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  • 1. Bengtsson, T.
    et al.
    Jaconi, ME
    Gustafsson, Mikael
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Heart and Medicine Center, Department of Cardiology in Linköping.
    Magnusson, Karl-Eric
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Theler, JM
    Lew, DP
    Stendahl, Olle
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Actin dynamics in human neutrophils during adhesion and phagocytosis is controlled by changes in intracellular free calcium1993In: European Journal of Cell Biology, ISSN 0171-9335, E-ISSN 1618-1298, Vol. 62, no 1, p. 19-58Article in journal (Refereed)
    Abstract [en]

    The role of changes in cytosolic free calcium concentration ([Ca2+]i) in the assembly and disassembly of actin during adhesion and phagocytosis was evaluated. Rhodamine-phalloidin staining combined with quantitative fluorescence and confocal laser scanning microscopy was used to measure local F-actin changes in single adherent human neutrophils phagocytosing yeast particles on different surfaces and under different calcium conditions. Cells were suspended in a) calcium-containing medium (CCM) or b) calcium-free medium (CFM) or c) were first depleted of calcium (i.e., MAPT/AM-loaded in CFM) and then suspended in CFM (MAPT). In parallel, local [Ca2+]i changes were monitored using a fura-2 ratio imaging system. In CCM or CFM, attachment to the substrate and formation of pseudopods around a yeast particle generated, within a few seconds, rises in [Ca2+]i, both around the phagosome and in the cell body. During continued phagocytosis, [Ca2+]i was more elevated around the phagosome compared to the rest of the cell. No [Ca2+]i fluctuations were observed in MAPT cells. Adhesion and phagocytosis led to a several-fold increase in F-actin. The increase was transient in cells in CCM and CFM, but remained high in Ca-depleted neutrophils. A distinct ring of F-actin was formed around a phagosome with a yeast particle. Twenty min after ingestion the amount of this actin decreased more than 50% in CCM and CFM cells but increased by 40 to 100% in MAPT cells. The accumulation of F-actin in MAPT cells was reduced to resting levels by adding Ca2+ and ionomycin after ingestion. This treatment reestablished the periphagosomal [Ca2+]i rises, as observed in CCM cells. In conclusion, the present study shows that the actin polymerization, occurring in human neutrophils during adhesion and phagocytosis, is not influenced by changes in [Ca2+]i, whereas the subsequent depolymerization is. The accumulation of actin filaments around the phagosome in calcium-depleted cells could be involved in the inhibition of phagolysosome fusion seen in the absence of [Ca2+]i changes (Jaconi et al., J. Cell Biol. 110, 1555-1564 (1990)). This suggests that the actin network, controlled by [Ca2+]i, regulates the movement of granules during phagocytosis.

  • 2.
    Carlén, Anna
    et al.
    Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Clinical Physiology in Linköping. Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine.
    Åström Aneq, Meriam
    Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Clinical Physiology in Linköping. Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine.
    Nylander, Eva
    Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Clinical Physiology in Linköping. Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine.
    Gustafsson, Mikael
    Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping. Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine.
    Loaded treadmill walking and cycle ergometry to assess work capacity: a retrospective comparison in 424 firefighters.2017In: Clinical Physiology and Functional Imaging, ISSN 1475-0961, E-ISSN 1475-097X, Vol. 37, no 1, p. 37-44Article in journal (Refereed)
    Abstract [en]

    The fitness of firefighters is regularly evaluated using exercise tests. We aimed to compare, with respect to age and body composition, two test modalities for the assessment work capacity. A total of 424 Swedish firefighters with cycle ergometer (CE) and treadmill (TM) tests available from Jan 2004 to Dec 2010 were included. We compared results from CE (6 min at 200 W, 250 W or incremental ramp exercise) with TM (6 min at 8° inclination, 4·5 km h(-1) or faster, wearing 24-kg protective equipment). Oxygen requirements were estimated by prediction equations. It was more common to pass the TM test and fail the supposedly equivalent CE test (20%), than vice versa (0·5%), P<0·001. Low age and tall stature were significant predictors of passing both CE and TM tests (P<0·05), while low body mass predicted accomplishment of TM test only (P = 0·006). Firefighters who passed the TM but failed the supposedly equivalent CE test within 12 months had significantly lower body mass, lower BMI, lower BSA and shorter stature than did those who passed both tests. Calculated oxygen uptake was higher in TM tests compared with corresponding CE tests (P<0·001). Body constitution affected approval differently depending on the test modality. A higher approval rate in TM testing suggests lower cardiorespiratory requirements compared with CE testing, even though estimated oxygen uptake was higher during TM testing. The relevance of our findings in relation to the occupational demands needs reconsidering.

  • 3.
    de Geer, Lina
    et al.
    Linköping University, Department of Medical and Health Sciences, Division of Drug Research. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Anaesthetics, Operations and Specialty Surgery Center, Department of Anaesthesiology and Intensive Care in Linköping.
    Oscarsson, Anna
    Linköping University, Department of Medical and Health Sciences, Division of Drug Research. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Anaesthetics, Operations and Specialty Surgery Center, Department of Anaesthesiology and Intensive Care in Linköping.
    Gustafsson, Mikael
    Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping. Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine.
    Lung ultrasound in quantifying lung water in septic shock patients2015In: Critical Care, ISSN 1364-8535, E-ISSN 1466-609X, Vol. 19, no 1, p. 140-Article in journal (Refereed)
    Abstract [en]

    Quantification of lung ultrasound (LUS) artifacts (B-lines) is used to assess pulmonary congestion in emergency medicine and cardiology [1,2]. We investigated B-lines in relation to extravascular lung-water index (EVLWI) from invasive transpulmonary thermodilution in septic shock patients. Our aim was to evaluate the role of LUS in an intensive care setting.

