Autophagic pathway and probable mechanism in degradation of mutant α-synuclein in PC12 cells / 中华神经科杂志

ABSTRACT

Objective To observe the effect of mutant α-synuclein(A30P)in autophagic programmed cell death by transfected PC12 cells and explore its probable role and pathway in PD.Methods The definite PC12 cells which were transfected mutant α-synuclein(A30P)were constructed at first and MPP+,Rapamycin and Wortmanin were administrated to transfected PC12 cells with mutant α-synuclein. Not only the proliferative activity of cells was detected with MTT method but also the ultrastructttre changes of cells and expression of α-synuclein in different circumstance were observed by transmission electron microscopy(TEM),Western Blot and the level of SOD.Results (1)The expression of α-synuclein in groups A30P+Wortmannin and A30P+MPP+was higher than that in group A30P(P＜0.01), particularly.there was more significant expression of α-synuclein in group A30P+Wortmannin.The expression of α-synuclein in group A30P+Rapamycin was weaker than that in group A30P(P＜0.01); (2)The results showed that the SOD level(group A30P+MPP+3 h97.49±13.8;12 h102.7±12.7; 24 h101.5±11.8;48 h104.3±12.4)was significantly decreased at various time points after MPP+ treatment compared that of group A30P(t=3.7721,P=0.0017).SOD level gradually increased in A30P +Rapamycin 12 h and showed significant difference at 24 h(121.2±13.0),48 h(124.3±14.1)and 72 h(127.7±13.7)after drug treatment compared with that in group A30P+Wortmannin(t2.9746, P=0.0083);(3)Mutant α-synuclein(A30P)leading to PC12 cells death by means of autophagy involved α-synuclein accumulation,membrane lipid oxidation,and loss of plasma membrane integrity.Mutant α- synuclein(A30P)mediated the toxicity of MPP+.Rapamycin,an inducer of autophagy,reduced the aggregation of α-synuclein in transfected cells.Meanwhile,Wortmanin,an inhibitor of autophagy,promoted the aggregation of α-synuclein in transfected cells and induced cells to die.Conclusions The abnormal aggregation of α-synuclein induces autophagic programmed cell death in PC12 cells and mutant α-synuclein (A30P)mediates the toxicity of MPP+.Meanwhile,Rapamycin may reduce the aggregation of α-synuclein in transfeeted cells by activation of autophagic pathway.