Homo sapiens
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ABSTRACT:
PROVIDER: PRJNA378945 | ENA |
REPOSITORIES: ENA
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Project description:To identify if platelet activation would result in altered platelet miRNA profile, not only in total RNA sample, also in AGO2-immunoprecipitation(AGO2-IP) products. To determinate if altered platelet miRNAs could regulate de novo protein synthesis of angiogenic factors in activated platelets, and even more importantly, if miRNA-regulated platelet angiogenic factor synthesis could result in changes of platelet angiogenic activities.
2017-03-14 | GSE96533 | GEO
Project description:Platelet activation is the key event triggering thrombus formation in physiological and pathological conditions, such as acute coronary syndromes. Current therapies using antiaggregants still fail to prevent thrombotic coronary events in a significant number of patients, indicating that the mechanisms modulating platelet response during activation need to be clarified. The evidence that platelets are capable of de novo protein synthesis in response to stimuli raised the issue of how the activity of megakaryocyte-derived mRNAs is regulated in these anucleate cell fragments. We applied a combined multi-omics approach to investigate this phenomenon in platelets from healthy donors activated in vitro with Collagen or Thrombin Receptor Activating Peptide. Combining HiRIEF LC-MS to transcriptome analysis by RNA-Seq allowed platelet proteome characterization at deep coverage, revealing a significant effect of either stimulus on proteome composition. In silico intron retention analysis was then applied to search for splicing events induced by platelet activation, coupled to unbiased proteogenomics, to correlate intron retention in resting platelets to intron removal by RNA splicing during activation. This allowed identification of a set of transcripts, specifically involved in platelet shape changes, showing reduced intron retention and high peptide representation at exon-exon junctions in activated vs resting platelets. These results indicate that RNA splicing events takes place in platelets during activation and that pre-mRNA maturation of specific transcripts is part of the activation cascade and could therefore provide novel molecular markers of platelet activation status in acute coronary syndromes and other pathological conditions.
2018-01-15 | E-MTAB-6148 | biostudies-arrayexpress
Project description:In an experimental model of tumor dormancy, heat shock protein 27 (HSP27) was up-regulated in angiogenic human breast cancer cells when compared with non-angiogenic cells. Stable down-regulation of HSP27 in angiogenic tumor cells was followed by long-term tumor dormancy in vivo and associated with reduced intra-tumoral endothelial cell proliferation, decreased secretion of VEGF and bFGF from tumor cells, and increased expression of thrombospondin-1. Phosphorylation of the transcription factor STAT3 and nuclear expression of NFκB were reduced following suppression of HSP27. In contrast, tumor cell proliferation and apoptosis were not affected. By clinical validation, high HSP27 expression was associated with markers of aggressive tumors and decreased survival in breast cancer and melanoma patients. Our present findings suggest a link between HSP27 and dormancy through tumor-vascular interactions. Targeting HSP27, a multifunctional cytoprotective protein, might offer a novel strategy in cancer treatment.
2010-02-28 | E-TABM-883 | biostudies-arrayexpress
Project description:A numerical simulation model to reproduce in vitro experiments of blood coagulation in microfluidic capillaries. A reduced one-equation model of thrombin distribution from the previous model under simplifying hypotheses and used to determine the conditions of clotting initiation on the platelet count, the shear rate, and the plasma composition.
2023-04-25 | MODEL2011030003 | BioModels
Project description:To identify the miRNA expressing profiles of Platelet microparticlesï¼PMPs, we have employed the Agilent Human miRNA 8Ã60K (Design ID:046064) microarray. Platelet microparticles. The platelets were derived from citrated blood of healthy human donors under an Institutional Review Board-approved protocol. Platelets were isolated after centrifugation of blood (1200r for 30 min at 21â), then the supernatant (platelet-rich plasma) was centrifuged at 2000r for 30 min at 21â, and the pellet containing platelets was resuspended in RPMI-1640 medium (HyClone, Logan, UT). Platelets were counted (Clinical Laboratory, Shanghai First Maternity and Infant Hospital, Shanghai) and adjusted to a density of 150 à 106 cells/mL before supplement with 1.5% ACD(sigma ) and stimulated with thrombin (1.0 u/mL; Takeda Austria) for 1 h. PMPs were in the supernatant after centrifugation at 4000r for 10 min at 4â,then the supernatants were centrifuged at 50,000 à g for 60 min at 4 °C. The pellets containing MPs were resuspended in RPMI-1640 medium and quantified by BCA method. The gene expressions of three independent paired PMPs from platelets stimulated by thrombin or apoptosis.
