The Mechanism Of Inflammasome NLRP6 Regulating P53 On Proliferation,Autophagy And Immune Escape Of Colorectal Cancer

Objective:To investigate the mechanism of inflammasome NLRP6 regulating p53 on proliferation,autophagy and immune escape of colorectal cancer.Methods:Lentivirus was used to knockdown NLRP6 in HCT116 p53+/+,HCT116 p53-/and SW620 colorectal cancer cells;NLRP6 knockdown efficiency was measured by Real-time PCR,Western blot,and immunofluorescence;The effect of NLRP6 on the cell proliferation was determined by CCK-8 assay;Immunofluorescence was performed to measure the location of NLRP6 and LC3B in cells,as well as autophagosome formation;The expression of p53,PDL1,LC3B,NF-κB and MDM2 was determined by Western blot;CD8+T cells were isolated by magnetic sorting for in vitro expansion and culture,and purity of CD8+T cells was confirmed by flow cytometry.;CD8+T cells were co-cultured with colorectal cancer cells,and the killing efficiency was measured by lactate dehydrogenase assay.Results:The stable transfected shNLRP6 cell lines were successfully established,and the NLRP6 knockdown efficiency is more than 70%at gene level.NLRP6 can suppress proliferation in p53 wild type,null and mutant colorectal cancer cells.NLRP6 and LC3B are mainly expressed in the cytosol.NLRP6 inhibits autophagosome formation in p53 wild type and promotes autophagosome formation in p53 null and mutant colorectal cancer cells.The CD8+T cells toxic efficiency was decreased in p53 wild type and increased in p53 null or mutant colorectal cancer cells with NLRP6 knockdown.Conclusion:1.NLRP6 can inhibit the proliferation of p53 wild type,null and mutant colorectal cancer cells.2.NLRP6 can affect colorectal cancer autophagy and immune escape through both p53 dependent and p53 independent pathways.NLRP6 can suppress colorectal cancer autophagy and immune escape through p53 dependent pathways in p53 wild type cell line,and NLRP6 promotes colorectal cancer autophagy and immune escape through p53 independent pathways in p53 null or mutant cell lines.