MPP~+Down-regulates Activation Of Autophagy In The PD Cell Model Possibly Via MTOR Pathway

Objective1. To observe the influence of different concentrations of MPP+and different timepoints on the survival rate of PC12cells,and searching for appropriate range ofconcentration and time for cellular model of Parkinson’s diseases;2. To observe the influence of MPP+on PC12cells survival rate、LDH leakage rateand the expression level of LC3-II,Beclin1involving in autophagy-lysosomal pathway,andview the phenomenon of RAPA protected PC12cells against MPP+-induced injury;3. To observe the influence of MPP+on PC12the expression level of mTOR(mammals rapamycin target protein) and other related protein in its signaling pathway, andview the effect of RAPA.MethodsSelect the appropriate concentration and treat time of MPP+, according to the effect ofdifferent concentration and time of MPP+on PC12cell activity, to make cellular model ofParkinson ’s diseases;And then giving the intervention of RAPA: The cells viability wasanalyzed by MTT assay; The cells injury was assessed by LDH assay; Western blot used tostudy autophagy mechanism of PD model by analyzing expression level of LC3-II,Beclin1,mTOR, Raptor,Rictor,p-p70s6k and p-4E-BP1.Results1. The relative survival rate of PC12cell showed a trend of decrease with MPP+concentration gradient. And cell relative survival rate decreased significantly (P<0.05～0.001) with MPP+during0.25~3.0mM; The relative survival rate of PC12showed atrend of decrease with the effect time of MPP+, and cell relative survival rate decreasedsignificantly (P<0.05～0.001) during8~48h;2. After24h, MPP+medium treatment resulted in significantly lower survival rate(P<0.001), the supernatant of LDH leakage was significantly higher (P<0.001), and the expression of LC3-II was inhibited significantly (P<0.01), the expression of Beclin1wasalso inhibited significantly (P<0.01), RAPA effectively improved the cell relative survivalrate (P<0.001), reduced LDH leakage rate (P<0.001), significantly increased theexpression of LC3-II (P<0.01) and the expression of Beclin1(P<0.01);3. After24h, MPP+medium treatment significantly increased the expression ofrelated protein in mTOR signaling pathway, while RAPA intervention coulddown-regulate mTOR signaling pathway.Conclusions1MPP+could down-regulate activation of autophagy of PC12cells and result in cellinjury, while RAPA showed antagonistic effect.2MPP+could down-regulate activation of Autophagy in the PD cell model possiblyvia mTOR pathway, while RAPA conld protect PC12cells by inhibiting mTOR Pathway.