Molecular Mechanism Of Autophagy Against Salinomycin Anti-tumor Effect Through The ATG3/Akt/mTOR Signaling Pathway

Objective: To clarify the salinomycin-induced SGC-7901 cells autophagy,and to explore the role of autophagy in salinomycin anti-tumor and its possible molecular mechanism.Methods: 1,SGC-7901 cells were treated with different concentrations of salinomycin for 24 hours,by MTT,flow cytometry,fluorescent spots experiments,immunofluorescence and Western Blot,Clear that salinomycin induced cell autophagy in SGC-7901 cells.2,To establish lentivirus-mediated autophagy key gene ATG3 overexpression and sh RNA interference SGC-7901 cell line,by flow cytometry and Western Blot,to explore salinomycin-induced autophagy resistant to the anti-tumor effect of salinomycin probably through the ATG3/AKT/m TOR signaling pathway.3,SGC-7901 cells were treated with autophagy inhibitor 3-methyladenosine(3-methyladenine,3-MA)and ATG5/ATG7 sh RNA interference lentivirus respectively,byflow cytometry and Western Blot,to investigate the role of autophagy in salinomycin anti-tumor.Results: 1,Salinomycin treatment SGC-7901 cells can inhibit cell proliferation and promote apoptosis;Increase the number of exogenous GFP-LC3 fluorescent spots and endogenous LC3 fluorescent spots;while down-regulating the expression of ATG3 protein and activating the AKT/m TOR signaling pathway.The differences were statistically significant.2,SGC-7901 stable cell line with ATG3 overexpression and sh RNA interference was successfully established.In ATG3 overexpression SGC7901 cells,ATG3 overexpression can activate the AKT/m TOR signaling pathway;After treatment with salinomycin,ATG3 overexpression can attenuate salinomycin-induced LC3-II protein levels,while enhancing salinomycin-induced apoptosis;the differences were statistically significant.In ATG3-interfered SGC7901 cells,ATG3 interference can reduce salinomycin-induced apoptosis,the difference was statistically significant.3,SGC-7901 cells were treated with 3-MA,inhibition of autophagy can promote salinomycin-induced apoptosis;while down-regulating the protein levels of Beclin-1,LC3-II and caspase 3 and Up-regulating the protein level of p62 and c-caspase 3;the differences were statistically significant.The SGC-7901 cell line that was interfered by ATG5-sh RNA and ATG7-sh RNA was successfully established.In SGC-7901 cell that was interfered by ATG5-sh RNA and ATG7-sh RNA,inhibition of autophagy can attenuate salinomycin-induced LC3-II protein expression,while increasing the protein level of c-Caspase3,the differences were statistically significant.Conclusions: 1,Salinomycin can induce autophagy in gastric cancer SGC-7901 cells.2,Autophagy resist the anti-tumor effects of salinomycin probably through the ATG3 / AKT / m TOR signaling pathway.3,inhibition of autophagy enhance the anti-tumor effects of salinomycin.