Effects Of MTOR Signaling Pathway In The Regulation Of Manganese Induced Apoptosis In MN9D Cells By Rapamycin

Objective: The mouse midbrain dopaminergic neuron(MN9D)was used as a model to investigate the role of rapamycin in the regulation of manganese-induced apoptosis of MN9 D cells and its possible mechanism.To explore the mTOR signaling pathway regulating Mn-induced MN9 D cells in rapamycin The role of apoptosis and its possible mechanisms provide the basis for prevention and treatment of manganese poisoning in occupational populations.Methods: 1.MN9 D cells were exposed to 1.2 and 2.4 m M manganese 24 h after pretreatment with rapamycin(0.1?g/ml)for 1 h.Then the morphology of the cells was observed.the number of viable cells was counted by trypan blue staining.the apoptosis rate was measured by flow cytometry after staining with Annexin V/PI double staining.and the activation of caspase-3 in MN9 D cells was analyzed by Western blot.2.MN9 D cells were exposed to 1.2 m M manganese for 24 h after 1 h of rapamycin RAP(0.1?g/ml)/mTOR agonist MHY(1?M).Then.the cell morphology was observed.The number of viable cells was counted by trypan blue staining.The apoptotic rate was detected by flow cytometry after Annexin V/PI double staining.The activation of caspase-3 and the expression of mTOR related protein in MN9 D cells were analyzed by western blot.Results: 1.Compared with the control group.manganese induced a decrease in MN9 D cell activity and morphological changes in a concentration-dependent manner.The apoptotic rate gradually increased.and the expression of cleaved-caspase-3 protein also increased gradually.The difference was statistically significant(P<0.05).Compared with the manganese exposure group.rapamycin pretreatment significantly reduced the manganeseinduced MN9 D cell activity and apoptosis.and blocked the manganese-induced caspase-3 activation.The difference was statistically significant(P<0.05).2.Compared with the control group.MN9 D cells in Mn-alone group decreased activity and morphological changes of apoptotic cells.the difference was statistically significant(P<0.05);cleaved-caspase-3 protein expression increased.the difference was statistically significant(P<0.05);AKT and S6K1?4E-BP1 protein expression also increased.the difference was statistically significant(P<0.05).Compared with the Mn-treated group.rapamycin pretreatment attenuated the expression of cleaved-caspase-3 protein(P<0.05).and weakened the protein expression of AKT and S6K1 and 4E-BP1.and the difference was statistically significant(P<0.05).Compared with the Mn-treated group.MHY pretreatment aggravated apoptosis and increased the expression of cleaved-caspase-3 protein(P<0.05);S6K1 and 4E-BP1 protein expression increased.the difference was statistically significant(P<0.05);AKT protein expression decreased.the difference was statistically significant(P<0.05).Conclusion: 1.Rapamycin has obvious resistance to manganese-induced apoptosis in MN9 D cells.2.Manganese can induce apoptosis of MN9 D cells through the mTOR signaling pathway.3.Rapamycin can attenuate manganese induced apoptosis of MN9 D cells via mTOR signaling pathway.