  • 4.
    Fernlund, Eva
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Division of Children's and Women's health. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Center of Paediatrics and Gynaecology and Obstetrics, H.K.H. Kronprinsessan Victorias barn- och ungdomssjukhus. Lund University, Sweden.
    Wålinder Österberg, Anna
    Linköping University, Department of Clinical and Experimental Medicine, Division of Children's and Women's health. Linköping University, Faculty of Medicine and Health Sciences.
    Kuchinskaya, Ekaterina
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
    Gustafsson, Mikael
    Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping.
    Jansson, Kjell
    Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping.
    Gunnarsson, Cecilia
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Center for Diagnostics, Clinical genetics. Region Östergötland, Center for Business support and Development.
    Novel Genetic Variants in BAG3 and TNNT2 in a Swedish Family with a History of Dilated Cardiomyopathy and Sudden Cardiac Death2017In: Pediatric Cardiology, ISSN 0172-0643, E-ISSN 1432-1971, Vol. 38, no 6, p. 1262-1268Article in journal (Refereed)
    Abstract [en]

    Familial dilated cardiomyopathy is a rare cause of dilated cardiomyopathy (DCM), especially in childhood. Our aim was to describe the clinical course and the genetic variants in a family where the proband was a four-month-old infant presenting with respiratory problems due to DCM. In the family, there was a strong family history of DCM and sudden cardiac death in four generations. DNA was analyzed initially from the deceased girl using next-generation sequencing including 50 genes involved in cardiomyopathy. A cascade family screening was performed in the family after identification of the TNNT2 and the BAG3 variants in the proband. The first-degree relatives underwent clinical examination including biochemistry panel, cardiac ultrasound, Holter ECG, exercise stress test, and targeted genetic testing. The index patient presented with advanced DCM. After a severe clinical course, the baby had external left ventricular assist as a bridge to heart transplantation. 1.5 months after transplantation, the baby suffered sudden cardiac death (SCD) despite maximal treatment in the pediatric intensive care unit. The patient was shown to carry two heterozygous genetic variants in the TNNT2 gene [TNNT2 c.518G amp;gt; A(p.Arg173Gln)] and BAG3 [BAG3 c.785C amp;gt; T(p.Ala262Val)]. Two of the screened individuals (two females) appeared to carry both the familial variants. All the individuals carrying the TNNT2 variant presented with DCM, the two adult patients had mild or moderate symptoms of heart failure and reported palpitations but no syncope or presyncopal attacks prior to the genetic diagnosis. The female carriers of TNNT2 and BAG3 variants had more advanced DCM. In the family history, there were three additional cases of SCD due to DCM, diagnosed by autopsy, but no genetic analysis was possible in these cases. Our findings suggest that the variants in TNNT2 and BAG3 are associated with a high propensity to life-threatening cardiomyopathy presenting from childhood and young adulthood.

  • 5.
    Gustafsson, Mikael
    Linköping University, Department of Medical and Health Sciences. Linköping University, Faculty of Health Sciences.
    Arbetsprov hos rök- och kemdykare2013In: Kliniska arbetsprov: metoder för diagnos och prognos / [ed] Lennart Jorfeldt, Olle Pahlm, Lund, Sverige: Studentlitteratur, 2013, 1, p. 433-438Chapter in book (Other academic)
    Abstract [sv]

    Kliniska arbetsprov – metoder för diagnos och prognos är en vidareutveckling av en serie kompendier och böcker om arbetsprov med första utgåvan 1987. Titeländringen från Det kliniska arbetsprovet till den nya markerar att fältet breddats. Ursprungligen var arbetsprovet inriktat på bedömning av fysisk arbetsförmåga och på hjärtischemidiagnostik med EKG. Under senare år har i allt större utsträckning tillkommit nya diagnostiska modaliteter, inte som ­ersättning för EKG utan som komplement. Exempel är myokardskintigrafi, lungmekanik, analys av andnings- och blodgaser, ekokardiografi, dator­tomografi och magnetresonanstomografi. Dessa modaliteter beskrivs i detalj 
i boken och deras roll i patientutredningar värderas och diskuteras.

    Boken vänder sig till alla som i sin vardagsgärning utför och bedömer arbetsprov, remitterar patienter till arbetsprov eller tar emot och utnyttjar undersökningsresultat; således ett brett spektrum av sjukvårdspersonal. Boken fyller också ett centralt behov i olika grundutbildningar, i första hand för BMA med inriktning mot klinisk fysiologi, sjukgymnaster och läkare.

    Kliniska arbetsprov – metoder för diagnos och prognos är en lärobok och ett ­referensverk. Som lärobok kan den läsas som en ”läxbok” från pärm till pärm, men den är även anpassad för dem som enligt moderna pedagogiska ­principer vill fördjupa sina kunskaper kring ett problem och tillgodogöra sig ett begränsat avsnitt.

  • 6.
    Gustafsson, Mikael
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Motility of human polymorphonuclear leukocytes: An image processing approach1993Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Cell motility is essential for polymorphonuclear leukocyte (PMN) function in the defense against invading microorganisms and in inflammatory processes. Using digital video microscopy and image processing, individual cells were studied during random locomotion, chemotactic locomotion and phagocytosis. When PMN moved in a specific direction, the lipid membrane was found to flow in the same direction, contradicting the rearward lipid flow model of cell locomotion. This was assessed with video photobleaching of a fluorescent lipid probe, Dii. The motility of PMN could not be blocked by ca2+ depletionand /or inhibition of myosin light chain kinase, suggesting that ea2+ and myosin are not necessary for locomotion. Temporal and spatial characteristics of intracellular free ea2+ and intracellular pH were assessed by loading the cells with esters of Fura-2 and the fluorcscein derivative BCECF, respectively. There was a distinct correlation between the direction of PMN locomotion and the slope of the Ca2+ gradient of the cells. PMN moving randomly on a surface often exhibited a rearward gradient with higher ca2+ concentration in the rear of the cell. Experimental reversal of the gradient, however, did not change the direction of motility. During phagocytosis of yeast particles, Saccharomyces cerevisiae, intracellular free calcium rose within seconds after contact with the prey. Intracellular pH varied between 7.1 and 7.3 and was uniform across the cells. After phagocytosis, phagosomal pH first decreased and then returned to neutraL A role for pH in phagosome-lysosome formation is proposed. PMN loaded with self-quenching concentrations of BCECF exhibited a strict correlation between pseudopod protrusions and increase in fluorescence, indicating water influx and dilution of the probe. This finding may be essential for the understanding of actin cytoskeleton dynamics during cell locomotion.

