title: Effects of endothelial microvesicles induced by A23187 on H9c2 cardiomyocytes.
abstract: OBJECTIVE
  To investigate the effects of endothelial microvesicles (EMVs) induced by calcium ionophore A23187 on H9c2 cardiomyocytes.
  
  
  METHODS
  Human umbilical vein endothelial cells (HUVECs) were treated with 10 micromol/L A23187 for 30 min. EMVs from HUVECs were isolated by ultracentrifugation from the conditioned culture medium. EMVs were characterized using 1 and 2 microm latex beads and anti-PE-CD144 antibody by flow cytometry. For functional research, EMVs at different concentrations were cocultured with H9c2 cardiomyocytes for 6 h. Cell viability of H9c2 cells and the activity of LDH leaked from H9c2 cells were tested by colorimetry. Moreover, apoptosis of H9c2 cells was observed through Hoechst 33258 staining and tested by FITC-Annexin V/PI double staining.
  
  
  RESULTS
  EMVs were induced by A23187 on HUVECs, and isolated by ultracentrifugation. We identified the membrane vesicles (< 1 microm) induced by A23187 were CD144 positive. In addition, the EMVs could significantly reduce the viability of H9c2 cells, and increase LDH leakage from H9c2 cells in a dose dependent manner (P < 0.05). Condensed nuclei could be observed with the increasing concentrations of EMVs through Hoechst 33258 staining. Furthermore, increased apoptosis rates of H9c2 cells could be assessed through FITC-Annexin V/PI double staining by flow cytometry.
  
  
  CONCLUSIONS
  Microvesicles could be released from HUVECs after induced by A23187 through calcium influx, and these EMVs exerted a pro-apoptotic effect on H9c2 cells by induction of apoptosis.
authors.forename: Man
authors.initials: M
authors.lastname: Shang
authors.forename: Qi
authors.initials: Q
authors.lastname: Zhang
authors.forename: Meng-Xiao
authors.initials: MX
authors.lastname: Zhang
authors.forename: Yao
authors.initials: Y
authors.lastname: Wang
authors.forename: Yan
authors.initials: Y
authors.lastname: Chen
authors.forename: Yan-Na
authors.initials: YN
authors.lastname: Wu
authors.forename: Jun-Qiu
authors.initials: JQ
authors.lastname: Song
authors.forename: Ming-Lin
authors.initials: ML
authors.lastname: Liu
authors.forename: Yan-Xia
authors.initials: YX
authors.lastname: Liu
mesh_terms: D017304:Annexin A5; D017209:Apoptosis; D000001:Calcimycin / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects*; D018920:Coculture Techniques; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D032383:Myocytes, Cardiac / Q000187:drug effects*; D013194:Staining and Labeling
chemical_list: D017304:Annexin A5; C492760:FITC-annexin A5; D000001:Calcimycin; D016650:Fluorescein-5-isothiocyanate; D002118:Calcium