The Preliminary Study Of Survivin On Related Genes Ragulation In The Formation Process Of Autophagy-lysosome In Eca-109 Cells

Objective: To study the regulatory mechanism of Survivin on autophagy and autophagic lysosome formation in cancer cells.Methods: The autophagy induction model was established with Rapamycin(RAPA),Survivin(SVV)overexpression plasmid,SVVshRNA plasmid and Survivin inhibitor YM155 were used to intervene in the autophagy model.The morphology of autophagy and the status of autophagy were observed by transmission electron microscope(SEM)and confocal laser scanning microscopy;Western blot was used to detect the expression changes of genes involved in the formation of autophagy.Results: 1)After RAPA induction in Eca-109 cells,increased autophage activities could be seen under transmission electron microscope;The increased level of LC3 expression in cells was observed by confocal laser scanning microscopy;Western blot results showed that after RAPA induced,LC-I and LC-II protein expression increased and LC-I/LC-II ratio decreased,the expression of p62 decreased.2)After overexpression of Survivin,the expression of LC3 and Rab7increased(P<0.05),the expression of ATG5 decreased(P<0.05),and the change of ATG16L1 was not obvious.3)The expression of Survivin and LC3 increased when Survivin over expressed(P<0.05),while the expression of Rab7,ATG5 and ATG16L1decreased(P < 0.05).After YM155 and SVVshRNA interfered in Eca-109 cells,the expression of p62,ATG5 and ATG16L1 was decreased(P<0.05).4.The distribution of Cathepsin-D in cells was analyzed by immunofluorescence staining.Cathepsin-D in cells remained diffuse after overexpression of Survivin.Conclusion: Rapamycin was used to induce and establish the autophagy model.RAPA increase the expression of Rab7,ATG5,and ATG16L1,while the increase of expression of ATG5 and ATG16L1 are dependent on the existence of SVV;SVV/RAPA’s induction of autophagy of tumor cells is a concerted regulation of certain target genes at specific times,which occurrs at the earlystage formation of autophagy and the late stage functioning of autophagic lysosome.ATG5/ATG16L1,p62 is the target of regulation for SVV during autophagy formations,which increase the expression of ATG5/ATG16L1 at the early stage of autophagy formation to promote the process and inhibit the degradation of lysosome;Overexpression of SVV may change the pH condition and membrane permissibility of lysosome,which leads to affection of functioning of lysosome.