Several data suggest that the inflammasome in infiltrating macrophages and renal dendritic cells promotes renal inflammation. However, the role of each inflammasome component (NLRP3, buy Everolimus ASC, and Caspase-1) in renal tubular cells remains unclear. We demonstrated that renal collecting duct (CD) epithelial cell was the major site of
ASC expression in mouse unilateral ureteral obstruction (UUO) model. The role of ASC in renal inflammation and fibrosis was investigated in vivo and in vitro. Methods: C57BL/6J-background wild-type (WT) and ASC-knockout (ASC-KO) mice underwent UUO. Primary mouse CD epithelial cells were used in in vitro study. Results: Expression of mRNA for inflammasome components, NLRP3, ASC, and Caspase-1 as well as IL-1β was time-dependently increased in the ligated kidney after UUO. ASC-KO showed significant amelioration in tubulointerstitial injury and fibrosis histologically. Flow cytometric analysis showed that increase in CD45+ leukocytes
was remarkably suppressed in ASC-KO, indicating that inflammatory response was ameliorated in ASC deficiency. Pregnenolone Immunohistochemistry showed ASC was upregulated in a portion Ibrutinib of renal tubules after UUO. Double immunofluorescence analysis showed that most ASC positive tubules were co-stained with aquaporin-2 (AQP2), collecting duct marker. In in vitro study, we identified the constitutive expression of NLRP3, ASC and Caspase-1 in primary mouse CD epithelial cells using western blotting. After extracellular ATP stimulation, active Caspase-1 and mature
form of IL-1β secretion were observed. CD epithelial cells from ASC-KO mice did not show the response to ATP. ATP-induced IL-1β release was inhibited significantly by P2X7R antagonist, blocking K+ efflux, or antioxidant N-acetyl cysteine (NAC). Conclusion: ASC was upregulated in CD epithelial cells after UUO and contributed to inflammation and fibrosis. Extracellular ATP stimulated IL-1β release in CD epithelial cells ASC-dependently. This inflammasome activation was mediated through P2X7R-potassium efflux and ROS-dependent pathways. These findings suggest that potential immunological role of CD by inflammasome activation.