2016-01-14 | E-GEOD-76789 | biostudies-arrayexpress
Project description:Cysteinyl leukotrienes (cysLT), i.e. LTC4, LTD4, and LTE4, are lipid mediators derived from the 5-lipoxygenase pathway. The cysLT receptors cysLT1-R and cysLT2-R are expressed on different target cells and mediate inflammatory reactions in tissue- and LT-R-specific ways. Though endothelial cells (ECs) predominantly express cysLT2-Rs, their role in vascular biology remains to be defined. To delineate cysLT2-R´s action, we stimulated human umbilical vein EC with 100 nM LTD4 for 60 min, determined gene signatures by microarrays, and characterized the resulting EC phenotypes. As controls, we compared LTD4-induced genes with those induced by 10 nM thrombin, a prototype vasoactive activator of EC that binds to protease-activated receptor 1 (PAR-1). Following application of stringent filters 37 LTD4-upregulated genes were identified (> 2.5fold stimulation). Surprisingly, most of the LTD4-regulated genes were also induced by thrombin and expression of cysLT2-R- and PAR-1-regulated genes strongly correlated (Pearson correlation coefficient: r = 0.90). Moreover, LTD4 + thrombin, when added together, augmented expression of LTD4- or thrombin-stimulated genes (Wilcoxon signed rank test: p < 0.01). Prominently induced genes that may play roles in vascular injury were studied in detail: Early growth response (EGR) and nuclear receptor subfamily 4 group A; E-selectin; CXC ligand 2; interleukin 8 (IL-8); a disintegrin-like and metalloprotease (reprolysin type) with thrombospondin type 1 motif 1 (ADAMTS-1); and tissue factor (TF). Transcripts of these genes peaked at approximately 60 min, were unaffected by the cysLT1-R antagonist montelukast, and were superinduced by cycloheximide. The EC phenotype was markedly altered: LTD4 induced de novo synthesis of EGR1 protein and EGR1 localized in the nucleus in LTD4-stimulated cells; LTD4 upregulated IL-8 formation and secretion; and LTD4 raised TF protein and TF-dependent EC pro-coagulant activity. These data show that cysLT2-R activation results in a pro-inflammatory EC phenotype through activation of immediate-early genes that resemble those induced by PAR-1. As LTD4 and thrombin are formed concomitantly during vascular injury and pro-thrombotic states, cysLT2-R and PAR-1 may collaborate in vivo to mediate vascular injury and repair. Keywords: Leukotriene Transcriptome, Thrombin Transcriptome, HUVEC, Immediate-Early Gene Expression, Cysteinyl Leukotriene 2 Receptor Gene Signature in HUVEC
2006-03-14 | GSE3589 | GEO
Project description:RNAi-mediated knockdown of DICER1 and DROSHA, enzymes critically involved in miRNA biogenesis, has been postulated to affect the homeostasis and the angiogenic capacity of human endothelial cells. To re-evaluate this issue, we reduced the expression of DICER1 or DROSHA by RNAi-mediated knockdown and subsequently investigated the effect of these interventions on the angiogenic capacity of human umbilical vein endothelial cells (HUVEC) in vitro (proliferation, migration, tube formation, endothelial cell spheroid sprouting) and in a HUVEC xenograft assay in immune incompetent NSGTM mice in vivo. In contrast to previous reports, neither knockdown of DICER1 nor knockdown of DROSHA profoundly affected migration or tube formation of HUVEC or the angiogenic capacity of HUVEC in vivo. Furthermore, knockdown of DICER1 and the combined knockdown of DICER1 and DROSHA tended to increase VEGF-induced BrdU incorporation and induced angiogenic sprouting from HUVEC spheroids. Consistent with these observations, global proteomic analyses showed that knockdown of DICER1 or DROSHA only moderately altered HUVEC protein expression profiles but additively reduced, for example, expression of the angiogenesis inhibitor thrombospondin-1. In conclusion, global reduction of miRNA biogenesis by knockdown of DICER1 or DROSHA does not inhibit the angiogenic capacity of HUVEC. Further studies are therefore needed to elucidate the influence of these enzymes in the context of human endothelial cell-related angiogenesis.
