Project description:Platelets are small anucleate cells derived from the fragmentation of megakaryocytes and are involved in different biological processes especially hemostasis, thrombosis and immune response. Platelet purification is a crucial step in transcriptomic analysis, and researchers usually encounter the problem of platelet contamination by leukocytes and erythrocytes. Leukocytes contain much more RNA than platelets, thus the presence of few contaminants in platelet preparation can strongly alter transcriptome results. Using microarray technique, we compared transcriptome of platelets from the same donor, purified by common centrifugation method or using magnetic microbeads to eliminate contaminating cells.
Project description:Viruses can directly interact with platelets and modulate their function. Viral impact on platelet activation, and platelet-mediated modulations of innate and adaptive immune responses. Human herpesvirus 4, also known as Epstein–Barr virus (EBV) interaction with platelets occurs via complement receptor 2 (CR2), but the exact mechanism of action with platelets is still poorly understood. Epstein–Barr virus (EBV), is extremely efficient at establishing a persistent life-long infection in human B cells. In the present study, GeneChips were performed in human platelets from three normal donors infected with the EBV-containing supernatant of the B95.8 marmoset cell line in vitro.
Project description:Viruses can directly interact with platelets and modulate their function. Viral impact on platelet activation, and platelet-mediated modulations of innate and adaptive immune responses. Human herpesvirus 4, also known as Epstein–Barr virus (EBV) interaction with platelets occurs via complement receptor 2 (CR2), but the exact mechanism of action with platelets is still poorly understood. Epstein–Barr virus (EBV), is extremely efficient at establishing a persistent life-long infection in human B cells. In the present study, GeneChips were performed in human platelets from three normal donors infected with the EBV-containing supernatant of the B95.8 marmoset cell line in vitro.
Project description:Patients with systemic lupus erythematosus (SLE) have a markedly increased risk to develop cardiovascular disease, and traditional cardiovascular risk factors fail to account for this increased risk. We used microarray to probe the platelet transcriptome in individuals with SLE and healthy controls, and the gene and protein expression of a subset of differentially expressed genes was further investigated and correlated to platelet activation status. Real-time PCR was used to confirm a type I interferon (IFN) gene signature in patients with SLE, and the IFN-regulated proteins PRKRA, IFITM1 and CD69 (p<0.0001) were found to be up-regulated in platelets from SLE patients as compared to healthy volunteers. Notably, patients with a history of vascular disease had increased expression of type I IFN-regulated proteins as well as more activated platelets as compared with patients without vascular disease. We suggest that interferogenic immune complexes stimulate production of IFNα which up-regulates the megakaryocytic type I IFN-regulated genes and proteins. This could affect platelet activation and contribute to development of vascular disease in SLE. In addition, platelets with type I IFN signature could be a novel marker for vascular disease in SLE.
Project description:Patients with systemic lupus erythematosus (SLE) have a markedly increased risk to develop cardiovascular disease, and traditional cardiovascular risk factors fail to account for this increased risk. We used microarray to probe the platelet transcriptome in individuals with SLE and healthy controls, and the gene and protein expression of a subset of differentially expressed genes was further investigated and correlated to platelet activation status. Real-time PCR was used to confirm a type I interferon (IFN) gene signature in patients with SLE, and the IFN-regulated proteins PRKRA, IFITM1 and CD69 (p<0.0001) were found to be up-regulated in platelets from SLE patients as compared to healthy volunteers. Notably, patients with a history of vascular disease had increased expression of type I IFN-regulated proteins as well as more activated platelets as compared with patients without vascular disease. We suggest that interferogenic immune complexes stimulate production of IFNα which up-regulates the megakaryocytic type I IFN-regulated genes and proteins. This could affect platelet activation and contribute to development of vascular disease in SLE. In addition, platelets with type I IFN signature could be a novel marker for vascular disease in SLE. Platelets were purified from SLE patients (n=10) and age and sex-matched healthy volunteers (n=10). cDNA was generated for each individual. For the microarray analysis, cDNA from the 10 SLE patients were pooled, as well as the cDNA from the 10 healthy volunteers to represent a mean mRNA expression level from 10 individuals.
Project description:Understanding the underlying mechanisms of the well-established platelet hyporeactivity in neonates, would be of great relevance for both improving the clinical management of neonates, a population with a higher bleeding risk than adults (especially among sick and preterm infants), and getting new insights onto the regulatory mechanisms of platelet biology. Transcriptome analysis is a useful tool to identify mRNA signature affecting platelet function. However, human fetal/neonatal platelet transcriptome analysis has never been reported. Here, we used, for the first time, mRNA expression array to compare the platelet transcriptome changes during development. Microarray analysis was performed in pure platelet RNA obtained from adult and cord blood, using the same platform in two independent laboratories. A high correlation was obtained between arrays results for both adult and neonate platelet samples. There was also a good agreement between our adult results and those previously reported in three different studies. Gene enrichment analysis demonstrated that immunity- and platelet function-related genes are highly expressed in either developmental stage. Remarkably, 201 genes were found to be differentially expressed along development. In particular, neonatal platelets contain higher levels of mRNA that are associated with protein synthesis and processing, while they carry significantly lower levels of genes related with calcium transport/metabolism and cell signaling (including GNAZ). Overall, our results highlight that variations in platelet transcriptome may underline the hypo-functional phenotype of neonatal platelets, and further support the role of platelets in cellular immune response. A better characterization of the platelet transcriptome across development may help to elucidate the implications of transcriptome changes in different pathological conditions.