Manuskript (Övrigt vetenskapligt)
    Identifikatorer
    urn:nbn:se:liu:diva-14631 (URN)
    Tillgänglig från: 2007-09-13 Skapad: 2007-09-13 Senast uppdaterad: 2010-01-13
    4. Inactivation of Cdc42 is nessecary for depolymerization of phagosomal F-actin and subsequent phagosomal maturation
    Öppna denna publikation i ny flik eller fönster >>Inactivation of Cdc42 is nessecary for depolymerization of phagosomal F-actin and subsequent phagosomal maturation
    Visa övriga...
    2007 (Engelska)Ingår i: Journal of Immunology, ISSN 0022-1767, Vol. 178, nr 11, s. 7357-7365Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Phagocytosis is a complex process involving the activation of various signaling pathways, such as the Rho GTPases, and the subsequent reorganization of the actin cytoskeleton. In neutrophils, Rac and Cdc42 are activated during phagocytosis but less is known about the involvement of these GTPases during the different stages of the phagocytic process. The aim of this study was to elucidate the role of Cdc42 in phagocytosis and the subsequent phagosomal maturation. Using a TAT-based protein transduction technique, we introduced dominant negative and constitutively active forms of Cdc42 into neutrophil-like HL60 (human leukemia) cells that were allowed to phagocytose IgG-opsonized yeast particles. Staining of cellular F-actin in cells transduced with constitutively active Cdc42 revealed that the activation of Cdc42 induced sustained accumulation of periphagosomal actin. Moreover, the fusion of azurophilic granules with the phagosomal membrane was prevented by the accumulated F-actin. In contrast, introducing dominant negative Cdc42 impaired the translocation per se of azurophilic granules to the periphagosomal area. These results show that efficient phagosomal maturation and the subsequent eradication of ingested microbes in human neutrophils is dependent on a strictly regulated Cdc42. To induce granule translocation, Cdc42 must be in its active state but has to be inactivated to allow depolymerization of the F-actin cage around the phagosome, a process essential for phagolysosome formation.

    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-14632 (URN)
    Tillgänglig från: 2007-09-13 Skapad: 2007-09-13
    5. Neutrophil activation status in stable coronary artery disease.
    Öppna denna publikation i ny flik eller fönster >>Neutrophil activation status in stable coronary artery disease.
    Visa övriga...
    2007 (Engelska)Ingår i: PLoS ONE, ISSN 1932-6203, Vol. 2, nr 10, s. e1056-Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Background: During the last years, neutrophils have emerged as important players in atherogenesis. They are highly activated in peripheral blood of patients with unstable angina. Moreover, a primed state of circulating neutrophils has been proposed in patients with stable angina. Our aim was to investigate the neutrophil activation status in patients with stable coronary artery disease (CAD) at conventional drug treatment.

    Methodology and principal findings: Thirty patients with stable CAD and 30 healthy controls were included using a paired design. The neutrophil expression of CD18 and high-affinity state of CD11b was analysed by flow cytometry before and after stimulation with chemoattractants. Also, the production of reactive oxygen species (ROS) was determined by chemiluminescence. During basal conditions, the neutrophil expression of CD18 or high-affinity state of CD11b did not differ between patients and controls. Chemoattractants (Interleukin-8 and Leukotriene B(4)) did not increase either the expression or the amount of high-affinity CD11b/CD18-integrins in CAD patients compared to controls, and had no effect on the production of ROS. On the other hand, the ROS production in response to C3bi-opsonised yeast particles and the neutrophils' inherent capacity to produce ROS were both significantly decreased in patients.

    Conclusion/Significance: We could not find any evidence that neutrophils in patients with stable CAD were primed, i.e. more prone to activation, compared to cells from healthy controls. According to our data, the circulating neutrophils in CAD patients rather showed an impaired activation status. It remains to be elucidated whether the neutrophil dysfunction in CAD is mainly a marker of chronic disease, an atherogenic factor or a consequence of the drug treatment.

    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-17246 (URN)10.1371/journal.pone.0001056 (DOI)17957240 (PubMedID)
    Tillgänglig från: 2009-03-12 Skapad: 2009-03-12 Senast uppdaterad: 2010-01-14
  • 20.
    Cantù, Claudio
    et al.
    Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy.
    Grande, Vito
    Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy.
    Alborelli, Ilaria
    Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy.
    Cassinelli, Letizia
    Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy.
    Cantù, Ileana
    Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy.
    Colzani, Maria Teresa
    Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy.
    Ierardi, Rossella
    San Raffaele Telethon Institute for Gene Therapy (HSR-TIGET), San Raffaele Scientific Institute, 20132 Milan, Italy.
    Ronzoni, Luisa
    Dipartimento di Medicina Interna, Università di Milano, Fondazione Policlinico Mangiagalli, Regina Elena, IRCCS, Milano, Italy.
    Cappellini, Maria Domenica
    Dipartimento di Medicina Interna, Università di Milano, Fondazione Policlinico Mangiagalli, Regina Elena, IRCCS, Milano, Italy.
    Ferrari, Giuliana
    San Raffaele Telethon Institute for Gene Therapy (HSR-TIGET), San Raffaele Scientific Institute, 20132 Milan // Vita-Salute San Raffaele University, Milan, Italy.
    Ottolenghi, Sergio
    Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy.
    Ronchi, Antonella
    Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy.
    A highly conserved SOX6 double binding site mediates SOX6 gene downregulation in erythroid cells2011Ingår i: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 39, nr 2, s. 486-501Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The Sox6 transcription factor plays critical roles in various cell types, including erythroid cells. Sox6-deficient mice are anemic due to impaired red cell maturation and show inappropriate globin gene expression in definitive erythrocytes. To identify new Sox6 target genes in erythroid cells, we used the known repressive double Sox6 consensus within the εy-globin promoter to perform a bioinformatic genome-wide search for similar, evolutionarily conserved motifs located within genes whose expression changes during erythropoiesis. We found a highly conserved Sox6 consensus within the Sox6 human gene promoter itself. This sequence is bound by Sox6 in vitro and in vivo, and mediates transcriptional repression in transient transfections in human erythroleukemic K562 cells and in primary erythroblasts. The binding of a lentiviral transduced Sox6FLAG protein to the endogenous Sox6 promoter is accompanied, in erythroid cells, by strong downregulation of the endogenous Sox6 transcript and by decreased in vivo chromatin accessibility of this region to the PstI restriction enzyme. These observations suggest that the negative Sox6 autoregulation, mediated by the double Sox6 binding site within its own promoter, may be relevant to control the Sox6 transcriptional downregulation that we observe in human erythroid cultures and in mouse bone marrow cells in late erythroid maturation.

  • 21.
    Chaabane, Wiem
    et al.
    Linköpings universitet, Medicinska fakulteten. Linköpings universitet, Institutionen för medicin och hälsa, Avdelningen för läkemedelsforskning.
    Lindqvist Appell, Malin
    Linköpings universitet, Institutionen för medicin och hälsa, Avdelningen för läkemedelsforskning. Linköpings universitet, Medicinska fakulteten.
    Interconnections between apoptotic and autophagic pathways during thiopurine-induced toxicity in cancer cells: the role of reactive oxygen species2016Ingår i: OncoTarget, ISSN 1949-2553, E-ISSN 1949-2553, Vol. 7, nr 46, s. 75616-75634Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Thiopurines (azathioprine, 6-mercaptopurine and 6-thioguanine) are a class of genotoxic drugs extensively used in the treatment of various illnesses including leukemia. Their underlying molecular mechanism of action involves the activation of apoptosis and autophagy but remains widely unclear. Here we present evidence that autophagy induction by thiopurines is a survival mechanism that antagonizes apoptosis and is involved in degrading damaged mitochondria through mitophagy. On the other hand, apoptosis is the main cell death mechanism by thiopurines as its inhibition prohibited cell death. Thus a tight interplay between apoptosis and autophagy controls cell fate in response to thiopurine treatment. Moreover, thiopurines disrupt mitochondrial function and induce a loss of the mitochondrial transmembrane potential. The involvement of the mitochondrial pathway in thiopurine-induced apoptosis was further confirmed by increased formation of reactive oxygen species (ROS). Inhibiting oxidative stress protected the cells from thiopurine-induced cell death and ROS scavenging prohibited autophagy induction by thiopurines. Our data indicate that the anticarcinogenic effects of thiopurines are mediated by complex interplay between cellular mechanisms governing redox homeostasis, apoptosis and autophagy.

  • 22. Chlichlia, K.
    et al.
    Los, Marek Jan
    Department of Immunology and Cell Biology, University of Muenster, Roentgenstr. 21, D-48149 Muenster, Germany.
    Schulze-Osthoff, Klaus
    Department of Immunology and Cell Biology, University of Muenster, Roentgenstr. 21, D-48149 Muenster, Germany.
    Gazzolo, L.
    INSERM U412, Ecole Normale S périeure de Lyon, 69367 Lyon, Cedex 07, France.
    Schirrmacher, V.
    Division of Cellular Immunology (G0100), Tumor Immunology Program, German Cancer Research Center, Im Neuenheimer Feld 280, D69120 Heidelberg, Germany.
    Khazaie, K.
    Division of Cellular Immunology (G0100), Tumor Immunology Program, German Cancer Research Center, Im Neuenheimer Feld 280, D69120 Heidelberg, Germany; 4Department of Cancer Immunology and AIDS, Dana Farber Cancer Institute, Boston, MA 02115, U.S.A..
    Redox events in HTLV-1 tax-induced apoptotic T-cell death2002Ingår i: Antioxidants and Redox Signaling, ISSN 1523-0864, E-ISSN 1557-7716, Vol. 4, nr 3, s. 471-477Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    A number of studies implicate reactive oxygen intermediates in the induction of DNA damage and apoptosis. Recent studies suggest that the human T-cell leukemia virus type I (HTLV-1) Tax protein induces oxidative stress and apoptotic T-cell death. Activation of the T-cell receptor/CD3 pathway enhances the Tax-mediated oxidative and apoptotic effects. Tax-mediated apoptosis and oxidative stress as well as activation of nuclear factor-kappaB can be potently suppressed by antioxidants. This review focuses on Tax-dependent changes in the intracellular redox status and their role in Tax-mediated DNA damage and apoptosis. The relevance of these observations to HTLV-1 virus-mediated T-cell transformation and leukemogenesis are discussed.