2022-02-17 | PXD027923 | Pride
Project description:Cysteinyl leukotrienes (cysLT), i.e. LTC4, LTD4, and LTE4, are lipid mediators derived from the 5-lipoxygenase pathway. The cysLT receptors cysLT1-R and cysLT2-R are expressed on different target cells and mediate inflammatory reactions in tissue- and LT-R-specific ways. Though endothelial cells (ECs) predominantly express cysLT2-Rs, their role in vascular biology remains to be defined. To delineate cysLT2-R´s action, we stimulated human umbilical vein EC with 100 nM LTD4 for 60 min, determined gene signatures by microarrays, and characterized the resulting EC phenotypes. As controls, we compared LTD4-induced genes with those induced by 10 nM thrombin, a prototype vasoactive activator of EC that binds to protease-activated receptor 1 (PAR-1). Following application of stringent filters 37 LTD4-upregulated genes were identified (> 2.5fold stimulation). Surprisingly, most of the LTD4-regulated genes were also induced by thrombin and expression of cysLT2-R- and PAR-1-regulated genes strongly correlated (Pearson correlation coefficient: r = 0.90). Moreover, LTD4 + thrombin, when added together, augmented expression of LTD4- or thrombin-stimulated genes (Wilcoxon signed rank test: p < 0.01). Prominently induced genes that may play roles in vascular injury were studied in detail: Early growth response (EGR) and nuclear receptor subfamily 4 group A; E-selectin; CXC ligand 2; interleukin 8 (IL-8); a disintegrin-like and metalloprotease (reprolysin type) with thrombospondin type 1 motif 1 (ADAMTS-1); and tissue factor (TF). Transcripts of these genes peaked at approximately 60 min, were unaffected by the cysLT1-R antagonist montelukast, and were superinduced by cycloheximide. The EC phenotype was markedly altered: LTD4 induced de novo synthesis of EGR1 protein and EGR1 localized in the nucleus in LTD4-stimulated cells; LTD4 upregulated IL-8 formation and secretion; and LTD4 raised TF protein and TF-dependent EC pro-coagulant activity. These data show that cysLT2-R activation results in a pro-inflammatory EC phenotype through activation of immediate-early genes that resemble those induced by PAR-1. As LTD4 and thrombin are formed concomitantly during vascular injury and pro-thrombotic states, cysLT2-R and PAR-1 may collaborate in vivo to mediate vascular injury and repair. Experiment Overall Design: HUVEC in passage 1 were cultured in serum-free EC medium until cells reached confluency. HUVEC were stimulated by 100 nM LTD4 (n = 4) or 10 nM thrombin (n = 4) or both agonists concomitantly (n = 3). The immediate-early gene programs of these experimental groups were identified at 60 minutes. In addition, one HUVEC preparation was stimulated with 100 nM LTD4 for 6 h and 24 h.
2007-10-27 | E-GEOD-3589 | biostudies-arrayexpress
Project description:Platelet activation induces the secretion of proteins that promote platelet aggregation and inflammation. However, detailed analysis of the released platelet proteome is hampered by platelets’ tendency to pre-activate during their isolation and a lack of sensitive protocols for low abundance releasate analysis. Here we detail the most sensitive analysis to date of the platelet releasate proteome with the detection of >1,300 proteins. Unbiased scanning for post-translational modifications within releasate proteins highlighted O-glycosylation as being a major component. For the first time, we detected O-fucosylation on previously uncharacterised sites including multimerin-1 (MMRN1), a major alpha granule protein that supports platelet adhesion to collagen and is a carrier for platelet factor V. The N-terminal elastin microfibril interface (EMI) domain of MMRN1, a key site for protein-protein interaction, was O-fucosylated at a conserved threonine within a new domain context. Our data suggest that either protein O-fucosyltransferase 1 (POFUT1), or a novel POFUT, may be responsible for this modification. Mutating this O-fucose site on the EMI domain led to a >50% reduction of MMRN1 secretion, supporting a key role of EMI O-fucosylation in MMRN1 secretion. By comparing releasates from resting and thrombin-treated platelets, 202 proteins were found to be significantly released after high-dose thrombin stimulation. Complementary quantification of the platelet lysates identified >3,800 proteins, which confirmed the platelet origin of releasate proteins by anti-correlation analysis. Low-dose thrombin treatment yielded a smaller subset of significantly regulated proteins with fewer secretory pathway enzymes. The extensive platelet proteome resource provided here (larancelab.com/platelet-proteome) allows identification of novel regulatory mechanisms for drug targeting to address platelet dysfunction and thrombosis.
2024-02-15 | PXD045535 | Pride
Project description:Here we characterized the proteins associated with detergent resistant membranes (DRMs) from resting and activated human platelets. We achieved a proteomic profile of 141 DRM proteins involved in many biological pathways including 'PAR-1-mediated thrombin signal events', 'Integrin family cell surface interactions', 'Platelet activation, signalling and aggregation' and 'Platelet degranulation'. Therefore, a high level of relevant platelet signalling and trafficking proteins were identified e.g., Rap 1b, Src, SNAP-23 and syntaxin-11, implicating platelet DRMs as concentrating platforms not only in the critical platelet function of activation but also in secretion/degranulation.
2017-03-28 | MSV000080708 | MassIVE