ABSTRACT
Objective To study the effect of siRNA-interfered endogenous Kir6.2 gene deletion on rotenone-induced cell damage and signal pathway according to the expression of siRNA-interfered endogenous Kir6.2 gene.Methods The expression of endogenous Kir6.2 gene in PC12 cells was detected by RT-PCR and Western bolt.The viability of PC12 cells was tested using WST-1.The expression of PKC and phosphorylated PKC in PC12 cells was detected by Western blot in negative control group,siRNA/Kir6.2 group,siRNA/Kir6.2 + rotenone group,siRNA/Kir6.2+ rotenone+PKC inhibitor group before and afer treatment with rotenone.Results RT-PCR showed that the interference was successful in siRNA/Kir6.2/group 1 and siRNA/Kir6.2/ group 2.Western blot displayed that the expression level of endogenous Kir 6.2 gene was significantly lower in siRNA/Kir6.2/ group 1 than in negative control group (0.55±0.07 vs 0.89±0.09,P＜ 0.05).WST-1 revealed that the viability of PC12 cells was significantly lower in siRNA/Kir6.2 group than in negative control group (P＜0.05).Western blot demonstrated that the expression of PKC and phosphorylated PKC was significantly lower in siRNA/Kir6.2+-rotenone+PKC inhibitor group than in the other three group (P＜0.05).Conclusion Endogenous Kir6.2 can protect PC12 cells against the toxicity of rotenone and plays an important role in regulating the rotenone-induced viability of PC12 cells by activating the phosphorylated PKC.
ABSTRACT
Objective To study the effect of siRNA-interfered endogenous Kir6.2 gene deletion on rotenone-induced cell damage and signal pathway according to the expression of siRNA-interfered endogenous Kir6.2 gene.Methods The expression of endogenous Kir6.2 gene in PC12 cells was detected by RT-PCR and Western bolt.The viability of PC12 cells was tested using WST-1.The expression of PKC and phosphorylated PKC in PC12 cells was detected by Western blot in negative control group,siRNA/Kir6.2 group,siRNA/Kir6.2 + rotenone group,siRNA/Kir6.2+ rotenone+PKC inhibitor group before and afer treatment with rotenone.Results RT-PCR showed that the interference was successful in siRNA/Kir6.2/group 1 and siRNA/Kir6.2/ group 2.Western blot displayed that the expression level of endogenous Kir 6.2 gene was significantly lower in siRNA/Kir6.2/ group 1 than in negative control group (0.55±0.07 vs 0.89±0.09,P＜ 0.05).WST-1 revealed that the viability of PC12 cells was significantly lower in siRNA/Kir6.2 group than in negative control group (P＜0.05).Western blot demonstrated that the expression of PKC and phosphorylated PKC was significantly lower in siRNA/Kir6.2+-rotenone+PKC inhibitor group than in the other three group (P＜0.05).Conclusion Endogenous Kir6.2 can protect PC12 cells against the toxicity of rotenone and plays an important role in regulating the rotenone-induced viability of PC12 cells by activating the phosphorylated PKC.