  • 23.
    Christoffersson, Jonas
    et al.
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Teknisk biologi. Linköpings universitet, Tekniska fakulteten.
    Aronsson, Christopher
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Molekylär fysik. Linköpings universitet, Tekniska fakulteten.
    Jury, Michael
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Molekylär fysik. Linköpings universitet, Tekniska fakulteten.
    Selegård, Robert
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Molekylär fysik. Linköpings universitet, Tekniska fakulteten.
    Aili, Daniel
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Molekylär fysik. Linköpings universitet, Tekniska fakulteten.
    Mandenius, Carl-Fredrik
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Teknisk biologi. Linköpings universitet, Tekniska fakulteten.
    Fabrication of modular hyaluronan-PEG hydrogels to support 3D cultures of hepatocytes in a perfused liver-on-a-chip device2018Ingår i: Biofabrication, ISSN 1758-5082, E-ISSN 1758-5090, Vol. 11, nr 1, s. 1-13, artikel-id 015013Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Liver cell culture models are attractive in both tissue engineering and for development of assays for drug toxicology research. To retain liver specific cell functions, the use of adequate cell types and culture conditions, such as a 3D orientation of the cells and a proper supply of nutrients and oxygen, are critical. In this article, we show how extracellular matrix mimetic hydrogels can support hepatocyte viability and functionality in a perfused liver-on-a-chip device. A modular hydrogel system based on hyaluronan and poly(ethylene glycol) (HA-PEG), modified with cyclooctyne moieties for bioorthogonal strain-promoted alkyne-azide 1, 3-dipolar cycloaddition (SPAAC), was developed, characterized, and compared for cell compatibility to hydrogels based on agarose and alginate. Hepatoma cells (HepG2) formed spheroids with viable cells in all hydrogels with the highest expression of albumin and urea in alginate hydrogels. By including an excess of cyclooctyne in the HA backbone, azide-modified cell adhesion motifs (linear and cyclic RGD peptides) could be introduced in order to enhance viability and functionality of human induced pluripotent stem cell derived hepatocytes (hiPS-HEPs). In the HA-PEG hydrogels modified with cyclic RGD peptides hiPS-HEPs migrated and grew in 3D and showed an increased viability and higher albumin production compared to when cultured in the other hydrogels. This flexible SPAAC crosslinked hydrogel system enabled fabrication of perfused 3D cell culture of hiPS-HEPs and is a promising material for further development and optimization of liver-on-a-chip devices.

  • 24.
    Clayton, Zoe E.
    et al.
    Heart Res Inst, Australia.
    Tan, Richard P.
    Heart Res Inst, Australia; Univ Sydney, Australia.
    Miravet, Maria M.
    Linköpings universitet, Medicinska fakulteten. Heart Res Inst, Australia.
    Lennartsson, Katarina
    Linköpings universitet, Medicinska fakulteten. Heart Res Inst, Australia.
    Cooke, John P.
    Houston Methodist Res Inst, TX 77030 USA.
    Bursill, Christina A.
    Heart Res Inst, Australia.
    Wise, Steven G.
    Heart Res Inst, Australia.
    Patel, Sanjay
    Heart Res Inst, Australia; Univ Sydney, Australia.
    Induced pluripotent stem cell-derived endothelial cells promote angiogenesis and accelerate wound closure in a murine excisional wound healing mode2018Ingår i: Bioscience Reports, ISSN 0144-8463, E-ISSN 1573-4935, Vol. 38, artikel-id BSR20180563Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Chronic wounds are a major complication in patients with cardiovascular diseases. Cell therapies have shown potential to stimulate wound healing, but clinical trials using adult stem cells have been tempered by limited numbers of cells and invasive procurement procedures. Induced pluripotent stem cells (iPSCs) have several advantages of other cell types, for example they can be generated in abundance from patients somatic cells (autologous) or those from a matched donor. iPSCs can be efficiently differentiated to functional endothelial cells (iPSC-ECs). Here, we used a murine excisional wound model to test the pro-angiogenic properties of iPSC-ECs in wound healing. Two full-thickness wounds were made on the dorsum of NOD-SCID mice and splinted. iPSC-ECs (5 x 10(5)) were topically applied to one wound, with the other serving as a control. Treatment with iPSC-ECs significantly increased wound perfusion and accelerated wound closure. Expression of endothelial cell (EC) surface marker, platelet endothelial cell adhesion molecule (PECAM-1) (CD31), and pro-angiogenic EC receptor, Tie1, mRNA was up-regulated in iPSC-EC treated wounds at 7 days post-wounding. Histological analysis of wound sections showed increased capillary density in iPSC-EC wounds at days 7 and 14 post-wounding, and increased collagen content at day 14. Anti-GFP fluorescence confirmed presence of iPSC-ECs in the wounds. Bioluminescent imaging (BLI) showed progressive decline of iPSC-ECs over time, suggesting that iPSC-ECs are acting primarily through short-term paracrine effects. These results highlight the pro-regenerative effects of iPSC-ECs and demonstrate that they are a promising potential therapy for intractable wounds.

  • 25.
    Domert, Jakob
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Rao, Sahana Bhima
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Agholme, Lotta
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Brorsson, Ann-Christin
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Kemi. Linköpings universitet, Tekniska högskolan.
    Marcusson, Jan
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för neurovetenskap. Linköpings universitet, Hälsouniversitetet.
    Hallbeck, Martin
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för inflammationsmedicin. Linköpings universitet, Hälsouniversitetet. Östergötlands Läns Landsting, Diagnostikcentrum, Klinisk patologi och klinisk genetik.
    Nath, Sangeeta
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Spreading of Amyloid-β Peptides via Neuritic Cell-to-cell Transfer Is Dependent on Insufficient Cellular Clearance2014Ingår i: Neurobiology of Disease, ISSN 0969-9961, E-ISSN 1095-953X, Vol. 65, s. 82-92Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The spreading of pathology through neuronal pathways is likely to be the cause of the progressive cognitive loss observed in Alzheimer's disease (AD) and other neurodegenerative diseases. We have recently shown the propagation of AD pathology via cell-to-cell transfer of oligomeric amyloid beta (Aβ) residues 1-42 (oAβ1-42) using our donor-acceptor 3-D co-culture model. We now show that different Aβ-isoforms (fluorescently labeled 1-42, 3(pE)-40, 1-40 and 11-42 oligomers) can transfer from one cell to another. Thus, transfer is not restricted to a specific Aβ-isoform. Although different Aβ isoforms can transfer, differences in the capacity to clear and/or degrade these aggregated isoforms result in vast differences in the net amounts ending up in the receiving cells and the net remaining Aβ can cause seeding and pathology in the receiving cells. This insufficient clearance and/or degradation by cells creates sizable intracellular accumulations of the aggregation-prone Aβ1-42 isoform, which further promotes cell-to-cell transfer; thus, oAβ1-42 is a potentially toxic isoform. Furthermore, cell-to-cell transfer is shown to be an early event that is seemingly independent of later appearances of cellular toxicity. This phenomenon could explain how seeds for the AD pathology could pass on to new brain areas and gradually induce AD pathology, even before the first cell starts to deteriorate, and how cell-to-cell transfer can act together with the factors that influence cellular clearance and/or degradation in the development of AD.

  • 26.
    Englund, Ulrika
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Brask, Johan
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Elinder, Fredrik
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Inhibition of SCN2A ortholog upregulation in Xenopus laevis oocytes prevents cell death2014Manuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    Transport of ions across the cell membrane is essential for the regulation of cell death and tissue homeostasis, and alterations in the function of voltage-gated ion channels and of the intracellular ionic compositions interfere with these processes. Opening of K, Na , or Cl channels have been linked to the apoptotic process and in many cases, opening of these channels precede caspase-3 activation and are thus early events in the apoptotic process. Consistent with the role of these channels in apoptosis, inhibition of these channels prevents or delays the apoptotic process. However, the role of ion channels during apoptosis has been difficult to explore, mainly due to unspecific/non-selective ion channe blockers. In the present investigation, the molecular identity of a  voltage-gated Na channel in oocytes from Xenopus laevis, which is crucial for the apoptotic response to mechanical stress, was identified. Specific down regulation of SCN2A Na channel expression by miRNA prevented apoptosis, suggesting that Na+ influx is essential for apoptosis in Xenopus oocytes.

  • 27.
    Ericsson, Maria
    et al.
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biologi. Linköpings universitet, Tekniska fakulteten.
    Henriksen, Rie
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biologi. Linköpings universitet, Tekniska fakulteten.
    Bélteky, Johan
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biologi. Linköpings universitet, Tekniska fakulteten.
    Sundman, Ann-Sofie
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biologi. Linköpings universitet, Tekniska fakulteten.
    Shionoya, Kiseko
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Medicinska fakulteten.
    Jensen, Per
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biologi. Linköpings universitet, Tekniska fakulteten.
    Long-Term and Transgenerational Effects of Stress Experienced during Different Life Phases in Chickens (Gallus gallus)2016Ingår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 11, nr 4, artikel-id e0153879Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Stress in animals causes not only immediate reactions, but may affect their biology for long periods, even across generations. Particular interest has been paid to perinatal stress, but also adolescence has been shown to be a sensitive period in mammals. So far, no systematic study has been performed of the relative importance of stress encountered during different life phases. In this study, groups of chickens were exposed to a six-day period of repeated stress during three different life phases: early (two weeks), early puberty (eight weeks) and late puberty (17 weeks), and the effects were compared to an unstressed control group. The short-term effects were assessed by behaviour, and the long-term and transgenerational effects were determined by effects on behavior and corticosterone secretion, as well as on hypothalamic gene expression. Short-term effects were strongest in the two week group and the eight week group, whereas long-term and transgenerational effects were detected in all three stress groups. However, stress at different ages affected different aspects of the biology of the chickens, and it was not possible to determine a particularly sensitive life phase. The results show that stress during puberty appears to be at least equally critical as the previously studied early life phase. These findings may have important implications for animal welfare in egg production, since laying hens are often exposed to stress during the three periods pinpointed here.

  • 28.
    Eriksson, Emma S. E.
    et al.
    Department of Chemistry and Molecular Biology, University of Gothenburg.
    Erdtman, Edvin
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Kemi. Linköpings universitet, Tekniska fakulteten.
    Eriksson, Leif A.
    Department of Chemistry and Molecular Biology, University of Gothenburg.
    Permeability of 5-aminolevulinic acid oxime derivatives in lipid membranes2016Ingår i: Theoretical Chemistry accounts, ISSN 1432-881X, E-ISSN 1432-2234, Vol. 135, nr 1, s. 1-9Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The endogenous molecule 5-aminolevulinic acid (5ALA) and its methyl ester (Me-5ALA) have been used as prodrugs in photodynamic treatment of actinic keratosis and superficial non-melanoma skin cancers for over a decade. Recently, a novel set of 5ALA derivatives based on introducing a hydrolyzable oxime functionality was proposed and shown to generate considerably stronger onset of the photoactive molecule protoporphyrin IX (PpIX) in the cells. In the current work, we employ molecular dynamics simulation techniques to explore whether the higher intercellular concentration of PpIX caused by the oxime derivatives is related to enhanced membrane permeability, or whether other factors contribute to this. It is concluded that the oximes show overall similar accumulation at the membrane headgroup regions as the conventional derivatives and that the transmembrane permeabilities are in general close to that of 5ALA. The highest permeability of all compounds explored is found for Me-5ALA, which correlates with a considerably lower fee energy barrier at the hydrophobic bilayer center. The high PpIX concentration must hence be sought in other factors, where slow hydrolysis of the oxime functionality is a plausible reason, enabling stronger buildup of PpIX over time.