  • 7.
    Gustafsson, Mikael
    et al.
    Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping. Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine.
    Alehagen, Urban
    Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping. Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine. Linköping University, Faculty of Medicine and Health Sciences.
    Johansson, Peter
    Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping. Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine. Linköping University, Faculty of Medicine and Health Sciences.
    Imaging congestion with a pocket ultrasound device - prognostic implications in patients with chronic heart failure.2015In: Journal of Cardiac Failure, ISSN 1071-9164, E-ISSN 1532-8414, Vol. 21, no 7, p. 548-554Article in journal (Refereed)
    Abstract [en]

    AIMS: Venous congestion is common in patients with chronic heart failure (HF). We used a pocket-sized ultrasound imaging device (PID) to assess the patient's congestive status and related our findings to prognosis.

    METHODS AND RESULTS: 104 consecutive outpatients from an HF outpatient clinic were studied. Interstitial lung water (ILW), pleural effusion (PE) and the diameter of the vena cava inferior (VCI) were assessed using a PID. ILW was assessed by demonstration of B-lines (comet tail artefact (CTA). Out of the 104 patients, 28 had CTA, and eight had PE. Median VCI diameter was 18 mm, ±14/22 mm (quartiles). Each of these parameters correlated weakly (r= 0.26-0.37, p< 0.05) with the HF biomarker NT-proBNP. During the median follow-up time of 530 days, 14 hospitalizations deaths and 7 deaths were registered. Findings of CTA, PE or a composite of both, increased the risk of death or hospitalization (hazard ratio 3-4, p< 0.05). After adjustment for age, cardiac systolic function and NT-proBNP, this difference remained significant for CTA alone and CTA + PE combined, but not for PE alone.

    CONCLUSION: By using a handheld ultrasound device, signs of pulmonary congestion could be demonstrated. When found, these had a significant prognostic impact in clinically stable HF.

  • 8.
    Gustafsson, Mikael
    et al.
    Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Heart and Medicine Center, Department of Cardiology in Linköping.
    Alehagen, Urban
    Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Heart and Medicine Center, Department of Cardiology in Linköping.
    Johansson, Peter
    Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Heart and Medicine Center, Department of Cardiology in Linköping.
    Pocket-sized ultrasound examination of fluid imbalance in patients with heart failure: A pilot and feasibility study of heart failure nurses without prior experience of ultrasonography.2015In: European journal of cardiovascular nursing : journal of the Working Group on Cardiovascular Nursing of the European Society of Cardiology, ISSN 1873-1953, Vol. 14, no 4, p. 294-302Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Detecting fluid imbalance in patients with chronic heart failure can be challenging. Use of a pocket-sized ultrasound device (PSUD) in addition to physical examination can be helpful to assess this important information.

    AIM: To evaluate the feasibility for nurses without prior experience of ultrasonography to examine fluid imbalance by the use of a PSUD on heart failure patients.

    METHOD: Four heart failure nurses and an expert cardiologist participated. The nurses underwent a four-hour PSUD training programme. One hundred and four heart failure outpatients were included. The examinations obtained information of pulmonary congestion, pleural effusion and the diameter of the vena cava inferior.

    RESULTS: Examinations took nine minutes on average. In 28% and 14% of the patients, pulmonary congestion and pleural effusion respectively were found by the nurses. The sensitivities and specificities for nurses' findings were 79% and 91%, and, 88% and 93% respectively. The inter-operator agreement between the nurses and the cardiologist reached a substantial level (kappa values: 0.71 and 0.66). The inter-operator agreement for vena cava inferior reached a fair level (kappa value=0.39). Bland-Altman plots of the level of agreement revealed a mean difference of vena cava inferior diameter of 0.11 cm, while the 95% lower and upper limits ranged from -0.78 cm to 1.00 cm.

    CONCLUSION: After brief training, heart failure nurses can reliably identify pulmonary congestion and pleural effusion with a PSUD. Assessment of vena cava inferior was less valid. PSUD readings, when added to the history and a physical examination, can improve nurse assessment of fluid status in patients with heart failure.

  • 9.
    Gustafsson, Mikael
    et al.
    Linköping University, Department of Medical and Health Sciences.
    Landtblom, Anne-Marie
    Linköping University, Department of Clinical and Experimental Medicine, Division of Neuro and Inflammation Science. Linköping University, Faculty of Health Sciences. Linköping University, Center for Medical Image Science and Visualization (CMIV). Östergötlands Läns Landsting, Local Health Care Services in Central Östergötland, Department of Neurology.
    Lundberg, Peter
    Linköping University, Department of Medical and Health Sciences, Division of Radiological Sciences. Linköping University, Faculty of Health Sciences. Linköping University, Center for Medical Image Science and Visualization (CMIV). Östergötlands Läns Landsting, Center for Surgery, Orthopaedics and Cancer Treatment, Department of Radiation Physics.
    Absolut kvantifiering av metaboliter i hjärnans vita substans hos patienter med MS och normal magnetkamerundersökning2000Conference paper (Other academic)
  • 10.
    Gustafsson, Mikael
    et al.
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Magnusson, Karl-Eric
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    A distributed image-processing system for measurements of intracellular calcium in living cells1991In: Computer Methods and Programs in Biomedicine, ISSN 0169-2607, E-ISSN 1872-7565, Vol. 36, no 4, p. 199-221Article in journal (Refereed)
    Abstract [en]

