Effects Of Homocysteine Induced The Apoptosis And Expression Of Caspase3, C-IAP2 Of Endothelial Progenitor Cells

Objective 1. To isolate single mononuclear cells from healthy adults'bone marrow, using inducing factor containing culture fluid to cause them to differentiate into endothelial progenitor cells (EPC), and to primary culture the EPC and identify them.2. The present study addresses the question how Hcy induces endothelial apoptosis and how folic acid antagonizes the proapoptotic effects of Hcy. therefore providing theoretical evidence for further exploring the relationship between EPC and diabetic angiopathy.Methods 1. Mononuclear cells from healthy volunteers'bone marrow were obtained by density gradient centrifugation and incubated into culture bottles (plates) coated with human fibronectin. After cultured in vitro, the adherent cells were collected. EPC were characterized as adherent cells which were double positive for Dil-acLDL uptake and FITC-UEA., and their surface markers CD 133 and CD34 were detected by applying flow cytometry to make a further identification of the primary cultured cells are EPCs.2.Endothelial progenitor cells (EPC) were treated with Hcy, with or without folic acid, for 48 hours. Cell apoptosis was evaluated by Annexin V staining and flow cytometery. The mRNA levels of caspase3 and c-IAP2 were analyzed by RT-PCR.Results Treatment with both low (0.3mmol/L)and high (3.0mmol/L) concentrations of Hcy induced EPC apoptosis, with higher apoptotic rate at the latter case.Hcy-induced EPC apoptosis was accompanied by an increased level of caspase3 expression, together with decreased c-IAP2 level. Folic acid upregulated c-IAP2 expression.Conclusions 1. We successfully isolated single mononuclear cell from human bone marrow and induced them to EPCs, which were identified and subcultured afterwords.2. Hcy may induce EPC apoptosis via a pathway involving caspase3, which can be partially antagonized by folic acid, possibly through upregulated c-IAP2 expression.