Background:The nucleotide-binding and oligomerization domain-like receptor protein 3 (NLRP3) inflammasome composed of NLRP3,apoptosis-associated speck-like protein containing CARD (ASC),and caspase-1 is engaged in the...Background:The nucleotide-binding and oligomerization domain-like receptor protein 3 (NLRP3) inflammasome composed of NLRP3,apoptosis-associated speck-like protein containing CARD (ASC),and caspase-1 is engaged in the inflammatory response of many kidney diseases and can be activated by purinergic 2X7 receptor (P2X7R).This study was conducted to explore whether P2X7R plays a pathogenic role in the podocyte damage of obesity-related glomerulopathy (ORG) and whether this role is mediated by the activation ofNLRP3 inflammasome.Methods:A mouse model of ORG was established by high-fat diet feeding.The conditionally immortalized mouse podocytes were cultured with leptin or with leptin and P2X7R antagonist (KN-62 or A438079).The mRNA and protein expression of the P2X7R and NLRP3 inflammasome components including NLRP3,ASC,and caspase-1,as well as the podocyte-associated molecules including nephrin,podocin,and desmin in mouse renal cortex or cultured mouse podocytes were tested by real-time-polymerase chain reaction and Westem blot analysis,respectively.Results:The significantly upregulated expression of P2X7R and NLRP3 inflammasome components and the NLRP3 inflammasome activation were observed in the renal cortex (in fact their location in podocytes was proved by confocal microscopy) of ORG mice in vivo,which were accompanied with the morphological changes of podocyte damage and the expression changes of podocyte-associated molecules.Similar changes in the expression of P2X7R and NLRP3 inflammasome components as well as in the expression ofpodocyte-associated molecules were also observed in the cultured podocyte studies treated by leptin in vitro,and all of the above changes were significantly attenuated by the P2X7R antagonist KN-62 or A438079.Conclusions:P2X7R could trigger the activation ofNLRP3 inflammasome,and the activated P2X7R/NLRP3 inflammasome in podocytes might be involved in the podocyte damage of ORG.展开更多
AIM To validate the effects of receptor interacting protein kinase-3(RIP3) deletion in non-alcoholic fatty liver disease(NAFLD) and to clarify the mechanism of action.METHODS Wild-type(WT) and RIP3 knockout(KO) mice w...AIM To validate the effects of receptor interacting protein kinase-3(RIP3) deletion in non-alcoholic fatty liver disease(NAFLD) and to clarify the mechanism of action.METHODS Wild-type(WT) and RIP3 knockout(KO) mice werefed normal chow and high fat(HF) diets for 12 wk. The body weight was assessed once weekly. After 12 wk, the liver and serum samples were extracted. The liver tissue expression levels of RIP3, microsomal triglyceride transfer protein, protein disulfide isomerase, apolipoprotein-B, X-box binding protein-1, sterol regulatory element-binding protein-1c, fatty acid synthase, cluster of differentiation-36, diglyceride acyltransferase, peroxisome proliferator-activated receptor alpha, tumor necrosis factor-alpha(TNF-α), and interleukin-6 were assessed. Oleic acid treated primary hepatocytes from WT and RIP3 KO mice were stained with Nile red. The expression of inflammatory cytokines, including chemokine(C-X-C motif) ligand(CXCL) 1, CXCL2, and TNF-α, in monocytes was evaluated.RESULTS RIP3 KO HF diet fed mice showed a significant gain in body weight, and liver weight, liver to body weight ratio, and liver triglycerides were increased in HF diet fed RIP3 KO mice compared to HF diet fed WT mice. RIP3 KO primary hepatocytes also had increased intracellular fat droplets compared to WT primary hepatocytes after oleic acid treatment. RIP3 overexpression decreased hepatic fat content. Quantitative real-time polymerase chain reaction analysis showed that the expression of very-low-density lipoproteins secretion markers(microsomal triglyceride transfer protein, protein disulfide isomerase, and apolipoprotein-B) was significantly suppressed in RIP3 KO mice. The overall NAFLD Activity Score was the same between WT and RIP3 KO mice; however, RIP3 KO mice had increased fatty change and decreased lobular inflammation compared to WT mice. Inflammatory signals(CXCL1/2, TNF-α, and interleukin-6) increased after lipopolysaccharide and pancaspase inhibitor(necroptotic condition) treatment in monocytes. Neutrophil chemokines(CXCL1, and CXCL2) were decreased, and TNF-α was increased after RIP3 inhibitor treatment in monocytes.CONCLUSION RIP3 deletion exacerbates steatosis, and partially inhibits inflammation in the HF diet induced NAFLD model.展开更多
基金grants from the National Natural Science Foundation of China (No.81573745and No.8160140274)Beijing Municipal Natural Science Foundation (No.7172066) Beijing Development Foundation of Traditional Chinese Medicine (QN2016-23).