  • 29.
    Eriksson, Ida
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Medicinska fakulteten.
    Nath, Sangeeta
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Medicinska fakulteten.
    Bornefall, Per
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för neuro- och inflammationsvetenskap. Linköpings universitet, Medicinska fakulteten.
    Villamil Giraldo, Ana Maria
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Medicinska fakulteten.
    Öllinger, Karin
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Medicinska fakulteten. Region Östergötland, Diagnostikcentrum, Klinisk patologi och klinisk genetik.
    Impact of high cholesterol in a Parkinsons disease model: Prevention of lysosomal leakage versus stimulation of alpha-synuclein aggregation2017Ingår i: European Journal of Cell Biology, ISSN 0171-9335, E-ISSN 1618-1298, Vol. 96, nr 2, s. 99-109Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Parkinsons disease is characterized by accumulation of intraneuronal cytoplasmic inclusions, Lewy bodies, which mainly consist of aggregated alpha-synuclein. Controversies exist as to whether high blood cholesterol is a risk factor for the development of the disease and whether statin treatment could have a protective effect. Using a model system of BE(2)-M17 neuroblastoma cells treated with the neurotoxin 1-methyl-4-phenylpyridinium (MPP+), we found that MPP+-induced cell death was accompanied by cholesterol accumulation in a lysosomal-like pattern in pre-apoptotic cells. To study the effects of lysosomal cholesterol accumulation, we increased lysosomal cholesterol through pre-treatment with U18666A and found delayed leakage of lysosomal contents into the cytosol, which reduced cell death. This suggests that increased lysosomal cholesterol is a stress response mechanism to protect lysosomal membrane integrity in response to early apoptotic stress. However, high cholesterol also stimulated the accumulation of alpha-synuclein. Treatment with the cholesterol-lowering drug lovastatin reduced MPP+-induced cell death by inhibiting the production of reactive oxygen species, but did not prevent lysosomal cholesterol increase nor affect alpha-synuclein accumulation. Our study indicates a dual role of high cholesterol in Parkinsons disease, in which it acts both as a protector against lysosomal membrane permeabilization and as a stimulator of alpha-synuclein accumulation. (C) 2017 Elsevier GmbH. All rights reserved.

  • 30.
    Eriksson, Ida
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Medicinska fakulteten.
    Öllinger, Karin
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Medicinska fakulteten. Region Östergötland, Diagnostikcentrum, Klinisk patologi och klinisk genetik.
    Appelqvist, Hanna
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Kemi. Linköpings universitet, Tekniska fakulteten.
    Analysis of Lysosomal pH by Flow Cytometry Using FITC-Dextran Loaded Cells2017Ingår i: Lysosomes: Methods and Protocols / [ed] Karin Öllinger;Hanna Appelqvist, Humana Press, 2017, Vol. 1594, s. 179-189Kapitel i bok, del av antologi (Refereegranskat)
    Abstract [en]

    The acidic environment of the lysosomal lumen provides an optimal milieu for the acid hydrolases and is also essential for fusion/fission of endo-lysosomal compartments and sorting of cargo. Evidence suggests that maintaining lysosomal acidity is essential to avoid disease. In this chapter, we describe a protocol for analyzing the lysosomal pH in cultured cells using the fluorescent probe fluorescein isothiocyanate (FITC)-dextran together with a dual-emission ratiometric technique suitable for flow cytometry. Fluorescence-labeled dextran is endocytosed and accumulated in the lysosomal compartment. FITC shows a pH-dependent variation in fluorescence when analyzed at maximum emission wavelength and no variation when analyzing at the isosbestic point, thereby the ratio can be used to determine the lysosomal pH. A standard curve is obtained by equilibrating intralysosomal pH with extracellular pH using the ionophore nigericin. The protocol also includes information regarding procedures to induce lysosomal alkalinization and lysosomal membrane permeabilization.

  • 31.
    Everett, Jake
    et al.
    Department of Surgery, Texas Tech University Health Sciences Center, MS8312, 3601 4th Street, Lubbock, USA.
    Gabrilska, Rebecca
    Department of Surgery, Texas Tech University Health Sciences Center, MS8312, 3601 4th Street, Lubbock, USA.
    Rumbaugh, Kendra P.
    Department of Surgery, Texas Tech University Health Sciences Center, MS8312, 3601 4th Street, Lubbock, USA.
    Vikström, Elena
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för mikrobiologi och molekylär medicin. Linköpings universitet, Medicinska fakulteten.
    Assessing Pseudomonas aeruginosa Autoinducer Effects on Mammalian Epithelial Cells2018Ingår i: Quorum Sensing: Methods and Protocols / [ed] Livia LeoniGiordano Rampioni, Humana Press, 2018, Vol. 1673, s. 213-225Kapitel i bok, del av antologi (Refereegranskat)
    Abstract [en]

    The human mucosal environment in the gut is rich with interactions between microbiota and mammalian epithelia. Microbes such as the Gram-negative bacterium Pseudomonas aeruginosa may use quorum sensing to communicate with other microorganisms and mammalian cells to alter gene expression. Here, we present methodologies to evaluate the effects of P. aeruginosa N-(3-oxo-dodecanoyl)-L-homoserine lactone (3O-C12-HSL) on Caco-2 cell monolayers. First, we describe a method for assessing barrier function and permeability of epithelial cells when exposed to 3O-C12-HSL by measuring transepithelial electrical resistance (TER) and paracellular flow using fluorescently labeled dextran. Secondly, we detail methods to investigate the effect of 3O-C12-HSL on protein-protein interactions of epithelial junction proteins. Lastly, we will detail imaging techniques to visualize Caco-2 barrier disruption following exposure to 3O-C12-HSL through the use of confocal laser scanning microscopy (CLSM) and a super resolution technique, stimulated emission depletion (STED) microscopy, to achieve nanoscale visualization of Caco-2 monolayers.

  • 32.
    Fernández-Gago, Rocío
    et al.
    Department of Medicine, Surgery and Veterinary Anatomy, University of León, 24071 León, Spain.
    Alvarez-Rodriguez, Manuel
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Alonso, Marta E
    Department of Animal Production, University of León, 24071 León, Spain.
    González, J Ramiro
    Department of Medicine, Surgery and Veterinary Anatomy, University of León, 24071 León, Spain.
    Alegre, Beatriz
    Department of Medicine, Surgery and Veterinary Anatomy, University of León, 24071 León, Spain.
    Domínguez, Juan C
    Department of Medicine, Surgery and Veterinary Anatomy, University of León, 24071 León, Spain.
    Martínez-Pastor, Felipe
    INDEGSAL (Institute for Animal Health and Cattle Development), University of León, 24071 León, Spain /Molecular Biology (Cell Biology), University of León, 24071 León, Spain/ .
    Thawing boar semen in the presence of seminal plasma improves motility, modifies subpopulation patterns and reduces chromatin alterations.2017Ingår i: Reproduction, fertility, and development, ISSN 1031-3613, Vol. 29, nr 8, s. 1576-1584Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Seminal plasma could have positive effects on boar semen after thawing. In the present study we investigated changes in the motility and chromatin structure in spermatozoa over 4h incubation (37°C) of boar semen thawed in the presence of 0%, 10% or 50% seminal plasma from good-fertility boars. Cryopreserved doses were used from seven males, three of which were identified as susceptible to post-thawing chromatin alterations. Motility was analysed by computer-aided sperm analysis every hour, and data were used in a two-step clustering, yielding three subpopulations of spermatozoa (slow non-linear, fast non-linear, fast linear). Chromatin structure was analysed using a sperm chromatin structure assay and flow cytometry to determine the DNA fragmentation index (%DFI) as a percentage, the standard deviation of the DFI (SD-DFI) and the percentage of high DNA stainability (%HDS), indicating chromatin compaction. Thawing without seminal plasma resulted in a rapid loss of motility, whereas seminal plasma helped maintain motility throughout the incubation period and preserved the subpopulation comprising fast and linear spermatozoa. The incidence of chromatin alterations was very low in samples from non-susceptible males, whereas samples from males susceptible to post-thawing chromatin alterations exhibited marked alterations in%DFI and%HDS. Seminal plasma partly prevented these alterations in samples from susceptible males. Overall, 50% seminal plasma was the most efficient concentration to protect motility and chromatin. Some changes were concomitant with physiological events reported previously (e.g., semen thawed with 50% seminal plasma increased the production of reactive oxygen species and yielded higher fertility after AI). Thawing in the presence of seminal plasma could be particularly useful in the case of samples susceptible to post-thawing chromatin damage.

  • 33.
    Ferrari, D.
    et al.
    Department of Internal Medicine I, Eberhard-Karls University, D-72076 Tübingen, Germany.
    Stepczynska, A.
    Department of Internal Medicine I, Eberhard-Karls University, D-72076 Tübingen, Germany.
    Los, Marek Jan
    Department of Internal Medicine I, Eberhard-Karls University, D-72076 Tübingen, Germany.
    Wesselborg, Sebastian
    Department of Internal Medicine I, Eberhard-Karls University, D-72076 Tübingen, Germany.
    Schulze-Osthoff, Klaus
    Department of Internal Medicine I, Eberhard-Karls University, D-72076 Tübingen, Germany.
    Differential regulation and ATP requirement for caspase-8 and caspase-3 activation during CD95- and anticancer drug-induced apoptosis1998Ingår i: Journal of Experimental Medicine, ISSN 0022-1007, E-ISSN 1540-9538, Vol. 188, nr 5, s. 979-984Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Apoptosis is induced by different stimuli, among them triggering of the death receptor CD95, staurosporine, and chemotherapeutic drugs. In all cases, apoptosis is mediated by caspases, although it is unclear how these diverse apoptotic stimuli cause protease activation. Two regulatory pathways have been recently identified, but it remains unknown whether they are functionally independent or linked to each other. One is mediated by recruitment of the proximal regulator caspase-8 to the death receptor complex. The other pathway is controlled by the release of cytochrome c from mitochondria and the subsequent ATP-dependent activation of the death regulator apoptotic protease-activating factor 1 (Apaf-1). Here, we report that both pathways can be dissected by depletion of intracellular ATP. Prevention of ATP production completely inhibited caspase activation and apoptosis in response to chemotherapeutic drugs and staurosporine. Interestingly, caspase-8, whose function appeared to be restricted to death receptors, was also activated by these drugs under normal conditions, but not after ATP depletion. In contrast, inhibition of ATP production did not affect caspase activation after triggering of CD95. These results suggest that chemotherapeutic drug-induced caspase activation is entirely controlled by a receptor-independent mitochondrial pathway, whereas CD95-induced apoptosis can be regulated by a separate pathway not requiring Apaf-1 function.

  • 34.
    Franz, F
    et al.
    Clinic of Endocrinology and Nephrology, Department of Internal Medicine, Neurology and Dermatology, University of Leipzig, Leipzig, Germany.
    Weidinger, C
    Clinic of Endocrinology and Nephrology, Department of Internal Medicine, Neurology and Dermatology, University of Leipzig, Leipzig, Germany.
    Krause, K
    Clinic of Endocrinology and Nephrology, Department of Internal Medicine, Neurology and Dermatology, University of Leipzig, Leipzig, Germany.
    Gimm, Oliver
    Department of General, Visceral, and Vascular Surgery, Martin-Luther-University of Halle-Wittenberg, Halle/Saale, Germany.
    Dralle, H
    Department of General, Visceral, and Vascular Surgery, Martin-Luther-University of Halle-Wittenberg, Halle/Saale, Germany.
    Führer, D
    Department of Endocrinology & Metabolism and Division of Laboratory Research, University Hospital Essen, Essen, Germany.
    The Transcriptional Regulation of FOXO Genes in Thyrocytes.2016Ingår i: Hormone and Metabolic Research, ISSN 0018-5043, E-ISSN 1439-4286, Vol. 48, nr 9, s. 601-6Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    FOXO transcription factors are key regulators of DNA damage repair, proliferation and apoptosis in thyrocytes. Thyroid malignancies show impaired FOXO function. In this study, we investigated the transcriptional regulation of FOXO isoforms in thyroid epithelial cells. mRNA expression of FOXO isoforms (FOXO1, 3 and 4) was determined in FRTL-5 cells stimulated with different growth factors and H2O2. Furthermore, the impact of PI3K/AKT signalling on FOXO transcription was investigated in PI3K p110α mutant FRTL-5 cells and regulatory dependence of FOXO transcription on FOXO was studied in FRTL-5 cells with hFOXO3 overexpression. Finally, mRNA expression levels of FOXO isoforms were determined in human epithelial thyroid tumours. Growth factor deprivation induced transcription of FOXO1, 3 and 4, whereas insulin stimulation decreased FOXO1 and FOXO4 transcription in FRTL-5 cells. Inhibition of the PI3K/AKT cascade amplified FOXO1 and FOXO4 expression. In contrast, H2O2 and TSH did not influence FOXO transcription in thyrocytes. Overexpression of PI3K p110α inhibited FOXO3 and induced FOXO4 transcription. In human thyroid tumours, FOXO1 and FOXO3 mRNA levels were significantly downregulated in papillary thyroid carcinoma when compared to normal tissues. In contrast, follicular thyroid carcinomas showed significant upregulation of FOXO4 mRNA.In this paper, we demonstrate an influence of PI3K signalling on FOXO transcription in thyrocytes. Moreover, we show that thyroid cancers exhibit alterations in FOXO transcription besides the previously reported alterations in posttranslational FOXO3 regulation. These findings may add to the concept of targeting the PI3K pathway in advanced thyroid cancers.