    During the last decade, image-processing techniques have been introduced as a valuable tool in biologically oriented research. In combination with novel fluorescent probes, these techniques permit assessment of subcellular distributions of several intracellularly important cations, such as free calcium ions and protons. Typically, systems used for image processing are located centrally around the experimental setup. This configuration has drawbacks, mainly because the laborious extraction and processing of data that generally follow an experimental session limits the access to the system for other investigators. We describe here the principles of a distributed image processing system, based on IBM-compatible personal computers (PCs), that without extra hardware can cope with all the necessary image processing involved in imaging of intracellular cations. The potential of the PC as an image processor, however, reaches beyond this specific application and many image processing tasks can be carried out successfully on a standard PC. Thus, the centrally located dedicated image processor is used only for image acquisition in the experimental situation. This in turn optimizes the utilization of expensive resources and increases efficiency. The mouse-operated software is described in detail, so that interested investigators can extract useful parts for integration into their own applications and experimental environment.

  • 11.
    Gustafsson, Mikael
    et al.
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Magnusson, Karl-Eric
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    A novel principle for quantitation of fast intracellular calcium changes using Fura-2 and a modified image processing system: applications in studies of neutrophil motility and phagocytosis1992In: Cell Calcium, ISSN 0143-4160, E-ISSN 1532-1991, Vol. 13, no 8, p. 473-486Article in journal (Refereed)
    Abstract [en]

    A new principle is described for imaging intracellular free calcium [Ca2+]i changes in single, living cells utilizing the fluorescent probe Fura-2. It is based upon video color mixing in real time and allows high-speed visualization, at maximum image resolution, of [Ca2+]i changes without digital image ratioing. The epifluorescence images produced by 340 and 380 nm excitations are stored in two memory buffers of a personal computer-based image processing system. Two video signals are generated independently from each buffer and connected to the red and green inputs of a video display. An image is this way created, in which [Ca2+]i shows up as a specific hue, whereas changes in dye concentration, light intensity, cell thickness show up as variations in brightness of the imaged cells. The method has advantages over conventional ratio imaging, notably simplicity and speed, since no calculations are made. Yet it can be combined with traditional digital image processing. The imaging technique allows monitoring of [Ca2+]i changes in rapidly moving cells, like neutrophils. It is demonstrated that during random locomotion on serum-coated glass surfaces, [Ca2+]i levels appeared to oscillate and that the frequency of the oscillations are related to locomotive activity. Furthermore, in Ca2+ free medium, the cells continue to move and phagocytose in the presence of Ca2+ ionophore (ionomycin) and 2 mM EGTA. In the presence of 1 mM extracellular Ca2+, ionomycin-treated cells were not able to move or phagocytose.

  • 12.
    Gustafsson, Mikael
    et al.
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Heart and Medicine Center, Department of Cardiology in Linköping.
    Sundqvist, Tommy
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Magnusson, Karl-Eric
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Lateral diffusion of the secretory component (SC) in the basolateral membrane of the human colon carcinoma cell line HT29 assessed with fluorescence recovery after photobleaching1988In: Journal of Cellular Physiology, ISSN 0021-9541, E-ISSN 1097-4652, Vol. 137, no 3, p. 608-611Article in journal (Refereed)
    Abstract [en]

    The lateral diffusion of the secretory component (SC), acting as a receptor for dimeric IgA in the basolateral side of intestinal epithelial cells, was studied in the human colonic carcinoma cell line HT29. The HT29 cells were grown in Dulbecco's modified Eagle's medium in which galactose had been substituted for glucose to promote development of small intestine-like cells, with a distinct separation of the basolateral side from the apical surface. The SC was stained with rhodamine-labeled polyclonal anti-human SC rabbit antibodies (Ig) or Fab fragments, and the lateral mobility was assessed with the fluorescence recovery after photobleaching technique. The average lateral diffusion was consistent with a diffusion constant of 7.7 ± 2.0 (mean value ± SD; n = 29) and 7.1 ± 2.3 (n = 30) × 10−10 cm2s−1 for Ig-and Fab-labeled receptors, respectively, which is slower than lipid diffusion but is similar to that found for other membrane receptors. The corresponding values for the fraction of mobile receptors were 66 ± 13% and 71 ± 12%, respectively. Cells were labeled from the top of the culture plate, and cells adjacent to a mechanically made rift or a natural opening in the cell monolayer were labeled more strongly, confirming the microscope-based impression that the basolateral surface primarily harboured the SC receptor.

  • 13.
    Jacobson, Ken
    et al.
    Department of Cell Biology and Anatomy, University of North Carolina, Chapel Hill, USA.
    Lee, Juliet
    Department of Cell Biology and Anatomy, University of North Carolina, Chapel Hill, USA.
    Gustafsson, Mikael
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Magnusson, Karl-Eric
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Lipid flow in locomoting cells: Response1991In: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 251, no 4991, p. 318-318Article in journal (Refereed)
    Abstract [en]

    n/a

  • 14.
    Lee, Juliet
    et al.
    Department of Cell Biology and Anatomy, University of North Carolina, Chapel Hill, USA.
    Gustafsson, Mikael
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Magnusson, Karl-Eric
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Jacobson, Ken
    Department of Cell Biology and Anatomy, University of North Carolina, Chapel Hill, USA.
    The direction of membrane lipid flow in locomoting polymorphonuclear leukocytes1990In: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 247, no 4947, p. 1229-1233Article in journal (Refereed)
    Abstract [en]

