Effect of Mitochondrial Fission Factor on Mitochondrial Apoptosis Pathway in OGD/R-stimulated PC12 Cells
Objective To explore the expression level and effects of mitochondrial fission factor(MFF)on mitochondrial function and apoptosis of MFF in oxygen glucose deprivation/re-oxygenation(OGD/R)-stimulated PC12 cells.Methods PC12 cells were used to establish ischemic neuron injury model in vitro.First,quantitative real time-PCR(qRT-PCR)and Western blot assays were performed to determine the mRNA and protein levels of MFF at different time points after OGD/R treatment.Then,MFF was knocked down by RNA interference technology,the mRNA and protein levels were measured by qRT-PCR and Western blot assay.Cell counting kit-8(CCK-8)and TdT-mediated dUTP nick end labeling(TUNEL)assays were used to determine the effect of MFF silencing on cell survival.The levels of mitochondrial membrane potential,adenosine triphosphate(ATP)and reactive oxygen species(ROS)were detected by JC-1 and corresponding kits,respectively.The cellular localization of cytochrome C(Cyt-C)and protein levels of Bax and Bc12 were quantified by Western blotting.The activities of Caspase-3 and Caspase-9 were assessed by using enzyme linked immunosorbent assay(ELISA)assay.Results Both the mRNA and protein levels of MFF in OGD/R-stimulated PC12 cells were significantly increased.Further experimental data showed that silence of MFF obviously enhanced the cell viability and inhibited cell apoptosis in OGD/R-induced PC12 cells,along with in-creased Bcl2 protein level and reduced Bax protein level.After MFF knockdown,mitochondrial membrane potential and ATP synthesis were greatly elevated,while ROS generation,Cyt-C release and the activities of Caspase-3 and Caspase-9 were reduced in OGD/R-stimulated PC12 cells.Conclusion Silencing of MFF increases cell survival and improves mitochondrial function.
NETL
NSTL
万方数据