  • 35.
    Gardela, Jaume
    et al.
    Univ Autonoma Barcelona, Spain.
    Ruiz-Conca, Mateo
    Univ Autonoma Barcelona, Spain.
    Alvarez-Rodriguez, Manuel
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten. Univ Autonoma Barcelona, Spain.
    Mogas, Teresa
    Univ Autonoma Barcelona, Spain.
    Lopez-Bejar, Manel
    Univ Autonoma Barcelona, Spain.
    Induction of CIRBP expression by cold shock on bovine cumulus-oocyte complexes2019Ingår i: Reproduction in domestic animals, ISSN 0936-6768, E-ISSN 1439-0531, Vol. 54, s. 82-85Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The aim of this study was to induce the cold-inducible RNA-binding protein (CIRBP) expression on cumulus-oocyte complexes (COCs) through exposure to a sub-lethal cold shock and determine the effects of hypothermic temperatures during the in vitro maturation of bovine oocytes. Nuclear maturation, cortical granule redistribution and identification of cold-inducible RNA-binding protein (CIRBP) were assessed after 24 hr of in vitro maturation of control (38.5 degrees C) and cold-stressed oocytes (33.5 degrees C). The presence of CIRBP was assessed by Western blot in COCs or denuded oocytes and their respective cumulus cells. Based on the odds ratio, cold-stressed oocytes presented higher abnormal cytoplasmic distribution of cortical granules and nuclear maturation than the control group. Although CIRBP was detected in both control and cold-stressed groups, cold-stressed COCs had 2.17 times more expression of CIRBP than control COCs. However, when denuded oocytes and cumulus cells were assessed separately, CIRBP only was detected in cumulus cells in both groups. In conclusion, cold shock induced CIRBP expression, but it negatively affected nuclear maturation and cortical granule distribution of bovine oocytes. Moreover, the expression of CIRBP was only identified in cumulus cells but not in oocytes.

  • 36.
    Ghavami, Saeid
    et al.
    Department of Biochemistry and Medical Genetics, Manitoba Institute of Cell Biology, Cancer Care Manitoba, Winnipeg, Manitoba, Canada.
    Asoodeh, Ahmad
    Department of Biophysics and Biochemistry, Faculty of Basic Sciences, Tarbiat Modares University, Tehran, Iran; Department of Chemistry Faculty of Science, Ferdowsi University of Mashhad, Mashhad, Iran .
    Klonisch, Thomas
    Department of Human Anatomy and Cell Sciences, and Manitoba Institute of Child Health, Winnipeg, Canada.
    Halayko, Andrew J.
    Department of Physiology, University of Manitoba, Winnipeg, MB, Canada; Manitoba Institute of Child Health, Winnipeg, MB, Canada.
    Kadkhoda, Kamran
    Department of Biochemistry and Medical Genetics, Manitoba Institute of Cell Biology, Cancer Care Manitoba, Winnipeg, Manitoba, Canada.
    Kroczak, Tadeusz J.
    Department of Biochemistry and Medical Genetics, Manitoba Institute of Cell Biology, Cancer Care Manitoba, Winnipeg, Manitoba, Canada.
    Gibson, Spencer B.
    Department of Biochemistry and Medical Genetics, Manitoba Institute of Cell Biology, Cancer Care Manitoba, Winnipeg, Manitoba, Canada; BioApplications Enterprises, Winnipeg, Canada.
    Booy, Evan P.
    Department of Biochemistry and Medical Genetics, Manitoba Institute of Cell Biology, Cancer Care Manitoba, Winnipeg, Manitoba, Canada.
    Naderi-Manesh, Hossein
    Department of Biophysics and Biochemistry, Faculty of Basic Sciences, Tarbiat Modares University, Tehran, Iran.
    Los, Marek Jan
    BioApplications Enterprises, Winnipeg, MB, Canada.
    Brevinin-2R semi-selectively kills cancer cells by a distinct mechanism, which involves the lysosomal-mitochondrial death pathway2008Ingår i: Journal of Cellular and Molecular Medicine (Print), ISSN 1582-1838, E-ISSN 1582-4934, Vol. 12, nr 3, s. 1005-1022Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Brevinin-2R is a novel non-hemolytic defensin that was isolated from the skin of the frog Rana ridibunda. It exhibits preferential cytotoxicity towards malignant cells, including Jurkat (T-cell leukemia), BJAB (B-cell lymphoma), HT29/219, SW742 (colon carcinomas), L929 (fibrosarcoma), MCF-7 (breast adenocarcinoma), A549 (lung carcinoma), as compared to primary cells including peripheral blood mononuclear cells (PBMC), T cells and human lung fibroblasts. Jurkat and MCF-7 cells overexpressing Bcl2, and L929 and MCF-7 over-expressing a dominant-negative mutant of a pro-apoptotic BNIP3 (ΔTM-BNIP3) were largely resistant towards Brevinin-2R treatment. The decrease in mitochondrial membrane potential (ΔΨm), or total cellular ATP levels, and increased reactive oxygen species (ROS) production, but not caspase activation or the release of apoptosis-inducing factor (AIF) or endonuclease G (Endo G), were early indicators of Brevinin-2R-triggered death. Brevinin-2R interacts with both early and late endosomes. Lysosomal membrane permeabilization inhibitors and inhibitors of cathepsin-B and cathepsin-L prevented Brevinin-2R-induced cell death. Autophagosomes have been detected upon Brevinin-2R treatment. Our results show that Brevinin-2R activates the lysosomalmitochondrial death pathway, and involves autophagy-like cell death.

  • 37.
    Ghavami, Saeid
    et al.
    Manitoba Institute of Cell Biology, and Department of Biochemistry and Medical Genetics, University of Manitoba, Canada.
    Eshraghi, Mehdi
    Manitoba Institute of Cell Biology, and Department of Biochemistry and Medical Genetics, University of Manitoba, Canada.
    Kadkhoda, Kamran
    Manitoba Institute of Cell Biology, and Department of Biochemistry and Medical Genetics, University of Manitoba, Canada.
    Mutawe, Mark M.
    Department of Physiology, University of Manitoba, Canada; Manitoba Institute of Child's Health, University of Manitoba, Canada.
    Maddika, Subbareddy
    Manitoba Institute of Cell Biology, and Department of Biochemistry and Medical Genetics, University of Manitoba, Canada.
    Bay, Graham H.
    Manitoba Institute of Cell Biology, and Department of Biochemistry and Medical Genetics, University of Manitoba, Canada.
    Wesselborg, Sebastian
    Department of Internal Medicine I, University of Tübingen, Tübingen, Germany.
    Halayko, Andrew J.
    Department of Physiology, University of Manitoba, Winnipeg, MB, Canada; Manitoba Institute of Child Health, Winnipeg, MB, Canada; Department of Internal Medicine, University of Manitoba, Winnipeg, MB, Canada .
    Klonisch, Thomas
    Department of Human Anatomy and Cell Sciences, and Manitoba Institute of Child Health, Winnipeg, Canada.
    Los, Marek Jan
    Interfaculty Institute for Biochemistry, University of Tübingen, Germany; BioApplications Enterprises, Winnipeg, MB, Canada.
    Role of BNIP3 in TNF-induced cell death - TNF upregulates BNIP3 expression2009Ingår i: Biochimica et Biophysica Acta. Molecular Cell Research, ISSN 0167-4889, E-ISSN 1879-2596, Vol. 1793, nr 3, s. 546-560Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Tumor necrosis factor alpha (TNF) is a cytokine that induces caspase-dependent (apoptotic) and caspase-independent (necrosis-like) cell death in different cells. We used the murine fibrosarcoma cell line model L929 and a stable L929 transfectant over-expressing a mutated dominant-negative form of BNIP3 lacking the C-terminal transmembrane (TM) domain (L929-ΔTM-BNIP3) to test if TNF-induced cell death involved pro-apoptotic Bcl2 protein BNIP3. Treatment of cells with TNF in the absence of actinomycin D caused a rapid fall in the mitochondrial membrane potential (ΔΨm) and a prompt increase in reactive oxygen species (ROS) production, which was significantly less pronounced in L929-ΔTM-BNIP3. TNF did not cause the mitochondrial release of apoptosis inducing factor (AIF) and Endonuclease G (Endo-G) but provoked the release of cytochrome c, Smac/Diablo, and Omi/HtrA2 at similar levels in both L929 and in L929-ΔTM-BNIP3 cells. We observed TNF-associated increase in the expression of BNIP3 in L929 that was mediated by nitric oxide and significantly inhibited by nitric oxide synthase inhibitor N5-(methylamidino)-l-ornithine acetate. In L929, lysosomal swelling and activation were markedly increased as compared to L929-ΔTM-BNIP3 and could be inhibited by treatment with inhibitors to vacuolar H+-ATPase and cathepsins −B/−L. Together, these data indicate that TNF-induced cell death involves BNIP3, ROS production, and activation of the lysosomal death pathway.

