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Osman, A., Hitzler, W. E., Ameur, A. & Provost, P. (2015). Differential Expression Analysis by RNA-Seq Reveals Perturbations in the Platelet mRNA Transcriptome Triggered by Pathogen Reduction Systems. PLOS ONE, 10(7), e0133070
Open this publication in new window or tab >>Differential Expression Analysis by RNA-Seq Reveals Perturbations in the Platelet mRNA Transcriptome Triggered by Pathogen Reduction Systems
2015 (English)In: PLOS ONE, E-ISSN 1932-6203, Vol. 10, no 7, p. e0133070-Article in journal (Refereed) Published
Abstract [en]

Platelet concentrates (PCs) are prepared at blood banks for transfusion to patients in certain clinical conditions associated with a low platelet count. To prevent transfusion-transmitted infections via PCs, different pathogen reduction (PR) systems have been developed that inactivate the nucleic acids of contaminating pathogens by chemical cross-linking, a mechanism that may also affect platelets nucleic acids. We previously reported that treatment of stored platelets with the PR system Intercept significantly reduced the level of half of the microRNAs that were monitored, induced platelet activation and compromised the platelet response to physiological agonists. Using genome-wide differential expression (DE) RNA sequencing (RNA-Seq), we now report that Intercept markedly perturbs the mRNA transcriptome of human platelets and alters the expression level of greater than800 mRNAs (Pless than0.05) compared to other PR systems and control platelets. Of these, 400 genes were deregulated with DE corresponding to fold changes (FC) greater than= 2. At the p-value less than 0.001, as many as 147 genes were deregulated by greater than= 2-fold in Intercept-treated platelets, compared to none in the other groups. Finally, integrated analysis combining expression data for microRNA (miRNA) and mRNA, and involving prediction of miRNA-mRNA interactions, disclosed several positive and inverse correlations between miRNAs and mRNAs in stored platelets. In conclusion, this study demonstrates that Intercept markedly deregulates the platelet mRNA transcriptome, concomitant with reduced levels of mRNA-regulatory miRNAs. These findings should enlighten authorities worldwide when considering the implementation of PR systems, that target nucleic acids and are not specific to pathogens, for the management of blood products.

Place, publisher, year, edition, pages
Public Library of Science, 2015
National Category
Clinical Medicine
Identifiers
urn:nbn:se:liu:diva-120648 (URN)10.1371/journal.pone.0133070 (DOI)000358194900119 ()26172280 (PubMedID)
Note

Funding Agencies|Regional Council of Ostergotland, Sweden [LIO-353011]; Canadian Blood Services/Canadian Institutes of Health Research (CIHR) Blood Utilization and Conservation Initiative via Health Canada [293933]

Available from: 2015-08-20 Created: 2015-08-20 Last updated: 2022-04-19
Osman, A., Hitzler, W. E., Meyer, C. U., Landry, P., Corduan, A., Laffont, B., . . . Provost, P. (2015). Effects of pathogen reduction systems on platelet microRNAs, mRNAs, activation, and function.. Platelets, 26(2), 154-163
Open this publication in new window or tab >>Effects of pathogen reduction systems on platelet microRNAs, mRNAs, activation, and function.
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2015 (English)In: Platelets, ISSN 0953-7104, E-ISSN 1369-1635, Vol. 26, no 2, p. 154-163Article in journal (Refereed) Published
Abstract [en]

Pathogen reduction (PR) systems for platelets, based on chemically induced cross-linking and inactivation of nucleic acids, potentially prevent transfusion transmission of infectious agents, but can increase clinically significant bleeding in some clinical studies. Here, we documented the effects of PR systems on microRNA and mRNA levels of platelets stored in the blood bank, and assessed their impact on platelet activation and function. Unlike platelets subjected to gamma irradiation or stored in additive solution, platelets treated with Intercept (amotosalen + ultraviolet-A [UVA] light) exhibited significantly reduced levels of 6 of the 11 microRNAs, and 2 of the 3 anti-apoptotic mRNAs (Bcl-xl and Clusterin) that we monitored, compared with platelets stored in plasma. Mirasol (riboflavin + UVB light) treatment of platelets did not produce these effects. PR neither affected platelet microRNA synthesis or function nor induced cross-linking of microRNA-sized endogenous platelet RNA species. However, the reduction in the platelet microRNA levels induced by Intercept correlated with the platelet activation (p < 0.05) and an impaired platelet aggregation response to ADP (p < 0.05). These results suggest that Intercept treatment may induce platelet activation, resulting in the release of microRNAs and mRNAs from platelets. The clinical implications of this reduction in platelet nucleic acids secondary to Intercept remain to be established.

