Connexin 43 Prevents the Progression of Diabetic Renal Tubulointerstitial Fibrosis by Regulating the SIRT1-HIF-1α Signaling Pathway

Abstract

Background: Diabetic renal fibrosis is a combination of glomerular sclerosis and renal tubulointerstitial fibrosis (RIF). Our previous studies have shown that connexin 43 (Cx43) activation could inhibit the development of diabetic renal glomerular sclerosis. Renal epithelial to mesenchymal transition (EMT) is a key pathological factor in diabetic RIF. Whether Cx43 affect the EMT and the underlying mechanism remains to be elucidated. Methods: The effects of Cx43 on EMT were assessed by transfections of plasmids and short hairpin RNAs respectively in NRK-52E cells and caudal vein injection of Cx43 adenovirus in the db/db spontaneous diabetic model mice. The downstream factors were screened through western blot, Immunofluorescence, co-immunoprecipitation, EMSA assay and dual luciferase reporter assay. Findings: Cx43 protein expression was decreased in the kidney tissues of db/db mice and in NRK-52E cells cultured in HG. Cx43 overexpression inhibited the HG-induced EMT process and reduced the expression of the extracellular matrix (ECM) components. Depletion of Cx43 further promoted the EMT process and increased those ECM components. Cx43, in a carboxyl terminal signal transduction-dependent manner, could up-regulate SIRT1 expression and enhance SIRT1-dependent deacetylation of HIF-1α to reduce HIF-1α activity, which eventually ameliorated diabetic RIF. Caudal vein injection of Cx43 adenovirus alleviated renal injury in db/db spontaneous diabetic model mice, accompanied by increased SIRT1 levels, decreased HIF-1α expression, and reduced production of EMT markers and ECM components. Interpretation: Cx43 ameliorates renal EMT and diabetic RIF by regulating the SIRT1-HIF-1α signaling pathway, which provides an experimental basis for Cx43 as a potential target of diabetic nephropathy. Funding Statement: This work was supported by research grants from the National Natural Foundation of China (No. 81603168, No. 81573477, No. 81770816, No.81973375), the Key Projects of Guangdong Natural Science Foundation (No. 2017A030311036), Guangdong Provincial Key Laboratory of Construction Foundation (No. 2017B030314030), and the Natural Science Foundation of Guangdong Province, China (No. 2017A030313678). Declaration of Interests: The authors have declared that no conflicts of interest exist. Ethics Approval Statement: All animal care and experimental procedures complied with the Guide for the Care and Use of Laboratory Animals (NIH Publication No. 85-23, revised 1996), in line with the China Animal Welfare Legislation, and were approved by the Ethics Committee on the Care and Use of Laboratory Animals of Sun Yat-sen University, Guangzhou, Guangdong, China.