  • 38.
    Ghavami, Saeid
    et al.
    Department of Physiology, University of Manitoba, Winnipeg, MB, Canada; National Training Program in Allergy and Asthma, University of Manitoba, Winnipeg, MB, Canada; Biology of Breathing Group, Manitoba Institute of Child Health, Winnipeg, MB, Canada.
    Eshragi, Mehdi
    Department of Biochemistry and Medical Genetics, Manitoba Institute of Cell Biology, CancerCare Manitoba, Winnipeg, Canada.
    Ande, Sudharsana R
    Department of Biochemistry and Medical Genetics, Manitoba Institute of Cell Biology, CancerCare Manitoba, Winnipeg, Canada.
    Chazin, Walter J
    Department of Biochemistry and Chemistry, Center for Structural Biology, Vanderbilt University, Nashville, USA.
    Klonisch, Thomas
    Department of Human Anatomy and Cell Science, University of Manitoba, Winnipeg, Canada.
    Halayko, Andrew J.
    Department of Physiology, University of Manitoba, Winnipeg, MB, Canada; Manitoba Institute of Child Health, Winnipeg, MB, Canada; Department of Internal Medicine, University of Manitoba, Winnipeg, MB, Canada .
    Mcneill, Karol D
    Manitoba Institute of Child Health, University of Manitoba, Winnipeg, Canada.
    Hashemi, Mohammad
    Department of Clinical Biochemistry, Zahedan University of Medical Sciences, Zahedan, Iran.
    Kerkhoff, Claus
    Institute of Immunology, University of Muenster, Roentgenstr. 21, Muenster, Germany.
    Los, Marek Jan
    Interfaculty Institute for Biochemistry, University of Tübingen, Germany.
    S100A8/A9 induces autophagy and apoptosis via ROS-mediated cross-talk between mitochondria and lysosomes that involves BNIP32010Ingår i: Cell Research, ISSN 1001-0602, E-ISSN 1748-7838, Vol. 20, nr 3, s. 314-331Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The complex formed by two members of the S100 calcium-binding protein family, S100A8/A9, exerts apoptosis-inducing activity in various cells of different origins. Here, we present evidence that the underlying molecular mechanisms involve both programmed cell death I (PCD I, apoptosis) and PCD II (autophagy)-like death. Treatment of cells with S100A8/A9 caused the increase of Beclin-1 expression as well as Atg12-Atg5 formation. S100A8/A9-induced cell death was partially inhibited by the specific PI3-kinase class III inhibitor, 3-methyladenine (3-MA), and by the vacuole H+-ATPase inhibitor, bafilomycin-A1 (Baf-A1). S100A8/A9 provoked the translocation of BNIP3, a BH3 only pro-apoptotic Bcl2 family member, to mitochondria. Consistent with this finding, ΔTM-BNIP3 overexpression partially inhibited S100A8/A9-induced cell death, decreased reactive oxygen species (ROS) generation, and partially protected against the decrease in mitochondrial transmembrane potential in S100A8/A9-treated cells. In addition, either ΔTM-BNIP3 overexpression or N-acetyl-L-cysteine co-treatment decreased lysosomal activation in cells treated with S100A8/A9. Our data indicate that S100A8/A9-promoted cell death occurs through the cross-talk of mitochondria and lysosomes via ROS and the process involves BNIP3.

  • 39.
    Ghavami, Saeid
    et al.
    Department of Biochemistry and Medical Genetics, Manitoba Institute of Cell Biology, Cancer Care Manitoba, Winnipeg, Manitoba, Canada.
    Hashemi, M.
    Manitoba Institute of Cell Biology, CancerCare Manitoba, Winnipeg, Canada.
    Kadkhoda, K.
    Manitoba Institute of Cell Biology, CancerCare Manitoba, Winnipeg, Canada.
    Alavian, S. M.
    Manitoba Institute of Cell Biology, CancerCare Manitoba, Winnipeg, Canada.
    Bay, G. H.
    Manitoba Institute of Cell Biology, CancerCare Manitoba, Winnipeg, Canada.
    Los, Marek Jan
    Manitoba Institute of Cell Biology, Cancer Care Manitoba; Manitoba Institute of Child Health; Department of Biochemistry and Medical Genetics; Department of Human Anatomy and Cell Science, University Manitoba, Winnipeg, Canada, .
    Apoptosis in liver diseases - detection and therapeutic applications2005Ingår i: Medical Science Monitor, ISSN 1234-1010, E-ISSN 1643-3750, Vol. 11, nr 11, s. RA337-RA345Artikel, forskningsöversikt (Refereegranskat)
    Abstract [en]

    The liver is continuously exposed to a large antigenic load that includes pathogens, toxins, tumor cells and dietary antigens. Amongst the hepatitis viruses, only hepatitis B virus (HBV) and hepatitis C virus (HCV) cause chronic hepatitis, which can progress to cirrhosis and hepatocellular carcinoma. Of the different antiviral defense systems employed by the tissue, apoptosis significantly contributes to the prevention of viral replication, dissemination, and persistence. Loss of tolerance to the liver autoantigens may result in autoimmune hepatitis (AIH). This review outlines the recent findings that highlight the role and mechanisms of apoptotic processes in the course of liver diseases. Among factors that contribute to liver pathology, we discuss the role of tumor necrosis factor (TNF)-alpha, HBx, ds-PKR, TRAIL, FasL, and IL-1 alpha. Since TNF and FasL-induced hepatocyte apoptosis is implicated in a wide range of liver diseases, including viral hepatitis, alcoholic hepatitis, ischemia/reperfusion liver injury, and fulminant hepatic failure, these items will be discussed in greater detail in this review. We also highlight some recent discoveries that pave the way for the development of new therapeutic strategies by protecting hepatocytes (for example by employing Bcl-2, Bcl-X-L or A1/Bfl-1, IAPs, or synthetic caspase inhibitors), or by the induction of apoptosis in stellate cells. The assessment of the severity of liver disease, as well as monitoring of patients with chronic liver disease, remains a major challenge in clinical hepatology practice. Therefore, a separate chapter is devoted to a novel cytochrome c - based method useful for the diagnosis and monitoring of fulminant hepatitis.

  • 40.
    Ghavami, Saeid
    et al.
    Tarbiat Modarres University, Tehran, Iran.
    Kerkhoff, C.
    Institute of Experimental Dermatology, Münster, Germany .
    Los, Marek Jan
    Institute of Experimental Dermatology, Münster, Germany .
    Hashemi, M.
    Tarbiat Modarres University, Tehran, Iran.
    Sorg, C.
    Institute of Experimental Dermatology, Münster, Germany .
    Karami-Tehrani, F.
    Tarbiat Modarres University, Tehran, Iran.
    Mechanism of apoptosis induced by S100A8/A9 in colon cancer cell lines: the role of ROS and the effect of metal ions2004Ingår i: Journal of Leukocyte Biology, ISSN 0741-5400, E-ISSN 1938-3673, Vol. 76, nr 1, s. 169-175Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The protein complex S100A8/A9, abundant in the cytosol of neutrophils, is secreted from the cells upon cellular activation and induces apoptosis in tumor cell lines and normal fibroblasts in a zinc-reversible manner. In the present study, we present evidence that the S100A8/A9 also exerts its apoptotic effect by a zinc-independent mechanism. Treatment of the colon carcinoma cells with different concentrations of human SI00A8/A9 or the metal ion chelator diethylenetriaminepentacetic acid (DTPA) resulted in a significant increase of cell death. Annexin V/phosphatidylinositol and Hoechst 33258 staining revealed that cell death was mainly of the apoptotic type. A significant increase in the activity of caspase-3 and -9 was observed in both cell lines after treatment. Caspase-8 activation was negligible in both cell lines. The cytotoxicity/apoptotic effect of human SI00A8/A9 and DTPA was inhibited significantly 2 2 (P<0.05) by Zn+2 and Cu+2, more effectively than by Ca2+ and Mg2+. The antioxidant N-acetyl-L-cysteine inhibited the cytotoxicity/apoptotic effect of SI00A8/A9 and DTPA. However, as a result of the different time-courses of both agents and that the S100A8/A9-induced apoptosis was not completely reversed, we conclude that S100A8/A9 exerts its apoptotic effect on two colon carcinoma cell lines through a dual mechanism: one via zinc exclusion from the target cells and the other through a yet-undefined mechanism, probably relaying on the cell-surface receptor(s).

  • 41.
    Ghavami, Saeid
    et al.
    Department of Biochemistry and Medical Genetics, Manitoba Institute of Cell Biology, Cancer Care Manitoba, Winnipeg, Manitoba, Canada.
    Kerkhoff, Claus
    Institute of Experimental Dermatology, Münster, Germany.
    Chazin, Walter J.
    Department of Biochemistry Vanderbilt University, Nashville, USA; Department of Chemistry, Vanderbilt University, Nashville, USA; Center for Structural Biology, Vanderbilt University, Nashville, TN 37232-8725, USA.
    Kadhoka, Kamran
    Manitoba Institute of Cell Biology, Canada; Department of Biochemistry and Medical Genetics, University of Manitoba, Winnipeg, Canada.
    Xiao, Wenyan
    Manitoba Institute of Cell Biology, Canada.
    Zusea, Anne
    Manitoba Institute of Cell Biology, Canada; Department of Biochemistry and Medical Genetics, University of Manitoba, Winnipeg, Canada.
    Hashemi, Mohammad
    Department of Clinical Biochemistry, School of Medicine, Zahedan University of Medical Science, Zahedan, Iran.
    Eshraghi, Mehdi
    Manitoba Institute of Cell Biology, Canada b Department of Biochemistry and Medical Genetics, University of Manitoba, Winnipeg, Canada.
    Schulze-Osthoff, Klaus
    Institute of Molecular Medicine, University of Düsseldorf, Düsseldorf, Germany .
    Klonisch, Thomas
    Department of Human Anatomy and Cell Sciences, and Manitoba Institute of Child Health, Winnipeg, Canada.
    Los, Marek Jan
    Manitoba Institute of Cell Biology, Canada; Department of Biochemistry and Medical Genetics, University of Manitoba, Winnipeg, Canada; BioApplications Enterprises, Winnipeg, MB, Canada.
    S100A8/9 induces cell death via a novel, RAGE-independent pathway that involves selective release of Smac/DIABLO and Omi/HtrA22008Ingår i: Biochimica et Biophysica Acta. Molecular Cell Research, ISSN 0167-4889, E-ISSN 1879-2596, Vol. 1783, nr 2, s. 297-311Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    A complex of two S100 EF-hand calcium-binding proteins S100A8/A9 induces apoptosis in various cells, especially tumor cells. Using several cell lines, we have shown that S100A8/A9-induced cell death is not mediated by the receptor for advanced glycation endproducts (RAGE), a receptor previously demonstrated to engage S100 proteins. Investigation of cell lines either deficient in, or over-expressing components of the death signaling machinery provided insight into the S100A8/A9-mediated cell death pathway. Treatment of cells with S100A8/A9 caused a rapid decrease in the mitochondrial membrane potential (ΔΨm) and activated Bak, but did not cause release of apoptosis-inducing factor (AIF), endonuclease G (Endo G) or cytochrome c. However, both Smac/DIABLO and Omi/HtrA2 were selectively released into the cytoplasm concomitantly with a decrease in Drp1 expression, which inhibits mitochondrial fission machinery. S100A8/A9 treatment also resulted in decreased expression of the anti-apoptotic proteins Bcl2 and Bcl-XL, whereas expression of the pro-apoptotic proteins Bax, Bad and BNIP3 was not altered. Over-expression of Bcl2 partially reversed the cytotoxicity of S100A8/A9. Together, these data indicate that S100A8/A9-induced cell death involves Bak, selective release of Smac/DIABLO and Omi/HtrA2 from mitochondria, and modulation of the balance between pro- and anti-apoptotic proteins.

