The Role Of NLRP6 Inflammasome On Salmonella Typhimurium Infection In Adipocytes

BackgroundStudies have shown that fat cells play an important role in fighting off bacterial infections,but the mechanism is unclear.Inflammasome is a multi-protein complex assembled by intracytoplasmic pattern recognition receptors,which can recruit and activate the pro-inflammatory cysteine aspartate proteolytic enzyme pro-caspase1.Activated Caspase1 cleaves IL-1? and IL-18 into mature cytokines and induces the occurrence of cell pyrotosis.In our study,inflammasome receptor molecule NLRP6 was found to have the highest expression level in adipocytes.Since the role of NLRP6 in adipocytes infected by bacteria is not clear,we mainly studied the role of NLRP6 in adipocytes infected by bacteria.ObjectiveBy studying the changes in the expression of NLRP6 and the activation of Caspase1 in adipocytes after LTA stimulation or S.typhimurium infection,we can provide a theoretical basis for the role of adipocytes in S.typhimurium infection.Methods1.The preadipocytes 3T3-L1 were differentiated into mature adipocytes,and the expressions of different inflammasome receptor molecules in adipocytes were detected by q PCR after the mature adipocytes were treated with S.typhimurium at different concentrations.2.3T3-L1 cells were differentiated into mature adipocytes and treated with NLRP6 ligand LTA.The expression of NLRP6 and its downstream signaling inflammatory cytokines were detected by q PCR,Western blot and ELISA.3.The expression of NLRP6 in adipocytes infected with S.typhimurium at different concentrations was directly proportional to the number of bacterial infections,and the expression of Caspase1 and IL-1? downstream signaling molecules were significantly increased.4.NLRP6 overexpression plasmid was constructed and transfected into 3T3-L1 differentiated mature adipocytes.Cells were treated with different concentrations of wild S.typhimurium.The expressions of NLRP6,ASC,Caspase1,IL-1? and IL-18 were detected by q PCR,Western blot and ELISA.And measured changes in the number of bacteria entering the cells and how the cells died.5.Primary gonadal adipocytes from NLRP6 knockout mice and WT mice were isolated and treated with wild S.typhimurium.The expression of NLRP6,ASC,Caspase1,IL-1? and IL-18 were detected by q PCR,Western blot and ELISA,and the changes in the number of bacteria entering the cells and the way of cell death were detected.Results1.After treatment of mature adipocytes with different concentrations of S.typhimurium,the expression of inflammasome receptors was increased,among which the expression of NLRP6 was the most significant.2.After LTA stimulation of adipocytes,the expression of NLRP6 and its downstream inflammatory factors ASC,caspase1,IL-1?,and IL-18 increased significantly,and the expression of caspase1 and IL-1? showed a dose-dependent relationship with LTA.3.The expression of NLRP6 in adipocytes infected with S.typhimurium at different concentrations was directly proportional to the number of bacterial infections,and the expression of Caspase1 and IL-1? downstream signaling molecules were significantly increased.4.After the overexpression of NLRP6,mature adipocytes were infected with S.typhimurium,the number of bacteria entering the cells was significantly reduced,and the expression of downstream signaling molecules was significantly increased.5.After NLRP6 was knocked out,mature adipocytes were infected with S.typhimurium and the number of bacteria entering the cells was significantly increased.Conclusions1.S.typhimurium will highly express NLRP6 and some other inflammasome recept or molecules after infecting fat cells.2.LTA is recognized by NLRP6 and causes the expression of downstream signal molecules to increase.3.S.typhimurium activates NLRP6 in fat cells and plays an important role in resistin g S.typhimurium infection.