Effects Of Heat Shock Protein 70 On Membrane Calcium Homeostasis Of PC12 Cells Induced By Ischemic

Objective To investigate the effects and manchanisms of Lentivirus-mediated HSP70 gene on voltage-gated calcium channels on the cell membrane of PC12 cells induced by ischemic and hypoxia.Methods PC12 cells were subjected ischemie/hypoxia for 8h and reoxygenation for 12 h stimulating the nerve cells suffered ischemia/reperfusion pathological process in vivo. PC12 cells were divided into non-infection group, lentivirus control group [infected by lentivirus containing green fluorescent protein(GFP) without HSP70 gene] and recombined lentiviral infection group(infected by lentivirus containing HSP70 and GFP gene). Western-blot and RT-PCR assay were performed to evaluate the effects of HSP70 on the expression of cav1.2、cav1.3、NR1、NR2 and NCX, the key subunits of LCC and NMDA in PC12 cells.Result(1)After being challenged with ischemia and hypoxia, an decreased level of cav1.2, NR1 and NR2 m RNA and protein in the PC12 cells was showed in recombined lentiviral infection group notably, compared with ones of non-infection group and lentivirus control group[cav1.2 m RNA(A value): 3.13±0.46 vs.5.12±0.52、5.13±0.66, NR1 m RNA(A value): 1.61±0.44 vs.3.23±0.82、3.31±0.78, NR2 m RNA(A value): 2.09±0.41 vs.3.91±0.64、3.88±0.62, cav1.2 protein(gray value): 2.82±0.39 vs.3.98±0.23、3.96±0.24, NR1 protein(gray value): 84±0.35 vs.2.79±0.21、2.86±0.23, NR2 protein(gray value): 0.87±0.24 vs.1.57±0.31、1.33±0.44, all P<0.01]; But there was no statistical difference in the expression of cav1.2,NR1 and NR2 m RNA and protein between lentivirus control group and non-infection group(all P>0.01).(2) There was no statistical difference in the expression of cav1.3 and NCX m RNA and protein among non-infection group、lentivirus control group and recombined lentiviral infection group [cav1.3m RNA(A value): 4.82±0.29、4.72±0.36、4.82±0.32, NCXm RNA(A value): 3.49±0.78、3.47±0.71、3.56±0.65, cav1.3 protein(gray value): 2.63±0.40、2.64±0.39、2.68±0.39, NCX protein(gray value): 3.27±0.48、3.34±0.48、3.31±0.42, all P>0.01].Conclusion Exogenous HSP70 affects the expression of cav1.2、NR1 and NR2, the key subunits of LCC and NMDA, which may protects PC12 cells from the injury caused by ischemic and hypoxia.