Inhibition Of Autophagy Activity Enhances The Effect Of Proliferation And Apoptosis In Colon Cancer Cell SW620

Objective: Through the study on the effect of autophagy inhibitor combined with Ce6-PDT on colorectal cancer SW620 cell proliferation and apoptosis,to make clear inhibition of tumor cell autophagy could enhance the inhibition rate and apoptosis rate of colorectal cancer cells induced by Ce6-PDT,to provide theoretical and practical basis for autophagy inhibitors improve sensitivity of Ce6-PDT treatment of colorectal cancer.Methods: 1.Using laser scanning confocal microscope combined with organelle probe to observe the subcellular localization of photosensitizer Ce6 in SW620 colon cancer cell line.2.The morphological and structural changes in Ce6-PDT cells were observed by fluorescence microscopy.3.Using MTT to detect the cell viability in Ce6-PDT and 3MA-Ce6-PDT.4.Using nnexinV-PE/7-ADD combined with flow cytometry to detect cellular apoptosis in Ce6-PDT and 3MA-Ce6-PDT.5.Acridine orange(AO)staining combined with fluorescence microscopy to observe the situation of autophagy in Ce6-PDT and 3MA-Ce6-PDT.6.Western blot was used to detect the expression of apoptosis related proteins Caspase-3,Bcl-2 and the expression of autophagy related protein LC3II/I in Ce6-PDT and 3MA-Ce6-PDT.Results: 1.Ce6 mainly localized in the endoplasmic reticulum,followed by the mitochondria and lysosomes,and almost no distribution in the nucleus.2.Ce6-PDT can lead to the normal morphology and structural changes of SW620 cells.3.After Ce6-PDT,cell viability was negatively correlated with Ce6 concentration(r=-0.820,P<0.01).4.After Ce6-PDT,celluar apoptosis rate was positively correlated with the concentration of Ce6(r=0.845,P<0.01).5.After Ce6-PDT,the expression of apoptosis related protein caspase-3,Bcl-2 and the ratio of autophagy related protein LC3II/I increased with Ce6 concentration(P<0.05).6.Compared to the cells in Ce6-PDT,cell viability decreased(P<0.05),celluar apoptosis rate increased(P<0.05),mitochondrial membrane potential decreased(P<0.05),the expression of autophagy related protein decreased(P<0.05)and the expression of apoptosis related protein increased(P<0.05)in 3MA-Ce6-PDT.Conclusions: 1.Ce6 was mainly localized in the endoplasmic reticulum and mitochondria of SW620 cells,which can induce endoplasmic reticulum and mitochondria stress,and finally induce celluar apoptosis and autophagy 2.After Ce6-PDT,with the increase of Ce6 concentration,the normal morphology of the cells changed obviously,and the typical morphological features of apoptosis was observed.3.Ce6-PDT can inhibit the proliferation and induce cell apoptosis in SW620 colorectal cancer cells.4.Ce6-PDT increased the expression of apoptosis and autophagy related proteins,further indicated that it could induce cell apoptosis and autophagy.5.3MA combined with Ce6-PDT lead to an increase in the rate of apoptosis and the decrease of cell viability and mitochondrial membrane potential.Moreover,the expression of apoptosis related proteins was increased,these results suggest that autophagy may play an important role in protecting SW620 cells from apoptosis induced by Ce6-PDT in human colon cancer cells.