  • 42.
    Ghavami, Saeid
    et al.
    Department of Physiology, University of Manitoba, Winnipeg, MB, Canada; National Training Program in Allergy and Asthma, University of Manitoba, Winnipeg, MB, Canada; Biology of Breathing Group, Manitoba Institute of Child Health, Winnipeg, MB, Canada.
    Mutawe, Mark M.
    Department of Physiology, University of Manitoba, Winnipeg, MB, Canada; Biology of Breathing Group, Manitoba Institute of Child Health, Winnipeg, MB, Canada.
    Hauff, Kristin
    Department of Pharmacology, University of Manitoba, Winnipeg, MB, Canada.
    Stelmack, Gerald L.
    Department of Physiology, University of Manitoba, Winnipeg, MB, Canada; Biology of Breathing Group, Manitoba Institute of Child Health, Winnipeg, MB, Canada.
    Schaafsma, Dedmer
    Department of Physiology, University of Manitoba, Winnipeg, MB, Canada; Biology of Breathing Group, Manitoba Institute of Child Health, Winnipeg, MB, Canada.
    Sharma, Pawan
    Department of Physiology, University of Manitoba, Winnipeg, MB, Canada; National Training Program in Allergy and Asthma, University of Manitoba, Winnipeg, MB, Canada; Biology of Breathing Group, Manitoba Institute of Child Health, Winnipeg, MB, Canada.
    McNeill, Karol D.
    Department of Physiology, University of Manitoba, Winnipeg, MB, Canada; Biology of Breathing Group, Manitoba Institute of Child Health, Winnipeg, MB, Canada.
    Hynes, Tyler S.
    Department of Physiology, University of Manitoba, Winnipeg, MB, Canada; National Training Program in Allergy and Asthma, University of Manitoba, Winnipeg, MB, Canada; Biology of Breathing Group, Manitoba Institute of Child Health, Winnipeg, MB, Canada.
    Kung, Sam K.
    Department of Immunology, University of Manitoba, Winnipeg, MB, Canada.
    Unruh, Helmut
    Department of Internal Medicine, University of Manitoba, Winnipeg, MB, Canada.
    Klonisch, Thomas
    Department of Human Anatomy and Cell Science, University of Manitoba, Winnipeg, MB, Canada.
    Hatch, Grant M.
    Department of Pharmacology, University of Manitoba, Winnipeg, MB, Canada.
    Los, Marek Jan
    Interfaculty Institute for Biochemistry, University of Tübingen, Germany.
    Halayko, Andrew J.
    Department of Physiology, University of Manitoba, Winnipeg, MB, Canada; National Training Program in Allergy and Asthma, University of Manitoba, Winnipeg, MB, Canada; Biology of Breathing Group, Manitoba Institute of Child Health, Winnipeg, MB, Canada; Department of Internal Medicine, University of Manitoba, Winnipeg, MB, Canada .
    Statin-triggered cell death in primary human lung mesenchyrnal cells involves p53-PUMA and release of Smac and Omi but not cytochrome c2010Ingår i: Biochimica et Biophysica Acta, ISSN 0006-3002, E-ISSN 1878-2434, Vol. 1803, nr 4, s. 452-467Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Statins inhibit 3-hydroxy-3-methyl-glutarylcoenzyme CoA (HMG-CoA) reductase, the proximal enzyme forcholesterol biosynthesis. They exhibit pleiotropic effects and are linked to health benefits for diseasesincluding cancer and lung disease. Understanding their mechanism of action could point to new therapies,thus we investigated the response of primary cultured human airway mesenchymal cells, which play aneffector role in asthma and chronic obstructive lung disease (COPD), to simvastatin exposure. Simvastatininduced apoptosis involving caspase-9, -3 and -7, but not caspase-8 in airway smooth muscle cells andfibroblasts. HMG-CoA inhibition did not alter cellular cholesterol content but did abrogate de novocholesterol synthesis. Pro-apoptotic effects were prevented by exogenous mevalonate, geranylgeranylpyrophosphate and farnesyl pyrophosphate, downstream products of HMG-CoA. Simvastatin increasedexpression of Bax, oligomerization of Bax and Bak, and expression of BH3-only p53-dependent genes, PUMAand NOXA. Inhibition of p53 and silencing of p53 unregulated modulator of apoptosis (PUMA) expressionpartly counteracted simvastatin-induced cell death, suggesting a role for p53-independent mechanisms.Simvastatin did not induce mitochondrial release of cytochrome c, but did promote release of inhibitor ofapoptosis (IAP) proteins, Smac and Omi. Simvastatin also inhibited mitochondrial fission with the loss ofmitochondrial Drp1, an essential component of mitochondrial fission machinery. Thus, simvastatin activatesnovel apoptosis pathways in lung mesenchymal cells involving p53, IAP inhibitor release, and disruption ofmitochondrial fission.

  • 43.
    Ghavami, Saeid
    et al.
    Department of Biochemistry and Medical Genetics, Manitoba Institute of Cell Biology, Cancer Care Manitoba, Winnipeg, Manitoba, Canada.
    Rashedi, Iran
    Manitoba Institute of Cell Biology and Department of Biochemistry and Medical Genetics, University of Manitoba, Winnipeg, Manitoba, Canada.
    Dattilo, Brian M.
    Departments of Biochemistry, Physics and Chemistry, Center for Structural Biology, Vanderbilt University, Nashville, Tennessee, USA;.
    Eshraghi, Mehdi
    Manitoba Institute of Cell Biology and Department of Biochemistry and Medical Genetics, University of Manitoba, Winnipeg, Manitoba, Canada.
    Chazin, Walter J.
    Departments of Biochemistry, Physics and Chemistry, Center for Structural Biology, Vanderbilt University, Nashville, Tennessee, USA;.
    HashemI, Mohammad
    Department of Clinical Biochemistry, School of Medicine, Zahedan University of Medical Science, Zahedan, Iran.
    Wesselborg, Sebastian
    Department of Internal Medicine I, University of Tübingen, Tübingen, Germany; and BioApplications Enterprises, Winnipeg, Manitoba, Canada.
    Kerkhoff, Claus
    Institute of Experimental Dermatology, Münster, Germany.
    Los, Marek Jan
    BioApplications Enterprises, Winnipeg, MB, Canada.
    S100A8/A9 at low concentration promotes tumor cell growth via RAGE ligation and MAP kinase-dependent pathway2008Ingår i: Journal of Leukocyte Biology, ISSN 0741-5400, E-ISSN 1938-3673, Vol. 83, nr 6, s. 1484-1492Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The complex formed by two members of the S100 calcium-binding protein family, S100A8/A9, exerts apoptosis-inducing activity against various cells, especially tumor cells. Here, we present evidence that S100A8/A9 also has cell growth-promoting activity at low concentrations. Receptor of advanced glycation end product (RAGE) gene silencing and cotreatment with a RAGE-specific blocking antibody revealed that this activity was mediated via RAGE ligation. To investigate the signaling pathways, MAPK phosphorylation and NF-κB activation were characterized in S100A8/A9-treated cells. S100A8/A9 caused a significant increase in p38 MAPK and p44/42 kinase phosphorylation, and the status of stress-activated protein kinase/JNK phosphorylation remained unchanged. Treatment of cells with S100A8/A9 also enhanced NF-κB activation. RAGE small interfering RNA pretreatment abrogated the S100A8/A9-induced NF-κB activation. Our data indicate that S100A8/A9-promoted cell growth occurs through RAGE signaling and activation of NF-κB.

  • 44.
    Glogowska, Aleksandra
    et al.
    Department of Human Anatomy and Cell Science, Winnipeg, Manitoba, Canada.
    Pyka, Janette
    Clinics of Surgery, Medical Faculty, Martin Luther University Halle-Wittenberg, Halle, Germany.
    Kehlen, Astrid
    Probiodrug AG, Weinbergweg, Halle, Germany.
    Los, Marek Jan
    BioApplications Enterprises, Winnipeg, MB, Canada.
    Perumal, Paul
    Department of Human Anatomy and Cell Science, Winnipeg, Manitoba, Canada.
    Weber, Ekkehard
    Cheng, Sheue-yann
    Laboratory of Molecular Biology, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264, USA.
    Hoang-Vu, Cuong
    Clinics of Surgery, Medical Faculty, Martin Luther University Halle-Wittenberg, Halle, Germany.
    Klonisch, Thomas
    Department of Human Anatomy and Cell Science; Department of Medical Microbiology and Infectious Diseases, Faculty of Medicine, University of Manitoba, Winnipeg, Canada.
    The cytoplasmic domain of proEGF negatively regulates motility and elastinolytic activity in thyroid carcinoma cells2008Ingår i: Neoplasia, ISSN 1522-8002, E-ISSN 1476-5586, Vol. 10, nr 10, s. 1120-1130Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The intracellular domains of the membrane-anchoring regions of some precursors of epidermal growth factor (EGF) family members have intrinsic biologic activities. We have determined the role of the human proEGF cytoplasmic domain (proEGFcyt) as part of the proEGF transmembrane-anchored region (proEGFctF) in the regulation of motility and elastinolytic invasion in human thyroid cancer cells. We found proEGFctF to act as a negative regulator of motility and elastin matrix penetration and the presence of proEGFcyt or proEGF22.23 resulted in a similar reduction in motility and elastinolytic migration. This activity was counteracted by EGF-induced activation of EGF receptor signaling. Decreased elastinolytic migratory activity in the presence of proEGFctF and proEGFcyt/proEGF22.23 coincided with decreased secretion of elastinolytic procathepsin L. The presence of proEGFctF and proEGFcyt/proEGF22.23 coincided with the specific transcriptional up-regulation of t-SNARE member SNAP25. Treatment with siRNA-SNAP25 resulted in motility and elastin migration being restored to normal levels. Epidermal growth factor treatment down-regulated SNAP25 protein by activating EGF receptor-mediated proteasomal degradation of SNAP25. These data provide first evidence for an important function of the cytoplasmic domain of the human proEGF transmembrane region as a novel suppressor of motility and cathepsin L-mediated elastinolytic invasion in human thyroid carcinoma cells and suggest important clinical implications for EGF-expressing tumors.

  • 45.
    Gomez-Carretero, Salvador
    et al.
    Dep. of Neuroscience, Karolinska Institutet, Stockholm, Sweden.
    Persson, Kristin M
    Linköpings universitet, Institutionen för teknik och naturvetenskap, Fysik och elektroteknik. Linköpings universitet, Tekniska högskolan.
    Libberton, Benjamin
    Dep. of Neuroscience, Karolinska Institutet, Stockholm, Sweden.
    Svennersten, Karl
    Dep. of Neuroscience, Karolinska Institutet, Stockholm, Sweden.
    Berggren, Magnus
    Linköpings universitet, Institutionen för teknik och naturvetenskap, Fysik och elektroteknik. Linköpings universitet, Tekniska högskolan.
    Rhen, Mikael
    Dep. of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden.
    Richter-Dahlfors, Agneta
    Dep. of Neuroscience, Karolinska Institutet, Stockholm, Sweden.
    Salmonella Biofilm Modulation with Electrically Conducting Polymers2014Manuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    Biofilms are ubiquitous in many human activities, constituting a threat or an advantage depending on the context of application. It is therefore of great interest to obtain new materials to study and control how biofilms are formed. Here, heparin and DBS (dodecylbenzenesulfonate) are incorporated as counter-ions to the PEDOT (poly(3,4-ethylenedioxythiophene)) backbone, forming conducting polymer thin-films. Polymer synthesis is based on electrodeposition, allowing for the adjustment, during fabrication, of properties like charge and hydrophobicity, important in bacterial adhesion. The electrochemical redox state of the polymer is of fundamental importance in Salmonella enterica Serovar Typhimurium biofilm modulation. Oxidized composites show increased levels of biofilm growth compared to reduced and pristine polymer films. As a result, biofilm formation is modulated by the application of a low electric voltage. Moreover, biofilm morphology and topology are affected by both the electrochemical redox state and the incorporated counter-ion, making these materials a useful tool in biofilm engineering.

