Alteration of apoptotic protease activating factor 1 expression and possible role in ONOO(-)-induced apoptosis in human cerebral vascular smooth muscle cells.

The study was to investigate the change of apoptotic protease activating factor 1 (Apaf1) expression in the human cerebral vascular smooth muscle cells (HCVSMCs) in response to peroxynitrite (ONOO(-)). HCVSMCs were cultured in vitro and ONOO(-) of different concentrations was directly added to the culture medium. The total proteins were extracted and the alteration of Apaf1 expression was examined by the means of Western-blot. Apaf-1 siRNA (h) was put into another plate of HCVSMCs for transfection. The transfected cells were also incubated in different concentrations of ONOO(-). The change of Apaf1 protein expression after siRNA transfection was examined by the means of Western-blot. The morphological changes were observed by acridine orange staining to determine whether cells experienced apoptosis in response to ONOO(-) before and after siRNA. The Flow cytometry analysis was used to examine the change of cells apoptotic rates in response to ONOO(-) before and after siRNA. Obviously, there was up-regulated Apaf1 expression at protein level in the course of HCVSMCs apoptosis induced by ONOO(-). When Apaf1 expression was suppressed, the apoptotic sum of HCVSMCs didn't change. This study demonstrates that Apaf1 gene is involved in ONOO(-)-induced apoptosis in HCVSMCs. Whether HCVSMCs treated by ONOO(-) undergo apoptosis depends on Apaf1 level.