文摘Background:The nucleotide-binding and oligomerization domain-like receptor protein 3 (NLRP3) inflammasome composed of NLRP3,apoptosis-associated speck-like protein containing CARD (ASC),and caspase-1 is engaged in the inflammatory response of many kidney diseases and can be activated by purinergic 2X7 receptor (P2X7R).This study was conducted to explore whether P2X7R plays a pathogenic role in the podocyte damage of obesity-related glomerulopathy (ORG) and whether this role is mediated by the activation ofNLRP3 inflammasome.Methods:A mouse model of ORG was established by high-fat diet feeding.The conditionally immortalized mouse podocytes were cultured with leptin or with leptin and P2X7R antagonist (KN-62 or A438079).The mRNA and protein expression of the P2X7R and NLRP3 inflammasome components including NLRP3,ASC,and caspase-1,as well as the podocyte-associated molecules including nephrin,podocin,and desmin in mouse renal cortex or cultured mouse podocytes were tested by real-time-polymerase chain reaction and Westem blot analysis,respectively.Results:The significantly upregulated expression of P2X7R and NLRP3 inflammasome components and the NLRP3 inflammasome activation were observed in the renal cortex (in fact their location in podocytes was proved by confocal microscopy) of ORG mice in vivo,which were accompanied with the morphological changes of podocyte damage and the expression changes of podocyte-associated molecules.Similar changes in the expression of P2X7R and NLRP3 inflammasome components as well as in the expression ofpodocyte-associated molecules were also observed in the cultured podocyte studies treated by leptin in vitro,and all of the above changes were significantly attenuated by the P2X7R antagonist KN-62 or A438079.Conclusions:P2X7R could trigger the activation ofNLRP3 inflammasome,and the activated P2X7R/NLRP3 inflammasome in podocytes might be involved in the podocyte damage of ORG.
基金Supported by National Research Foundation of Korea(NRF)funded by the South Korean Government,No.NRF-2017M3A9C8028794
文摘AIM To validate the effects of receptor interacting protein kinase-3(RIP3) deletion in non-alcoholic fatty liver disease(NAFLD) and to clarify the mechanism of action.METHODS Wild-type(WT) and RIP3 knockout(KO) mice werefed normal chow and high fat(HF) diets for 12 wk. The body weight was assessed once weekly. After 12 wk, the liver and serum samples were extracted. The liver tissue expression levels of RIP3, microsomal triglyceride transfer protein, protein disulfide isomerase, apolipoprotein-B, X-box binding protein-1, sterol regulatory element-binding protein-1c, fatty acid synthase, cluster of differentiation-36, diglyceride acyltransferase, peroxisome proliferator-activated receptor alpha, tumor necrosis factor-alpha(TNF-α), and interleukin-6 were assessed. Oleic acid treated primary hepatocytes from WT and RIP3 KO mice were stained with Nile red. The expression of inflammatory cytokines, including chemokine(C-X-C motif) ligand(CXCL) 1, CXCL2, and TNF-α, in monocytes was evaluated.RESULTS RIP3 KO HF diet fed mice showed a significant gain in body weight, and liver weight, liver to body weight ratio, and liver triglycerides were increased in HF diet fed RIP3 KO mice compared to HF diet fed WT mice. RIP3 KO primary hepatocytes also had increased intracellular fat droplets compared to WT primary hepatocytes after oleic acid treatment. RIP3 overexpression decreased hepatic fat content. Quantitative real-time polymerase chain reaction analysis showed that the expression of very-low-density lipoproteins secretion markers(microsomal triglyceride transfer protein, protein disulfide isomerase, and apolipoprotein-B) was significantly suppressed in RIP3 KO mice. The overall NAFLD Activity Score was the same between WT and RIP3 KO mice; however, RIP3 KO mice had increased fatty change and decreased lobular inflammation compared to WT mice. Inflammatory signals(CXCL1/2, TNF-α, and interleukin-6) increased after lipopolysaccharide and pancaspase inhibitor(necroptotic condition) treatment in monocytes. Neutrophil chemokines(CXCL1, and CXCL2) were decreased, and TNF-α was increased after RIP3 inhibitor treatment in monocytes.CONCLUSION RIP3 deletion exacerbates steatosis, and partially inhibits inflammation in the HF diet induced NAFLD model.
