Fig. 2

Expression of the NLRP3 inflammasome and markers of tubular injury are reduced in the UUO-Cx43^−/−mouse. Analysis of a published transcriptomic dataset shows that NLRP3 expression increases in kidneys with CKD (n = 48 patients) as compared to healthy controls (n = 5) (a) and that this positively correlates with proteinuria (b) in people with diabetic kidney disease (DKD), the leading cause of end stage renal failure in which tubulointerstitial fibrosis is the key underlying pathology. With evidence that Cx43 and NLRP3 are upregulated in CKD, combined with data that the heterozygous Cx43^+/− UUO mouse exhibits improved renal structure and diminished inflammation, we utilised our Pax8-rtTA-cre:cx43 flox Cx43^−/− model to determine if Cx43 plays a role in regulating expression of NLRP3 and principal mediators of inflammation. Quantitative PCR (qPCR) analysis and immunostaining of the renal cortex showed a marked downregulation of NLRP3 messenger RNA (mRNA) (c) and protein expression (d) in UUO-Cx43^−/− mice as compared with WT UUO. Similarly, deletion of Cx43 from the proximal tubules prior to induction of UUO protected against an upregulation of IL1β (e), IL6 (f) and downstream NLRP3 inflammatory mediators, monocyte chemoattractant protein (MCP1) (g) and injury marker neutrophil gelatinase-associated lipocalin (N-GAL) (h). An ANOVA and Tukey post-test were used for experimental comparisons except transcriptomic data where an unpaired t-test with Welch’s correction &/or a simple linear regression and Pearson’s correlation analysis was used. *P < 0.05, **P < 0.01, ***P < 0.001 and ****P < 0.0001