The Microtube-targeting Agents Induced Autophagy/autophagic Cell Death And Its Relationship With Apoptotic Sensibility In Human Gastric Cancer Cells

Background & Objective: The microtube-targeting agents such as Paliataxel and Vincristine, can induce apoptosis in many malignant tumor cells. However, the sensitivities of distinct cancer cells to such agents are quite different. Recent studies of autophagy, which is an evolutionarily conserved mechanism of protein and organelle degradation, suggest that this mechanism of proteolysis may affect the regulation of cancer cells' survival and death. On the one hand, autophagy may lead to the programmed cell death of certain form, such as autophagic cell death and apoptosis. On the other hand, autophagy can also be a protective mechanism that prevents cancer cells from being damage to lethal levels. Now, little is known about the possible role of autophagy /autophagic cell death on cancer cells, especially on gastric cells during the treatment of microtube-targeting agents. In this paper, we study the existence of autophagy/autophagic cell death and their effects on gastric cancer cells in the treatment of microtube-targeting agents, as well as the relationship between the autophagy and the apoptosis. The conclusion of the paper may help us form a new viewpoint of promoting the cancer cells sensitivity to such agents.Method: Apoptosis rates are measured by Annexin V/PI kit by FACS analysis and cell death rates are determined by MTT assay. Using fluorescent microscope, the non-apoptosis cell death is observed after DAPI staining. Also after the staining of acriding orange, autophagy and autoghagic cell death were detected using flow cytometry and fluorescent microscope. We employ immunoblotting assay to detect the expressions of Beclin 1. The relationship between autophagy and apoptosis induced by microtube-targeting agents is also studied by autophagy inhibitor 3-MA or caspase inhibitor Z-VAD-FMK co-treatment.Results: 1.Taxol and VCR both can induce apoptosis in gastric cancer cells and the apoptotic rates are dependent of dose and inducing-time. SGC-7901 cells are more sensitive to the apoptosis induced by the agents than BGC-823 cells. The difference between the rates of apoptosis and cell death rates, together with the results of DAPI staining in dead cells implies the existence of non-apoptosis cell death induced by Taxol or VCR. The results of acriding orange staining and Beclin 1 expressions prove the non-apoptosis cell death is of the kind of autoghagic cell death. 2. Taxol and VCR both can induce autophagy of gastric cancer cells in early moment. However, the perk of autohagy induced by Taxol is 3-6 hours which is a contrast to that of VCR, which is 24-36 hours. BGC-823 cells are more sensitive to the autophagy induced by such agents than SGC-7901 cells. 3-MA co-treament with Taxol can promote Taxol induced apoptosis, while Z-VAD-FMK co-treatment has no influence on Taxol induced autophagy in gastric cancer cells.Conclusion: Microtube-targeting agents, Taxol and VCR, can induce both apoptosis and autophagy/autoghagic cell death in gastric cancer cells. The autophagy/autoghagic cell death, which relates to the up-expression of Beclin 1, occurs before the apoptosis induced by Taxol or VCR, and even inhibits the occurring of apoptosis.