Objective:To investigate the effects of nitrogen permease regulator-like-2 (NPRL2) on autophagy in human colon cancer HT29 cells, and further explore the relationship between autophagy and apoptosis. Methods:The lentiviral vector expressing the NPRL2 was constructed and transfected into HT29 cells. Western blot was used to detect the expressions of NPRL2, LC3B and p62 protein in HT29 cells after 48 h transfection and acridine orange staining were performed to observe the autophagy under a fluorescence microscope. The transfected HT29 cells were treated with autophagy inhibitor 3-methyladenine(3-MA), cell proliferation was measured by CCK-8, cell death was detected by Trypan blue, and caspase-3 activity was measured by caspase-3 activity kit. Cell apoptosis was measured by Annexin V-FITC/PI kit. Western blot was used to detect the protein expressions of active caspase-3, Bax, Bcl-2, LC3B and p62. Results:Western blot results showed that NPRL2 gene was successfully overexpressed in HT29 cells. Western blot results also revealed that NPRL2 can activate HT29 autophagy. And acridine orange staining were performed to observe the autophagy. Our published results showed that NPRL2 could promote the apoptosis of HT29 cells. To further investigate the relationship between NPRL2-activated autophagy and NPRL2-induced apoptosis, 3-MA was to treat used HT29 cells. The results showed that 3-MA inhibited the autophagy induced by NPRL2, and further reduced the cell viability of HT29 cell line, inhibited its proliferation, promoted its apoptosis, promoted the expressions of active caspase 3 and Bax, inhibited the expression of Bcl-2 and attenuated its autophagy. Conclusion:NPRL2 can promote the autophagy of colon cancer HT29 cell line, but this autophagy inhibits the apoptosis caused by NPRL2, and the inhibition of autophagy induced NPRL2 by 3-MA can effectively promote the apoptosis of colon cancer cells.
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