    The objective of this study was to determine the direction of membrane lipid flow in locomoting cells. The plasma membrane of human polymorphonuclear leukocytes was stained with a fluorescent lipid analog dihexadecanoyl indocarbocyanine. A line was photobleached on the cell surface perpendicular to the direction of cell motion. Low-light-level fluorescence microscopy and digital image-processing techniques were used to analyze a series of images taken at short intervals after photobleaching. The bleached line remained visible for about 5 seconds before being erased by diffusional recovery. Examination of fluorescence intensity profiles allowed a comparison to be made between the velocities of line and cell movement. Results indicate that the bleached line moves forward with the same velocity as the cell during locomotion, refuting the retrograde lipid flow model of locomotion. Instead, the plasma membrane lipid appears to move forward according to either the unit movement of membrane or the tank track model of locomotion.

  • 15.
    Loitto, Vesa-Matti
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Forslund, Tony
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Sundqvist, Tommy
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Magnusson, Karl-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Gustafsson, Mikael
    Linköping University, Department of Medicine and Care. Linköping University, Faculty of Health Sciences.
    Neutrophil leukocyte motility requires directed water influx2002In: Journal of Leukocyte Biology, ISSN 0741-5400, E-ISSN 1938-3673, Vol. 71, no 2, p. 212-222Article in journal (Refereed)
    Abstract [en]

    The ability of neutrophils to sense and move to sites of infection is essential for our defense against pathogens. For motility, lamellipodium extension and stabilization are prerequisites, but how cells form such membrane protrusions is still obscure. Using contrast-enhanced video microscopy and Transwell® assays, we show that water-selective aquaporin channels regulate lamellipodium formation and neutrophil motility. Addition of anti-aquaporin-9 antibodies, HgCl2, or tetraethyl ammonium inhibited the function(s) of the channels and blocked motility-related shape changes. On human neutrophils, aquaporin-9 preferentially localized to the cell edges, where N-formyl peptide receptors also accumulated, as assessed with fluorescence microscopy. To directly visualize water fluxes at cell edges, cells were loaded with high dilution-sensitive, self-quenching concentrations of fluorophore. In these cells, motile regions always displayed increased fluorescence compared with perinuclear regions. Our observations provide the first experimental support for motility models where water fluxes play a pivotal role in cell-volume increases accompanying membrane extensions.

  • 16.
    Lundqvist, Helen
    et al.
    Phagocyte Research Laboratory, Department of Medical Microbiology and Immunology, University of Göteborg, Göteborg, Sweden.
    Gustafsson, Mikael
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Johansson, Agneta
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Särndahl, Eva
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Dahlgren, Claes
    Phagocyte Research Laboratory, Department of Medical Microbiology and Immunology, University of Göteborg, Göteborg, Sweden.
    Neutrophil control of formylmethionyl-leucyl-phenylalanine induced mobilization of secretory vesicles and NADPH-oxidase activation: Effect of an association of the ligand-receptor complex to the cytoskeleton1994In: Biochimica et Biophysica Acta. Molecular Cell Research, ISSN 0167-4889, E-ISSN 1879-2596, Vol. 1224, no 1, p. 43-50Article in journal (Refereed)
    Abstract [en]

    The stimulus formylmethionyl-leucyl-phenylalanine (FMLP) interacts with neutrophils and generates signal(s) in the cells that induces mobilization of the secretory vesicles as well as activation of the superoxide anion/hydrogen peroxide generating NADPH-oxidase. Binding, at 15°C, of FMLP to its neutrophil surface receptor is followed by an association of the ligand-receptor complex to the cell cytoskeleton, and this association occurs concomitant with a desensitization of the cells with respect to activation of the NADPH-oxidase. Other stimuli can still activate the oxidase (in fact even induce a primed response), indicating that the observed phenomenon is stimulus specific and could not be accounted for by an effect on the oxidase itself, but rather that the association of the ligand-receptor complex to the cytoskeleton eliminates the capacity of the complex to generate the signal(s) that activates the NADPH-oxidase. The cytoskeleton associated ligand-receptor complex generates, however, the signal(s) responsible for mobilization of the secretory vesicles, to the plasma membrane, and this mobilization occurs without any increase in the intracellular concentration of free Ca2+.

  • 17.
    Magnusson, Karl-Eric
    et al.
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Gustafsson, Mikael
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Heart and Medicine Center, Department of Cardiology in Linköping.
    Holmgren, Kajsa
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Johansson, Birgitta
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Small intestinal differentiation in human colon carcinoma HT29 cells has distinct effects on the lateral diffusion of lipids (ganglioside GM1) and proteins (HLA Class 1, HLA Class 2, and neoplastic epithelial antigens) in the apical cell membrane1990In: Journal of Cellular Physiology, ISSN 0021-9541, E-ISSN 1097-4652, Vol. 143, no 2, p. 381-390Article in journal (Refereed)
    Abstract [en]

    We have studied the effect of maturation to small intestinal-like epithelial cells of the human colonic calcinoma cell line HT29 on the lateral mobility of different representative membrane components (lipid, proteins), as assessed with fluorescence recovery after photobleaching (FRAP). Maturation was induced in vitro in the HT29 cells by replacing glucose (Glu) with galactose (Gal) in the growth medium (DMEM) during a 21-day period. Scanning electron microscopy revealed an increased number of microvilli in the apical cell membrane, and enzyme analyses (alkaline phosphatase, aminopeptidase) in combination with aqueous countercurrent distribution, indicated that maturation was induced with DMEM-Gal. In comparison to control cells grown in DMEM-Glu medium, the more small intestinal-like cells grown in DMEM-Gal displayed no alteration of the lateral mobility of either cholera toxin (B subuni)-labelled ganglioside GM1 (diffusion coefficient, D [x 108] = 0.8–0.9 cm2s−1; mobile fraction, R = 50-60%) or antibody-stained Class 2 histocompatibility (HLA-DR) antigen (D [x 109] = 2 cm2s−1; R = 60–70%). However, antibody-labelled β2-microglobulin of HLA Class 1 antigen displayed increased mobility in HT29-Gal cells; D was × 1.4 and R × 1.8 larger in the HT29-Gal cells. By contrast, the mobility of a neoplastic antigen was reduced; D and R were × 0.60 and × 0.69 of the values seen in HT29-Glu cells. It is thus concluded that DMEM-Gal-induced differentiation in confluent HT29 cells is accompanied by specific rather than general effects on the lateral mobil-ity of different membrane components.

