DMDD, Isolated from the Root of <i> Averrhoa carambola</i> L., Protects Against Diabetic Kidney Disease by Inhibiting TLR4/TGFβ Signaling Pathway

Abstract

Objective: Diabetic kidney disease (DKD) is the leading cause of death and disability of diabetes mellitus. However, there is still a lack of specific drugs for the treatment of DKD. Objective: The chief aim of this research is to investigate the role and mechanism of DMDD for DKD. Methods: Wild type and TLR4 knockout mice were induced to diabetes. After 4-week treatment with DMDD, blood sugar, renal function, blood lipid and pathological changes were assessed. Real-time PCR, western blotting, and immunohistochemistry were employed to detect the expressions of TLR4, TGFβ1 and Smad2/3 in the renal tissue. Results: DMDD improved the serum lipid and decreased fasting blood glucose levels in diabetic mice. CysC and urinary albumin levels increased markedly in the diabetic group, and they were obviously decreased after 4 weeks of DMDD treatment. Compared with the WT diabetic mice, the urinary albumin and CysC in the TLR4-/- mice were expressed at lower levels. HE and Masson's staining revealed that DMDD clearly ameliorated pathological changes and renal fibrosis. When TLR4 gene was knock out, the pathological was improved. Mechanistically, TLR4, TGF-β1 and Smad2/3 were obvious up-regulation in the renal tissues of diabetic mice. The expressions of these proteins were significantly down-regulated after DMDD treatment (p< 0.05). In the TLR4-/- mice, mRNA and protein levels of TGF-β1 and Smad2/3 were obviously lower than those in the WT mice. In addition, IHC revealed that a strong in situ expressions of TLR4, TGF-β1 and Smad2/3 were seen in the kidney tissues of diabetic mice, which were distinctly weakened in the DMDD-treated mice. In the TLR4-/- mice, however, expressions of TGF-β1 and Smad2/3 were not remarkable increase in the diabetic mice compared with normal mice. Conclusions: These results strongly indicate that TLR4 is essential for DMDD protection against renal dysfunction in diabetic mice. Its hypoglycemic and anti-fibrosis effects were likely mediated by inhibiting of the TLR4/TGFβ signaling pathway, and the downregulation of Smads in the renal interstitium. Funding Statement: This research was supported by the National Natural Science Foundation of China (No. 81760665, 81460205), and Guangxi Natural Science Foundation under Grant No. 2017GXNSFBA198177. In addition, this project was financially supported by the project of improving the basic ability of young teachers in colleges and universities in Guangxi (KY2016YB098). Declaration of Interests: The authors declare no conflicts of interest in this work. Ethics Approval Statement: All experimental procedures were approved by the Institutional Animal Care and Use Committee of Guangxi Medical University.