  • 46.
    Gopinath, Madhumala
    et al.
    Department of Allied Health Sciences, Chettinad Hospital and Research Institute (CHRI), Chettinad Academy of Research and Education (CARE), Kelambakkam, Chennai-603103, India.
    Di Liddo, Rosa
    Department of Pharmacology and Pharmaceutical Sciences, University of Padova, Padova, Italy.
    Marotta, Francesco
    ReGenera RandD International for Aging Intervention, Milano-Beijing, Italy-China, VCC Preventive Medical Promotion Foundation, Beijing, China.
    Murugesan, Ramachandran
    Department of Allied Health Sciences, Chettinad Hospital and Research Institute (CHRI), Chettinad Academy of Research and Education (CARE), Kelambakkam, Chennai-603103, India.
    Banerjee, Antara
    Department of Allied Health Sciences, Chettinad Hospital and Research Institute (CHRI), Chettinad Academy of Research and Education (CARE), Kelambakkam, Chennai-603103, India.
    Sriramulu, Sushmitha
    Department of Allied Health Sciences, Chettinad Hospital and Research Institute (CHRI), Chettinad Academy of Research and Education (CARE), Kelambakkam, Chennai-603103, India.
    Jothimani, Ganesan
    Department of Allied Health Sciences, Chettinad Hospital and Research Institute (CHRI), Chettinad Academy of Research and Education (CARE), Kelambakkam, Chennai-603103, India.
    Subramaniam, Vimala Devi
    Department of Allied Health Sciences, Chettinad Hospital and Research Institute (CHRI), Chettinad Academy of Research and Education (CARE), Kelambakkam, Chennai-603103, India.
    Narasimhan, Srinivasan
    Department of Allied Health Sciences, Chettinad Hospital and Research Institute (CHRI), Chettinad Academy of Research and Education (CARE), Kelambakkam, Chennai-603103, India.
    Priya K, Swarna
    Department of Gynecology and Pediatrics, Chettinad Hospital and Research Institute (CHRI), Chettinad Academy of Research and Education (CARE), Kelambakkam, Chennai-603103, India.
    Sun, Xiao-Feng
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för Kirurgi, Ortopedi och Onkologi. Linköpings universitet, Medicinska fakulteten. Region Östergötland, Centrum för kirurgi, ortopedi och cancervård, Onkologiska kliniken US.
    Pathak, Surajit
    Department of Allied Health Sciences, Chettinad Hospital and Research Institute (CHRI), Chettinad Academy of Research and Education (CARE), Kelambakkam, Chennai-603103, India.
    Role of Hippo Pathway Effector Tafazzin Protein in Maintaining Stemness of Umbilical Cord-Derived Mesenchymal Stem Cells (UC-MSC)2018Ingår i: International Journal of Hematology-Oncology and Stem Cell Research, ISSN 2008-3009, E-ISSN 2008-2207, Vol. 12, nr 2, s. 153-165Artikel, forskningsöversikt (Refereegranskat)
    Abstract [en]

    Tafazzin (TAZ) protein has been upregulated in various types of human cancers, although the basis for elevation is uncertain, it has been made definite that the effect of mutation in the hippo pathway, particularly when it is switched off, considerably activates tafazzin transcriptionally and thus this results in tissue or tumor overgrowth. Recent perceptions into the activity of tafazzin, have ascribed to it, a role as stem cell factor in mouse mesenchymal and as well as in neural stem cells. Being a downstream molecule in Hippo signalling, phosphorylation or dephosphorylation of tafazzin gene regulates its transcriptional activity and the stemness of mesenchymal stem cells. Commonly, extracellular matrix controls the stem cell fate commitment and perhaps tafazzin controls stemness through altering the extra cellular matrix. Extracellular matrix is generally made up of prime proteoglycans and the fate stabilization of the resulting lineages is surveilled by engineering these glycans. Tafazzin degradation and addition of proteoglycans affect physical attributes of the extracellular matrix that drives cell differentiation into various lineages. Thus, tafazzin along with major glycans present in the extracellular matrix is involved in imparting stemness. However, there are incoherent molecular events, wherein both tafazzin and the extracellular matrix components, together either activate or inhibit differentiation of stem cells. This review discusses about the role of tafazzin oncoprotein as a stemness factor.

  • 47.
    Greuel, Selina
    et al.
    Charite Univ Med Berlin, Germany.
    Hanci, Guengoer
    Charite Univ Med Berlin, Germany.
    Boehme, Mike
    Charite Univ Med Berlin, Germany.
    Miki, Toshio
    Univ Southern Calif, CA USA.
    Schubert, Frank
    StemCell Syst GmbH, Germany.
    Sittinger, Michael
    Charite Univ Med Berlin, Germany.
    Mandenius, Carl-Fredrik
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Teknisk biologi. Linköpings universitet, Tekniska fakulteten.
    Zeilinger, Katrin
    Charite Univ Med Berlin, Germany.
    Freyer, Nora
    Charite Univ Med Berlin, Germany.
    Effect of inoculum density on human-induced pluripotent stem cell expansion in 3D bioreactors2019Ingår i: Cell Proliferation, ISSN 0960-7722, E-ISSN 1365-2184, Vol. 52, nr 4, artikel-id e12604Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Objective For optimized expansion of human-induced pluripotent stem cells (hiPSCs) with regards to clinical applications, we investigated the influence of the inoculum density on the expansion procedure in 3D hollow-fibre bioreactors. Materials and Methods Analytical-scale bioreactors with a cell compartment volume of 3 mL or a large-scale bioreactor with a cell compartment volume of 17 mL were used and inoculated with either 10 x 10(6) or 50 x 10(6) hiPSCs. Cells were cultured in bioreactors over 15 days; daily measurements of biochemical parameters were performed. At the end of the experiment, the CellTiter-Blue (R) Assay was used for culture activity evaluation and cell quantification. Also, cell compartment sections were removed for gene expression and immunohistochemistry analysis. Results The results revealed significantly higher values for cell metabolism, cell activity and cell yields when using the higher inoculation number, but also a more distinct differentiation. As large inoculation numbers require cost and time-extensive pre-expansion, low inoculation numbers may be used preferably for long-term expansion of hiPSCs. Expansion of hiPSCs in the large-scale bioreactor led to a successful production of 5.4 x 10(9) hiPSCs, thereby achieving sufficient cell amounts for clinical applications. Conclusions In conclusion, the results show a significant effect of the inoculum density on cell expansion, differentiation and production of hiPSCs, emphasizing the importance of the inoculum density for downstream applications of hiPSCs. Furthermore, the bioreactor technology was successfully applied for controlled and scalable production of hiPSCs for clinical use.

  • 48.
    Grote, Jens
    et al.
    Institute of Experimental Dermatology, University of Muenster, Germany; Interdisciplinary Center for Clinical Research, Core Group Integrated Functional Genomics, Medical Faculty, University of Muenster, Muenster, Germany .
    Koenig, Simone
    Interdisciplinary Center for Clinical Research, Core Group Integrated Functional Genomics, Medical Faculty, University of Muenster, Muenster, Germany .
    Ackermann, Doreen
    Interdisciplinary Center for Clinical Research, Core Group Integrated Functional Genomics, Medical Faculty, University of Muenster, Muenster, Germany .
    Sopalla, Claudia
    Institute of Experimental Dermatology, University of Muenster, Germany; Interdisciplinary Center for Clinical Research (IZKF) Münster, Germany .
    Benedyk, Malgorzata
    Institute of Experimental Dermatology, University of Muenster, Germany .
    Los, Marek Jan
    Manitoba Institute of Cell Biology, Cancer Care Manitoba; Manitoba Institute of Child Health; Department of Biochemistry and Medical Genetics; Department of Human Anatomy and Cell Science, University Manitoba, Winnipeg, Canada, .
    Kerkhoff, Claus
    Interdisciplinary Center for Clinical Research (IZKF) Münster, Germany; Institute of Experimental Dermatology, University of Muenster, Germany .
    Identification of poly(ADP-ribose) polymerase-1 and Ku70/Ku80 as transcriptional regulators of S100A9 gene expression2006Ingår i: BMC Molecular Biology, ISSN 1471-2199, E-ISSN 1471-2199, Vol. 7Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Background: S100 proteins, a multigenic family of non-ubiquitous cytoplasmic Ca2+-binding proteins, have been linked to human pathologies in recent years. Dysregulated expression of S100 proteins, including S100A9, has been reported in the epidermis as a response to stress and in association with neoplastic disorders. Recently, we characterized a regulatory element within the S100A9 promotor, referred to as MRE that drives the S100A9 gene expression in a cell type-specific, activation- and differentiation-dependent manner (Kerkhoff et al. ( 2002) J. Biol. Chem. 277, 41879-41887). Results: In the present study, we investigated transcription factors that bind to MRE. Using the MRE motif for a pull-down assay, poly(ADP-ribose) polymerase-I (PARP-I) and the heterodimeric complex Ku70/Ku80 were identified by mass spectrometry and confirmed by chromatin immunoprecipitation. Furthermore, TPA-induced S100A9 gene expression in HaCaT keratinocytes was blocked after the pharmacologic inhibition of PARP-l with 1,5- isoquinolinediol (DiQ). Conclusion: The candidates, poly(ADP-ribose) polymerase-l (PARP-l) and the heterodimeric complex Ku70/ Ku80, are known to participate in inflammatory disorders as well as tumorgenesis. The latter may indicate a possible link between S100 and inflammation-associated cancer.

  • 49.
    Gréen, Anna
    Linköpings universitet, Institutionen för klinisk och experimentell medicin. Linköpings universitet, Hälsouniversitetet.
    Histone H1: Subtypes and phosphorylation in cell life and death2009Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The genetic information of a human diploid cell is contained within approximately 2 metres of linear DNA. The DNA molecules are compacted and organized in various ways to fit inside the cell nucleus. Various kinds of histones are involved in this compaction. One of these histones, histone H1 is the topic of the present thesis. In addition to its structural role, H1 histones have been implicated in various processes, for example gene regulation and inhibition of chromatin replication.

    H1 histones, also termed linker histones, are relatively conserved proteins, and the various subtypes seem to have different and important functions even though redundancy between the subtypes has been demonstrated. Despite the sequence conservation of H1 subtypes, two sequence variations were detected within the H1.2 and H1.4 subtypes using hydrophilic interaction liquid chromatographic separation of H1 proteins from K562 and Raji cell lines in Paper I in the present thesis. The variations were confirmed by genetic analysis, and the H1.2 sequence variation was also found in genomic DNA of normal blood donors, in an allele frequency of 6.8%. The H1.4 sequence variation was concluded to be Raji specific. The significance of H1 microsequence variants is unclear, since the physiological function of H1 histones remains to be established.

    H1 histones can be phosphorylated at multiple sites. Changes in H1 phosphorylation has been detected in apoptosis, the cell cycle, gene regulation, mitotic chromatin condensation and malignant transformation. Contradictory data have been obtained on H1 phosphorylation in apoptosis, and many results indicate that H1 dephosphorylation occurs during apoptosis. We and others hypothesized that cell cycle effects by the apoptosis inducers may have affected previous studies. In Paper II, the H1 phosphorylation pattern was investigated in early apoptosis in Jurkat cells, taking cell cycle effects into account. In receptor-mediated apoptosis, apoptosis occurs with a mainly preserved phosphorylation pattern, while Camptothecin induced apoptosis results in rapid dephosphorylation of H1 subtypes, demonstrating that H1 dephosphorylation is not a general event in apoptosis, but may occur upon apoptosis induction via the mitochondrial pathway. The dephosphorylation may also be a result of early cell cycle effects or signalling.Therefore, the H1 phosphorylation pattern in the cell cycle of normal activated T cells was investigated in Paper IV in this thesis. Some studies, which have been made using cancer cell lines from various species and cell synchronization, have indicated a sequential addition of phosphate groupsacross the cell cycle. Normal T cells and cell sorting by flow cytometry were used to circumvent side-effects from cell synchronization. The data demonstrate that a pattern with phosphorylated serines is established in late G1/early S phase, with some additional phosphorylation occurring during S, and further up-phosphorylation seems to occur during mitosis. Malignant transformation may lead to an altered G1 H1 phosphorylation pattern, as was demonstrated using sorted Jurkat T lymphoblastoid cells.