  • 18.
    Majeed, Meytham
    et al.
    Linköping University, Department of Molecular and Clinical Medicine, Clinical Microbiology. Linköping University, Faculty of Health Sciences.
    Gustafsson, Mikael
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Kihlström, Erik
    Department of clinical microbiology, Örebro medical center hospital, Örebro, Sweden.
    Stendahl, Olle
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Roles of Ca2+ and F-actin in intracellular aggregation of Chlamydia trachomatis in eucaryotic cells1993In: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 61, no 4, p. 1406-1414Article in journal (Refereed)
    Abstract [en]

    The effect of intracellular free Ca2+ ([Ca2+]i) on the intracellular aggregation of Chlamydia trachomatis serovars L2 and E in McCoy and HeLa cells is investigated. Loading the cells with the Ca2+ chelator MAPT/AM (1,2-bis-5-methyl-amino-phenoxylethane-N,N-n'-tetra-acetoxymethyl acetate), thereby decreasing the [Ca2+]i from 67 to 19 nM, decreased the number of cells with a local aggregation of chlamydiae in a dose-dependent manner. Neither the attachment nor the uptake of elementary bodies (EBs) was, however, affected after depletion of Ca2+ from the cells. There was no significant difference in the level of measured [Ca2+]i between infected and uninfected cells. Reducing the [Ca2+]i also significantly inhibited chlamydial inclusion formation. Differences in the organization of the actin filament network were observed in response to [Ca2+]i depletion. In Ca(2+)-depleted cells, where few EB aggregates were formed, few local accumulations of F-actin were observed in the cytosol. These results suggest that the aggregation of EBs in eucaryotic cells requires a normal homeostasis of intracellular Ca2+. By affecting F-actin reorganization and putatively certain Ca(2+)-binding proteins, [Ca2+]i plays a vital role in the infectious process of chlamydiae.

  • 19.
    Mourad, Ghassan
    et al.
    Linköping University, Department of Social and Welfare Studies, Division of Nursing Science. Linköping University, Faculty of Medicine and Health Sciences.
    Strömberg, Anna
    Linköping University, Department of Medical and Health Sciences, Division of Nursing Science. Linköping University, Faculty of Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping.
    Jonsbu, E
    More and Romsdal Hospital Trust, Department of Psychiatry, Molde, Norway.
    Gustafsson, Mikael
    Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping.
    Johansson, Peter
    Linköping University, Department of Medical and Health Sciences, Division of Nursing Science. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping.
    Jaarsma, Tiny
    Linköping University, Department of Social and Welfare Studies, Division of Nursing Science. Linköping University, Faculty of Medicine and Health Sciences.
    Guided Internet-delivered cognitive behavioural therapy in patients with non-cardiac chest pain: -a pilot randomized controlled study2016In: Abstract book, Sage Publications, 2016, Vol. 15 (S1), p. S44-S44, article id P116Conference paper (Other academic)
    Abstract [en]

    Background: Patients with recurrent episodes of non-cardiac chest pain (NCCP) suffer from cardiac anxiety as they misinterpret the pain as being cardiac-related and avoid physical activity that they think could threaten their life. This could lead to increased healthcare utilization and costs. These patients might benefit from help and support to evaluate the perception and management of their chest pain. 

    Objective: To test the feasibility of a short guided Internet-delivered cognitive behavioural intervention and the effects on chest pain, cardiac anxiety, fear of body sensations and depressive symptoms in patients with NCCP compared to usual care.

    Methods: A pilot randomized controlled study was conducted. Fifteen patients, 9 men and 6 women between the age of 22 and 76 (median age of 66 years, q1-q3 57-73) were randomly assigned to either intervention (n=7) or control (n=8) group. Patients had recurrent NCCP and suffered from cardiac anxiety or fear of body sensations. The intervention consisted of a 4-session guided Internet-delivered cognitive behavioural therapy program containing psychoeducation, exposure to physical activity, and relaxation. The control group received usual care. All patients completed a web-based questionnaire on socio-demographic variables, chest pain frequency, cardiac anxiety, fear of body sensations, and depressive symptoms. 

    Results: Five out of the 7 patients in the intervention group completed all sessions as planned and two completed only parts of the program. The program was perceived as user-friendly with comprehensible language, adequate and varied content, and manageable homework assignments. The patients were engaged in the program for about 45-60 minutes per day and about 22 minutes’ therapist time was required to guide, support and give feedback to each patient throughout the program. Participating in the program empowered and motivated many of the patients to be active and complete the program. In general, patients in both the intervention and control groups improved with regard to chest pain frequency, cardiac anxiety, fear of body sensations, and depressive symptoms, but there were no significant differences between the groups. 

    Conclusions: A short guided Internet-delivered cognitive behavioural therapy program was feasible. Patients in both the intervention and control groups improved with regard to chest pain frequency, cardiac anxiety, fear of body sensations, and depressive symptoms, but no significant differences were found between the groups. Patients should be followed-up for longer periods to measure the long-term effects of the intervention.