National Category
Clinical Medicine
Identifiers
urn:nbn:se:liu:diva-115266 (URN)10.3109/09537104.2014.898178 (DOI)000351740700009 ()24749844 (PubMedID)
Available from: 2015-03-11 Created: 2015-03-11 Last updated: 2022-04-19Bibliographically approved
Bastami, S., Gupta, A., Zackrisson, A. L., Ahlner, J., Osman, A. & Uppugunduri, S. (2014). Influence of UGT2B7, OPRM1 and ABCB1 gene polymorphisms on morphine use. Basic & Clinical Pharmacology & Toxicology, 115(5), 423-431
Open this publication in new window or tab >>Influence of UGT2B7, OPRM1 and ABCB1 gene polymorphisms on morphine use
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2014 (English)In: Basic & Clinical Pharmacology & Toxicology, ISSN 1742-7835, E-ISSN 1742-7843, Vol. 115, no 5, p. 423-431Article in journal (Refereed) Published
Abstract [en]

Therapeutic modulation of pain with morphine and other opioids is associated with significant variation in, both, effects and adverse effects in individual patients. Many factors including gene polymorphisms have been shown to contribute to the interindividual variability in the response to opioids. The aim of this study was to investigate the significance of UGT2B7, OPRM1 and ABCB1 polymorphisms for interindividual variability in morphine induced analgesia in patients undergoing hysterectomy. The frequency of these polymorphisms was also investigated in forensic autopsy cases as morphine is also a very commonly abused drug

Blood samples were collected from 40 patients following abdominal hysterectomy, 24 hours after initiation of analgesia through a PCA pump. Samples were genotyped and analysed for morphine and its metabolites. We also genotyped approximately 200 autopsy cases found positive for morphine in routine forensic analysis.

Patients homozygous for UGT2B7 802C needed significantly lower dose of morphine for pain relief. The same trend was observed for patients homozygous for ABCB1 1236T and 3435T, as well as to OPRM1 118A. Dose of morphine in patients included in this study was significantly related to variation in UGT2B7 T802C. Age was significantly related to both dose and concentration of morphine in blood.

Regression analysis showed that 30% of differences in variation in morphine dose could be explained by SNPs in these genes. The genotype distribution was similar between the forensic cases and the patients. However, the mean concentration of morphine was higher in forensic cases compared to patients.

We conclude that gene polymorphisms contribute significantly to the variation in morphine levels observed in individual patients.

Place, publisher, year, edition, pages
Wiley, 2014
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-96791 (URN)10.1111/bcpt.12248 (DOI)000344015300008 ()24703092 (PubMedID)
Available from: 2013-08-27 Created: 2013-08-27 Last updated: 2022-04-19Bibliographically approved
Saleiban, A., Faxälv, L., Claesson, K., Jönsson, J.-I. & Osman, A. (2014). miR-20b regulates expression of proteinase-activated receptor-1 (PAR-1) thrombin receptor in melanoma cells. Pigment Cell & Melanoma Research, 27(3), 431-441
Open this publication in new window or tab >>miR-20b regulates expression of proteinase-activated receptor-1 (PAR-1) thrombin receptor in melanoma cells
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2014 (English)In: Pigment Cell & Melanoma Research, ISSN 1755-1471, E-ISSN 1755-148X, Vol. 27, no 3, p. 431-441Article in journal (Refereed) Published
Abstract [en]

The proteinase-activated receptor 1 (PAR-1) plays a central role in melanoma progression and its expression level is believed to correlate with the degree of cancer invasiveness. Here, we show that PAR-1 is post-transcriptionally regulated by miR-20b microRNA in human melanoma cells. PAR-1 was found to be expressed in metastatic melanoma cells but was barely detectable in primary melanoma. By transducing primary melanoma cells with a lentivirus containing a 3-UTR construct of PAR-1 mRNA, we could show that endogenous melanoma microRNAs interacted with PAR-1 3-UTR and silenced a fused luciferase reporter. Transfection of an inhibitor against miR-20b into primary melanoma cells reversed this process. Finally, transfection of miR-20b mimic into metastatic melanoma cells caused downregulation of the luciferase reporter. We conclude that miR-20b regulates expression of melanoma PAR-1 receptor, which may explain the differential expression of PAR-1 observed in human melanoma.