    During mitosis, certain H1 subtypes may be relocated to the cytoplasm. In Paper III, the location of histones H1.2, H1.3 and H1.5 during mitosis was investigated. Histone H1.3 was detected in cell nuclei in all mitotic stages, while H1.2 was detected in the nucleus during prophase and telophase, and primarily in the cytoplasm during metaphase and early anaphase. H1.5 was located mostly to chromatin during prophase and telophase, and to both chromatin and cytoplasm during metaphase and anaphase. Phosphorylated H1 was located in chromatin in prophase, and in both chromatin and cytoplasm during metaphase, anaphase and telophase, indicating that the mechanism for a possible H1 subtype relocation to the cytoplasm is phosphorylation.

    In conclusion, data obtained during this thesis work suggest that H1 histones and their phosphorylation may participate in the regulation of events in the cell cycle, such as S-phase progression and mitosis, possibly through altered interactions with chromatin, and/or by partial or complete removal of subtypes or phosphorylated variants from chromatin.

    Delarbeten
    1. Characterization of sequence variations in human histone H1.2 and H1.4 subtypes
    Öppna denna publikation i ny flik eller fönster >>Characterization of sequence variations in human histone H1.2 and H1.4 subtypes
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    2005 (Engelska)Ingår i: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 272, nr 14, s. 3673 -3683Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    In humans, eight types of histone H1 exist (H1.1–H1.5, H1°, H1t and H1oo), all consisting of a highly conserved globular domain and less conserved N- and C-terminal tails. Although the precise functions of these isoforms are not yet understood, and H1 subtypes have been found to be dispensable for mammalian development, it is now clear that specific functions may be assigned to certain individual H1 subtypes. Moreover, microsequence variations within the isoforms, such as polymorphisms or mutations, may have biological significance because of the high degree of sequence conservation of these proteins. This study used a hydrophilic interaction liquid chromatographic method to detect sequence variants within the subtypes. Two deviations from wild-type H1 sequences were found. In K562 erythroleukemic cells, alanine at position 17 in H1.2 was replaced by valine, and, in Raji B lymphoblastoid cells, lysine at position 173 in H1.4 was replaced by arginine. We confirmed these findings by DNA sequencing of the corresponding gene segments. In K562 cells, a homozygous GCC→GTC shift was found at codon 18, giving rise to H1.2 Ala17Val because the initial methionine is removed in H1 histones. Raji cells showed a heterozygous AAA→AGA codon change at position 174 in H1.4, corresponding to the Lys173Arg substitution. The allele frequency of these sequence variants in a normal Swedish population was found to be 6.8% for the H1.2 GCC→GTC shift, indicating that this is a relatively frequent polymorphism. The AAA→AGA codon change in H1.4 was detected only in Raji cells and was not present in a normal population or in six other cell lines derived from individuals suffering from Burkitt's lymphoma. The significance of these sequence variants is unclear, but increasing evidence indicates that minor sequence variations in linker histones may change their binding characteristics, influence chromatin remodeling, and specifically affect important cellular functions.

    Ort, förlag, år, upplaga, sidor
    Wiley InterScience, 2005
    Nyckelord
    HILIC, linker histones, sequence variants, SNP, tumor cell lines
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-16381 (URN)10.1111/j.1742-4658.2005.04793.x (DOI)
    Tillgänglig från: 2009-01-20 Skapad: 2009-01-20 Senast uppdaterad: 2017-12-14Bibliografiskt granskad
    2. Histone H1 Dephosphorylation Is Not a General Feature in Early Apoptosis
    Öppna denna publikation i ny flik eller fönster >>Histone H1 Dephosphorylation Is Not a General Feature in Early Apoptosis
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    2008 (Engelska)Ingår i: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 47, s. 7539-7547Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Histone H1 is a family of nucleosomal proteins that exist in a number of subtypes. These subtypes can be modified after translation in various ways, above all by phosphorylation. Increasing levels of H1 phosphorylation has been correlated with cell cycle progression, while both phosphorylation and dephosphorylation of histone H1 have been linked to the apoptotic process. Such conflicting results may depend on which various apoptosis-inducing agents cause apoptosis via different apoptotic pathways and often interfere with cell proliferation. Therefore, we investigated the relation between apoptosis and H1 phosphorylation in Jurkat cells after apoptosis induction via both the extrinsic and intrinsic pathways and by taking cell cycle effects into account. After apoptosis induction by anti-Fas, no significant dephosphorylation, as measured by capillary electrophoresis, or cell cycle-specific effects were detected. In contrast, H1 subtypes were rapidly dephosphorylated when apoptosis was induced by camptothecin. We conclude that histone H1 dephosphorylation is not connected to apoptosis in general but may be coupled to apoptosis by the intrinsic pathway or to concomitant growth inhibitory signaling.

    Ort, förlag, år, upplaga, sidor
    ACS Publications, 2008
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-16382 (URN)10.1021/bi702311x (DOI)
    Tillgänglig från: 2009-01-20 Skapad: 2009-01-20 Senast uppdaterad: 2017-12-14Bibliografiskt granskad
    3. Translocation of Histone H1 Subtypes Between Chromatin and Cytoplasm During Mitosis in Normal Human Fibroblasts
    Öppna denna publikation i ny flik eller fönster >>Translocation of Histone H1 Subtypes Between Chromatin and Cytoplasm During Mitosis in Normal Human Fibroblasts
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    2010 (Engelska)Ingår i: Cytometry Part A, ISSN 1552-4922, E-ISSN 1552-4930, Vol. 77A, nr 5, s. 478-484Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Histone H1 is an important constituent of chromatin which undergoes major structural rearrangements during mitosis. However, the role of H1, multiple H1 subtypes and H1 phosphorylation is still unclear. In normal human fibroblasts, phosphorylated H1 was found located in nuclei during prophase and in both cytoplasm and condensed chromosomes during metaphase, anaphase and telophase as detected by immunocytochemistry. Moreover, we detected remarkable differences in the distribution of the histone H1 subtypes H1.2, H1.3 and H1.5 during mitosis. H1.2 was found in chromatin during prophase, and almost solely in the cytoplasm of metaphase and early anaphase cells. In late anaphase it appeared in both chromatin and cytoplasm, and again in chromatin during telophase. H1.5 distribution pattern resembled that of H1.2, but some H1.5 remained situated in chromatin during metaphase and early anaphase. H1.3 was detected in chromatin in all cell cycle phases. We propose therefore, that H1 subtype translocation during mitosis is controlled by phosphorylation, in combination with H1 subtype inherent affinity. We conclude that H1 subtypes, or their phosphorylated variants, may be signalling molecules in mitosis or that they leave chromatin in a regulated way to give access for chromatin condensing factors or transcriptional regulators during mitosis.

    Ort, förlag, år, upplaga, sidor
    John Wiley & Sons, 2010
    Nyckelord
    Histone H1, Chromatin, Cell cycle, Mitosis
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-16383 (URN)10.1002/cyto.a.20851 (DOI)000277174000009 ()20104577 (PubMedID)
    Tillgänglig från: 2009-01-20 Skapad: 2009-01-20 Senast uppdaterad: 2017-12-14Bibliografiskt granskad
    4. Histone H1 interphase phosphorylation pattern becomes largely established during G1/S transition in proliferating cells
    Öppna denna publikation i ny flik eller fönster >>Histone H1 interphase phosphorylation pattern becomes largely established during G1/S transition in proliferating cells
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    (Engelska)Manuskript (Övrigt vetenskapligt)
    Abstract [en]

    Histone H1 is an important constituent of chromatin, and is believed to be involved in regulation of chromatin structure. During the cell cycle, chromatin becomes locally decondensed in S phase, highly condensed during metaphase and again decondensed before re-entry into G1. This has been connected to increasing phosphorylation of H1 histones during the cell cycle. However, many of these experiments have been performed in non-human and human cancer   cell lines, and by the use of cell synchronization techniques and cell cycle-arresting drugs. In this study, we have investigated the H1 subtype composition and phosphorylation pattern in the cell cycle. Exponentially growing normal human activated T cells and Jurkat lymphoblastoid cells were sorted by fluorescence activated cell sorting into G1, S and G2/M populations, without the use of cell cycle arresting drugs. We found that the H1.5 protein level increased after T-cell activation. Our data indicate that serine phosphorylation of H1 subtypes occurred to a large extent in late G1 phase or early S, while some additional serine phosphorylation took place during S, G2 and M phases. Furthermore, our data suggest that the newly synthesized H1 molecules during S phase also achieve a similar phosphorylation pattern as the previous ones. Jurkat cells showed more extended H1.5 phosphorylation in G1 compared with T cells, a difference that can be explained by faster cell growth and/or the presence of enhanced H1 kinase activity in G1 in Jurkat cells. In conclusion, our data is consistent with a model where a major part of interphase H1 serine phosphorylation takes place within a narrow time window during the G1/Stransition. This implies that H1 serine phosphorylation may be coupled to changes in chromatin structure necessary for DNA replication.

    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:liu:diva-16384 (URN)
    Tillgänglig från: 2009-01-20 Skapad: 2009-01-20 Senast uppdaterad: 2010-01-14Bibliografiskt granskad
  • 50.
    Guan, Na N.
    et al.
    Department of Molecular Medicine and Surgery, Section of Urology, Karolinska Institutet, Stockholm, Sweden / Department of Urology, Karolinska University Hospital, Stockholm, Sweden / Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden.
    Sharma, Nimish
    Department of Molecular Medicine and Surgery, Section of Urology, Karolinska Institutet, Stockholm, Sweden / Department of Urology, Karolinska University Hospital, Stockholm, Sweden.
    Hallén‐Grufman, Katarina
    Department of Molecular Medicine and Surgery, Section of Urology, Karolinska Institutet, Stockholm, Sweden / Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden.
    Jager, Edwin W. H.
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Sensor- och aktuatorsystem. Linköpings universitet, Tekniska fakulteten.
    Svennersten, Karl
    Department of Molecular Medicine and Surgery, Section of Urology, Karolinska Institutet, Stockholm, Sweden / Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden.
    The role of ATP signalling in response to mechanical stimulation studied in T24 cells using new microphysiological tools2018Ingår i: Journal of Cellular and Molecular Medicine (Print), ISSN 1582-1838, E-ISSN 1582-4934, Vol. 22, nr 4, s. 2319-2328Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The capacity to store urine and initiate voiding is a valued characteristic of the human urinary bladder. To maintain this feature, it is necessary that the bladder can sense when it is full and when it is time to void. The bladder has a specialized epithelium called urothelium that is believed to be important for its sensory function. It has been suggested that autocrine ATP signalling contributes to this sensory function of the urothelium. There is well‐established evidence that ATP is released via vesicular exocytosis as well as by pannexin hemichannels upon mechanical stimulation. However, there are still many details that need elucidation and therefore there is a need for the development of new tools to further explore this fascinating field. In this work, we use new microphysiological systems to study mechanostimulation at a cellular level: a mechanostimulation microchip and a silicone‐based cell stretcher. Using these tools, we show that ATP is released upon cell stretching and that extracellular ATP contributes to a major part of Ca2+ signalling induced by stretching in T24 cells. These results contribute to the increasing body of evidence for ATP signalling as an important component for the sensory function of urothelial cells. This encourages the development of drugs targeting P2 receptors to relieve suffering from overactive bladder disorder and incontinence.

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