  • 20.
    Mourad, Ghassan
    et al.
    Linköping University, Department of Social and Welfare Studies, Division of Health, Activity and Care. Linköping University, Faculty of Medicine and Health Sciences.
    Strömberg, Anna
    Linköping University, Department of Medical and Health Sciences, Division of Nursing Science. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping.
    Jonsbu, Egil
    Department of Psychiatry, More and Romsdal Hospital Trust, Molde, and Department of Neuroscience, Norwegian University of Science and Technology, Trondheim, Norway.
    Gustafsson, Mikael
    Linköping University, Department of Medical and Health Sciences. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping.
    Johansson, Peter
    Linköping University, Department of Medical and Health Sciences. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping.
    Jaarsma, Tiny
    Linköping University, Department of Social and Welfare Studies, Division of Health, Activity and Care. Linköping University, Faculty of Medicine and Health Sciences.
    Guided Internet-delivered cognitive behavioral therapy in patients with non-cardiac chest pain: a pilot randomized controlled study2015Manuscript (preprint) (Other academic)
    Abstract [en]

    Background: Patients with recurrent episodes of non-cardiac chest pain (NCCP) may suffer from cardiac anxiety and avoidance behavior, leading to increased healthcare utilization. These patients might benefit from help and support to evaluate the perception and management of their chest pain.

    Objective: To test the feasibility of a short guided Internet-delivered CBT intervention and the effects on chest pain, cardiac anxiety, fear of body sensations and depressive symptoms in patients with NCCP compared to usual care.

    Methods: A pilot randomized controlled study was conducted. Fifteen patients, nine men and six women with a median age of 66 years (range 22-76), were randomly assigned to either intervention (n=7) or control (n=8) group. Patients had recurrent episodes of non-cardiac chest pain and suffered from cardiac anxiety and/or fear of body sensations. The intervention consisted of a four-session guided Internet-delivered CBT program containing psychoeducation, physical activity, and relaxation. The control group received usual care. All patients completed a web-based questionnaire on socio-demographic variables, chest pain frequency, cardiac anxiety, fear of body sensations, and depressive symptoms.

    Results: Five out of the seven patients in the intervention group completed all sessions as planned and two joined only parts of the program. The program was perceived as userfriendly with understandable language, adequate and varied content, and manageable homework assignments. The patients were engaged in the program for about 60 minutes per day and about 22 minutes’ therapist time was required to guide, support and give feedback to each patient through the program. Participating in the program, particularly being guided and supported, empowered and motivated many of the patients to be active and complete the program. In general, patients in both intervention and control groups improved with regard to chest pain frequency, cardiac  anxiety, fear of body sensations, and depressive symptoms, but there were no significant differences between the groups.

    Conclusions: A short guided Internet-delivered CBT program was feasible. Patients in both intervention and control groups improved with regard to chest pain frequency, cardiac anxiety, fear of body sensations, and depressive symptoms, but no significant differences were found between the groups. Patients should be followed-up for longer periods to measure the long-term effects of the intervention.

  • 21.
    Mourad, Ghassan
    et al.
    Linköping University, Department of Social and Welfare Studies, Division of Nursing Science. Linköping University, Faculty of Medicine and Health Sciences.
    Strömberg, Anna
    Linköping University, Department of Medical and Health Sciences, Division of Nursing Science. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping.
    Jonsbu, Egil
    More and Romsdal Hospital Trust, Norway; Norwegian University of Science and Technology, Norway.
    Gustafsson, Mikael
    Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping. Linköping University, Department of Medical and Health Sciences, Division of Cardiovascular Medicine.
    Johansson, Peter
    Linköping University, Department of Social and Welfare Studies, Division of Nursing Science. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Cardiology in Linköping.
    Jaarsma, Tiny
    Linköping University, Department of Social and Welfare Studies, Division of Nursing Science. Linköping University, Faculty of Medicine and Health Sciences.
    Guided Internet-delivered cognitive behavioural therapy in patients with non-cardiac chest pain - a pilot randomized controlled study2016In: Trials, ISSN 1745-6215, E-ISSN 1745-6215, Vol. 17, p. 1-12, article id 352Article in journal (Refereed)
    Abstract [en]

    Background: Patients with recurrent episodes of non-cardiac chest pain may experience cardiac anxiety and avoidance behavior, leading to increased healthcare utilization. These patients might benefit from help and support to evaluate the perception and management of their chest pain. The purpose of this study was to test the feasibility of a short guided Internet-delivered cognitive behavioural therapy (CBT) program and explore the effects on cardiac anxiety, fear of body sensations, depressive symptoms, and chest pain in patients with non-cardiac chest pain, compared with usual care. Methods: A pilot randomized controlled study was conducted. Fifteen patients with non-cardiac chest pain with cardiac anxiety or fear of body sensations, aged 22-76 years, were randomized to intervention (n = 7) or control (n = 8) groups. The four-session CBT program contained psychoeducation, physical activity, and relaxation. The control group received usual care. Data were collected before and after intervention. Results: Five of seven patients in the intervention group completed the program, which was perceived as user-friendly with comprehensible language, adequate and varied content, and manageable homework assignments. Being guided and supported, patients were empowered and motivated to be active and complete the program. Patients in both intervention and control groups improved with regard to cardiac anxiety, fear of body sensations, and depressive symptoms, but no significant differences were found between the groups. Conclusions: The Internet-delivered CBT program seems feasible for patients with non-cardiac chest pain, but needs to be evaluated in larger groups and with a longer follow-up period.