Place, publisher, year, edition, pages
Wiley, 2014
Keywords
melanoma; metastasis; proteinase-activated receptor-1; gene expression regulation; microRNAs
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-106844 (URN)10.1111/pcmr.12217 (DOI)000334170900014 ()
Available from: 2014-05-28 Created: 2014-05-23 Last updated: 2022-04-19
Osman, A., Uhlin, F., Frånlund, E., Fernström, A. & Magnusson, P. (2013). Exon resequencing of the gene encoding UCMA/GRP reveals a common carboxy-terminal 138Thr > Ser Polymorphism. Clinical Laboratory, 59(11-12), 1397-1401
Open this publication in new window or tab >>Exon resequencing of the gene encoding UCMA/GRP reveals a common carboxy-terminal 138Thr > Ser Polymorphism
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2013 (English)In: Clinical Laboratory, ISSN 1433-6510, Vol. 59, no 11-12, p. 1397-1401Article in journal (Refereed) Published
Abstract [en]

Background: The upper zone of growth plate and cartilage matrix-associated protein (UCMA), also called Gla-rich protein (GRP), is a novel protein found at sites af-fected by pathological calcifications.Methods: We performed a full exon resequencing on DNA samples from 17 chronic kid-ney disease (CKD) patients (stage 5) and compared the results with 121 healthy con-trols in a Swedish population.Results: A novel non-synonymous single nucleotide polymorphism (SNP) causing a car-boxy-terminal amino acid exchange was found. This SNP involves an alteration of the last ACC codon for threonine in exon 5 (adjacent to the stop codon) to an AGC ser-ine codon (138Thr > Ser). Six controls and two CKD patients were heterozygous for the 138Thr > Ser polymorphism. Both patients had histories of vascular calcifica-tion; however, it is uncertain whether this SNP has any significance for the func-tional domains of the UCMA protein. In addition, a heterozygous transversion muta-tion was found in a patient at SNP rs4750328 (A/G) in intron 2, involving an ex-change of the ancestral A allele to a T base.Conclusions: The 138Thr > Ser polymorphism seems to be the only non-synonymous SNP found in the UCMA gene in a Swedish population.

Place, publisher, year, edition, pages
Clinical Laboratory Publications, 2013
Keywords
chronic kidney disease, polymorphisms, vascular calcification
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-102330 (URN)10.7754/Clin.Lab.2013.130123 (DOI)000328725100024 ()
Available from: 2013-12-05 Created: 2013-12-05 Last updated: 2022-04-19Bibliographically approved
Kissopoulou, A., Jonasson, J., Lindahl, T. & Osman, A. (2013). Next Generation Sequencing Analysis of Human Platelet PolyA plus mRNAs and rRNA-Depleted Total RNA. PLOS ONE, 8(12), 81809
Open this publication in new window or tab >>Next Generation Sequencing Analysis of Human Platelet PolyA plus mRNAs and rRNA-Depleted Total RNA
2013 (English)In: PLOS ONE, E-ISSN 1932-6203, Vol. 8, no 12, p. 81809-Article in journal (Refereed) Published
Abstract [en]

Background: Platelets are small anucleate cells circulating in the blood vessels where they play a key role in hemostasis and thrombosis. Here, we compared platelet RNA-Seq results obtained from polyA+ mRNA and rRNA-depleted total RNA. Materials and Methods: We used purified, CD45 depleted, human blood platelets collected by apheresis from three male and one female healthy blood donors. The Illumina HiSeq 2000 platform was employed to sequence cDNA converted either from oligo(dT) isolated polyA+ RNA or from rRNA-depleted total RNA. The reads were aligned to the GRCh37 reference assembly with the TopHat/Cufflinks alignment package using Ensembl annotations. A de novo assembly of the platelet transcriptome using the Trinity software package and RSEM was also performed. The bioinformatic tools HTSeq and DESeq from Bioconductor were employed for further statistical analyses of read counts. Results: Consistent with previous findings our data suggests that mitochondrially expressed genes comprise a substantial fraction of the platelet transcriptome. We also identified high transcript levels for protein coding genes related to the cytoskeleton function, chemokine signaling, cell adhesion, aggregation, as well as receptor interaction between cells. Certain transcripts were particularly abundant in platelets compared with other cell and tissue types represented by RNA-Seq data from the Illumina Human Body Map 2.0 project. Irrespective of the different library preparation and sequencing protocols, there was good agreement between samples from the 4 individuals. Eighteen differentially expressed genes were identified in the two sexes at 10% false discovery rate using DESeq. Conclusion: The present data suggests that platelets may have a unique transcriptome profile characterized by a relative over-expression of mitochondrially encoded genes and also of genomic transcripts related to the cytoskeleton function, chemokine signaling and surface components compared with other cell and tissue types. The in vivo functional significance of the non-mitochondrial transcripts remains to be shown.

