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RIPK3 blockade attenuates tubulointerstitial fibrosis in a mouse model of diabetic nephropathy.

ABSTRACT: Receptor-interacting protein kinase-3 (RIPK3) is a multifunctional regulator of cell death and inflammation. RIPK3 controls cellular signalling through the formation of the domain-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome, which is recognised to mediate renal fibrogenesis. The role of RIPK3 in diabetic kidney disease (DKD) induced renal fibrosis has not been previously determined. To define the action of RIPK3 in the development of diabetic kidney disease, wild-type (WT), RIPK3 -/- and endothelium-derived nitric oxide synthase (eNOS)-/- mice were induced to develop diabetes mellitus using multiple low doses of streptozotocin and maintained for 24 weeks. RIPK3 activity and NLRP3 expression were upregulated and fibrotic responses were increased in the kidney cortex of WT mice with established diabetic nephropathy compared to control mice. Consistently, mRNA expression of inflammasome components, as well as transforming growth factor beta 1 (TGF?1), ? smooth muscle actin (?-SMA) and collagen deposition were increased in diabetic kidneys of WT mice compared to control mice. However, these markers were normalised or significantly reversed in kidneys of diabetic RIPK3 -/- mice. Renoprotection was also observed using the RIPK3 inhibitor dabrafenib in eNOS-/- diabetic mice as demonstrated by reduced collagen deposition and myofibroblast activation. These results suggest that RIPK3 is associated with the development of renal fibrosis in DKD due to the activation of the NLRP3 inflammasome. Inhibition of RIPK3 results in renoprotection. Thus, RIPK3 may be a potential target for therapeutic intervention in patients with diabetic kidney disease.

SUBMITTER: Shi Y

PROVIDER: S-EPMC7319952 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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RIPK3 blockade attenuates tubulointerstitial fibrosis in a mouse model of diabetic nephropathy.

Shi Ying Y Huang Chunling C Zhao Yongli Y Cao Qinghua Q Yi Hao H Chen Xinming X Pollock Carol C

Scientific reports 20200626 1

Receptor-interacting protein kinase-3 (RIPK3) is a multifunctional regulator of cell death and inflammation. RIPK3 controls cellular signalling through the formation of the domain-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome, which is recognised to mediate renal fibrogenesis. The role of RIPK3 in diabetic kidney disease (DKD) induced renal fibrosis has not been previously determined. To define the action of RIPK3 in the development of diabetic kidney disease, wild-type (WT ...[more]

PMID: 32591618

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Project description:BackgroundRenal tubulointerstitial fibrosis is the hallmark of various chronic kidney diseases. Symmetric dimethylarginine (SDMA) is an independent cardiovascular risk factor in patients with chronic kidney diseases, which is mostly excreted through renal tubules. However, the effect of SDMA on kidneys in a pathological condition is currently unknown. In this study, we investigated the role of SDMA in renal tubulointerstitial fibrosis and explored its underlying mechanisms.MethodsMouse unilateral ureteral obstruction (UUO) and unilateral ischemia-reperfusion injury (UIRI) models were established to study renal tubulointerstitial fibrosis. SDMA was injected into kidneys through ureter retrogradely. TGF-β stimulated human renal epithelial (HK2) cells were used as an in vitro model and treated with SDMA. Signal transducer and activator of transcription-4 (STAT4) was inhibited by berbamine dihydrochloride or siRNA or overexpressed by plasmids in vitro. Masson staining and Western blotting were performed to evaluate renal fibrosis. Quantitative PCR was performed to validate findings derived from RNA sequencing analysis.ResultsWe observed that SDMA (from 0.01 to 10 µM) dose-dependently inhibited the expression of pro-fibrotic markers in TGF-β stimulated HK2 cells. Intrarenal administration of SDMA (2.5 µmol/kg or 25 µmol/kg) dose-dependently attenuated renal fibrosis in UUO kidneys. A significant increase in SDMA concentration (from 19.5 to 117.7 nmol/g, p < 0.001) in mouse kidneys was observed after renal injection which was assessed by LC-MS/MS. We further showed that intrarenal administration of SDMA attenuated renal fibrosis in UIRI induced mouse fibrotic kidneys. Through RNA sequencing analysis, we found that the expression of STAT4 was reduced by SDMA in UUO kidneys, which was further confirmed by quantitative PCR and Western blotting analysis in mouse fibrotic kidneys and renal cells. Inhibition of STAT4 by berbamine dihydrochloride (0.3 mg/ml or 3.3 mg/ml) or siRNA reduced the expression of pro-fibrotic markers in TGF-β stimulated HK2 cells. Furthermore, blockage of STAT4 attenuated the anti-fibrotic effect of SDMA in TGF-β stimulated HK2 cells. Conversely, overexpression of STAT4 reversed the anti-fibrotic effect of SDMA in TGF-β stimulated HK2 cells.ConclusionTaken together, our study indicates that renal SDMA ameliorates renal tubulointerstitial fibrosis through inhibition of STAT4.

Dataset Information

RIPK3 blockade attenuates tubulointerstitial fibrosis in a mouse model of diabetic nephropathy.

Publications

RIPK3 blockade attenuates tubulointerstitial fibrosis in a mouse model of diabetic nephropathy.

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