  • 22.
    Pettersson, Jonas
    et al.
    Department of Cell and Molecular Biology, University of Umeå, Umeå, Sweden.
    Nordfelth, Roland
    Department of Cell and Molecular Biology, University of Umeå, Umeå, Sweden.
    Dubinina, Elena
    Department of Cell and Molecular Biology, University of Umeå, Umeå, Sweden.
    Bergman, Tomas
    Department of Cell and Molecular Biology, University of Umeå, Umeå, Sweden.
    Gustafsson, Mikael
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Magnusson, Karl Eric
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Wolf-Watz, Hans
    Department of Cell and Molecular Biology, University of Umeå, Umeå, Sweden.
    Modulation of Virulence Factor Expression by Pathogen Target Cell Contact1996In: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 273, no 5279, p. 1231-1233Article in journal (Refereed)
    Abstract [en]

    Upon contact with the eukaryotic cell, Yersinia pseudotuberculosis increased the rate of transcription of virulence genes (yop), as determined by in situ monitoring of light emission from individual bacteria expressing luciferase under the control of the yopE promoter. The microbe-host interaction triggered export of LcrQ, a negative regulator of Yop expression, via the Yop-type III secretion system. The intracellular concentration of LcrQ was thereby lowered, resulting in increased expression of Yops. These results suggest a key role for the type III secretion system of pathogenic bacteria to coordinate secretion with expression of virulence factors after physical contact with the target cell.

  • 23.
    Wilsson, Åsa
    et al.
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Lundqvist, Helen
    Department of Medical Microbiology and Immunology, University of Göteborg.
    Gustafsson, Mikael
    Linköping University, Department of Medicine and Care, Internal Medicine. Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Stendahl, Olle
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Killing of phagocytosed Staphylococcus aureus by human neutrophils requires intracellular free calcium1996In: Journal of Leukocyte Biology, ISSN 0741-5400, E-ISSN 1938-3673, Vol. 59, no 6, p. 902-907Article in journal (Refereed)
    Abstract [en]

    The mobilization of intracellular calcium plays an important role in regulating neutrophil activation. With this in mind we investigated the effect of intra- and extracellular calcium on the ability of human neutrophils to kill complement-opsonized Staphylococcus aureus. We found that a rise in intracellular calcium is necessary for efficient killing of phagocytosed S. aureus. In the presence of extracellular calcium, killing of ingested bacteria in calcium-buffered neutrophils compared with normal cells was slightly reduced. Calcium buffering had no effect on phagocytic uptake by the neutrophils, but did decrease the generation of toxic oxygen metabolites, measured as chemiluminescence (CL). In nondepleted and calcium-depleted cells, removal of extracellular calcium did not affect ingestion but did cause a marked decrease in the ability to kill the bacteria. In parallel, the CL response was substantially reduced or completely blocked. These data show that calcium is not a prerequisite for phagocytosis of S. aureus by human neutrophils, but does play a vital role in the post-ingestion killing of the bacteria by regulating the generation of toxic oxygen metabolites.

  • 24.
    Zimmerli, Stefan
    et al.
    Division of Infectious Diseases and The Rosalind Russell Arthritis Research Laboratory, San Francisco General Hospital, University of California at San Francisco, San Francisco, California, USA.
    Majeed, Meytham
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Gustafsson, Mikael
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Stendahl, Olle
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Sanan, David A.
    Gladstone Institute of Cardiovascular Disease, San Francisco, California, USA.
    Ernst, Joel D.
    Division of Infectious Diseases and The Rosalind Russell Arthritis Research Laboratory, San Francisco General Hospital, University of California at San Francisco, San Francisco, California, USA.
    Phagosome-Lysosome Fusion Is a Calcium-independent Event in Macrophages1996In: Journal of Cell Biology, ISSN 0021-9525, E-ISSN 1540-8140, Vol. 132, no 1-2, p. 49-61Article in journal (Refereed)
    Abstract [en]

    Phagosome-lysosome membrane fusion is a highly regulated event that is essential for intracellular killing of microorganisms, Functionally, it represents a form of polarized regulated secretion, which is classically dependent on increases in intracellular ionized calcium ([Ca2+](i)). Indeed, increases in [Ca2+](i) are essential for phagosome-granule (lysosome) fusion in neutrophils and for lysosomal fusion events that mediate host cell invasion by Trypanosoma cruzi trypomastigotes, Since several intracellular pathogens survive in macrophage phagosomes that do not fuse with lysosomes, we examined the regulation of phagosome-lysosome fusion in macrophages. Macrophages (MO) were treated with 12.5 mu M bis-(2-amino-S-methylphenoxy) ethane-N,N,N',N',-tetraacetic acid tetraacetoxymethyl ester (MAPT/AM), a cell-permeant calcium chelator which reduced resting cytoplasmic [Ca2+]; from 80 nM to less than or equal to 20 nM and completely blocked increases in [Ca2+](i) in response to multiple stimuli, even in the presence of extracellular calcium, Subsequently, MO phagocytosed serum-opsonized zymosan, staphylococci, or Mycobacterium bovis, Microbes were enumerated by 4',6-diamidino-2-phenylindole, dihydrochloride (DAPI) staining, and phagosome-lysosome fusion was scored using both lysosome-associated membrane protein (LAMP-1) as a membrane marker and rhodamine dextran as a content marker for lysosomes, Confirmation of phagosomelysosome fusion by electron microscopy validated the fluorescence microscopy findings, We found that phagosome-lysosome fusion in MO occurs normally at very low [Ca2+](i) (less than or equal to 20 nM), Kinetic analysis showed that in MO none of the steps leading from particle binding to eventual phagosome-lysosome fusion are regulated by [Ca2+](i) in a rate-limiting way. Furthermore, confocal microscopy revealed no difference in the intensity of LAMP-1 immunofluorescence in phagolysosome membranes in calcium-buffered vs, control macrophages, We conclude that neither membrane recognition nor fusion events in the phagosomal pathway in macrophages are dependent on or regulated by calcium.

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