Place, publisher, year, edition, pages
Public Library of Science, 2013
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-103875 (URN)10.1371/journal.pone.0081809 (DOI)000328730300052 ()
Available from: 2014-01-31 Created: 2014-01-30 Last updated: 2022-04-19
Osman, A. (2012). MicroRNAs in Health and Disease - Basic Science and Clinical Applications. CLINICAL LABORATORY, 58(5-6), 393-402
Open this publication in new window or tab >>MicroRNAs in Health and Disease - Basic Science and Clinical Applications
2012 (English)In: CLINICAL LABORATORY, ISSN 1433-6510, Vol. 58, no 5-6, p. 393-402Article, review/survey (Refereed) Published
Abstract [en]

MicroRNAs (miRs) are small RNAs that fine-tune gene expression at posttranscriptional level. They are involved in virtually all physiological processes, such as development and differentiation, heart function, metabolism, haemostasis, and apoptosis. Anucleated human platelets were revealed to contain comparatively many different miR species relative to nucleated cells, suggesting a functional impact of miRs on these small cells. In recent years, numerous studies have established the significance of miRs in different physiological states. Sufficient evidence exists to suggest that miRs are involved in a range of diseases, including cancer and cardiovascular disorders. Aberrant expression levels of miRs are found in tissues and in sera from patients with different forms of malignant tumours. Utilising these miRs as biomarkers for disease is a valuable diagnostic strategy. In addition, a novel class of synthetic oligonucleotides, called antagomirs, has recently been introduced as efficient silencers of overexpressed miRs in cancer. Overall, miRs represent an important class of small RNAs with a huge impact on health and disease. Future studies will further illuminate the potential value of miRs in diagnostics and therapeutics.

Place, publisher, year, edition, pages
CLIN LAB PUBL, 2012
Keywords
MicroRNAs; haematopoiesis; platelets; cancer; cardiovascular disease
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-79720 (URN)000305711100003 ()
Available from: 2012-08-13 Created: 2012-08-13 Last updated: 2022-04-19
Nylander, M., Osman, A., Ramström, S., Åklint, E., Larsson, A. & Lindahl, T. (2012). The role of thrombin receptors PAR1 and PAR4 for PAI-1 storage, synthesis and secretion by human platelets. Thrombosis Research, 129(4), E51-E58
Open this publication in new window or tab >>The role of thrombin receptors PAR1 and PAR4 for PAI-1 storage, synthesis and secretion by human platelets
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2012 (English)In: Thrombosis Research, ISSN 0049-3848, E-ISSN 1879-2472, Vol. 129, no 4, p. E51-E58Article in journal (Refereed) Published
Abstract [en]

INTRODUCTION:

Arterial thrombi contain more platelets than venous thrombi and are more resistant to fibrinolysis. This resistance could partly be due to plasminogen activator inhibitor 1 (PAI-1) secreted by platelets. The aim of this study was to elucidate differences between thrombin receptors protease-activated receptor (PAR) 1 and 4 and platelet storage, secretion and synthesis of platelet PAI-1, as compared to other platelet α-granule proteins such as VEGF and endostatin.

MATERIALS AND METHODS:

Human isolated platelets were incubated with thrombin (0.5U/ml), PAR1-activating peptide (AP) (0.4-30μM) or PAR4-AP (1.5-300μM) for up to 24hours. ELISA, western blot and fluorescence microscopy were used to measure secretion, contents and localization of PAI-1, VEGF and endostatin.

RESULTS:

Our results show that PAI-1 and VEGF might be co-localized and that endostatin does not co-localize with either PAI-1 or VEGF. PAI-1 and VEGF show a similar secretion pattern, being more sensitive to low grade PAR1 activation, but secretion was also observed with higher concentrations of PAR4-APs. PAI-1 is secreted in an active form. PAI-1 mRNA was found in platelets, and elevated levels of PAI-1 were detected after 24hours incubation of platelets.

CONCLUSIONS:

PAI-1 and VEGF, but not endostatin, might be stored in the same α-granule in human platelets. PAI-1 and VEGF also show a similar secretion pattern, being more sensitive to PAR1 than to PAR4 activation, but the secretion is not exclusively selective. Our results also show that platelet PAI-1 is increased if incubated for 24hours, both with addition of PAR1-activating peptide and without activation, which could indicate de novo synthesis.

Place, publisher, year, edition, pages
Elsevier, 2012
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-71393 (URN)10.1016/j.thromres.2011.12.021 (DOI)000301587400010 ()
Note

funding agencies|Swedish Research Council| K2010-65X-15060-07-3 |strategic research area Cardiovascular Inflammation Research Center (CIRC)||County Council of Ostergotland||University of Linkoping||

Available from: 2011-10-14 Created: 2011-10-14 Last updated: 2022-04-19Bibliographically approved
Osman, A. & Fälker, K. (2011). Characterization of human platelet microRNA by quantitative PCR coupled with an annotation network for predicted target genes. Platelets, 22(6), 433-441
Open this publication in new window or tab >>Characterization of human platelet microRNA by quantitative PCR coupled with an annotation network for predicted target genes
2011 (English)In: Platelets, ISSN 0953-7104, E-ISSN 1369-1635, Vol. 22, no 6, p. 433-441Article in journal (Refereed) Published
Abstract [en]

latelets are anucleate blood cells that play a crucial role in thrombosis and hemostasis. Despite their lack of nuclear DNA, platelets contain significant amounts of microRNA (miRNA) that may have vital functions in post-transcriptional gene regulation. Here, we combined comprehensive miRNA expression profiling by quantitative PCR with target prediction analysis for the most abundant miRNAs in human platelets. A network composed of predicted platelet miRNA target genes was then constructed, using annotations available in Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. In addition, we evaluated possible differences in miRNA levels between resting and thrombin-stimulated platelets. We identified 281 transcripts, including 228 mature miRNAs and 53 minor miRNAs (or miR(star)), of which six miRNAs (miR-15 a, miR-339-3 p, miR-365, miR-495, miR-98, and miR-361-3 p) were up-or down-regulated in activated human platelets (P andlt;= 0.001). A redundancy-reduced network was established that encompassed 246 genes in five statistically significant functional clusters representing platelet miRNA regulating pathways. Comparison of the 246 network genes with the platelet mRNA expression data available at ArrayExpress database confirmed that most of these genes (89%) are expressed in human platelets. In conclusion, this work affirms a recent microarray study reporting a wide-spread existence of miRNAs in human platelets. Further, we observed that thrombin stimulation was associated with altered levels of some miRNAs in platelets. The proposed functional network, combining computational prediction analysis with annotations from experimental observations, may in addition provide some information about probable miRNA target pathways in human platelets.

Place, publisher, year, edition, pages
Informa Healthcare, 2011
Keywords
Platelets, microRNA, gene expression, annotation network
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-70114 (URN)10.3109/09537104.2011.560305 (DOI)000293600300005 ()
Available from: 2011-08-19 Created: 2011-08-19 Last updated: 2022-04-19
Pfister, A. & Osman, A. (2010). Letter: The VKORC1 promoter is occupied by c-Myc transcription factor in HepG2 cells [Letter to the editor]. Thrombosis Research, 126(2), E150-E151
Open this publication in new window or tab >>Letter: The VKORC1 promoter is occupied by c-Myc transcription factor in HepG2 cells
2010 (English)In: Thrombosis Research, ISSN 0049-3848, E-ISSN 1879-2472, Vol. 126, no 2, p. E150-E151Article in journal, Letter (Other academic) Published
Abstract [en]

n/a

Place, publisher, year, edition, pages
Elsevier Science B.V., Amsterdam., 2010
Keywords
VKORC1; HepG2; c-Myc; transcription factors
National Category
Engineering and Technology
Identifiers
urn:nbn:se:liu:diva-58158 (URN)10.1016/j.thromres.2010.01.050 (DOI)000280311300033 ()
Available from: 2010-08-11 Created: 2010-08-09 Last updated: 2022-04-19Bibliographically approved
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0002-5143-8315

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