1,25(OH)2D3 provides protection against diabetic kidney disease by downregulating the TLR4-MyD88-NF-κB pathway

Abstract

The over-activation of Toll-like receptors (TLRs) is a typical immune response to injury. Previous work has suggested that controlling the over-activation of TLR4-MyD88-NF-κB may represent a new therapeutic option for diabetic kidney disease (DKD). 1,25(OH)₂D₃ has also been shown to exert a protective effect on DKD, although the mechanism involved has yet to be elucidated. The aim of this study was to investigate whether 1,25(OH)₂D₃ protects against DKD by down-regulating the innate immune TLR-NF-κB pathway. NRK-52E cells were cultured under normal or high-glucose conditions. We then used siRNA to knock down TLR4 expression under high-glucose conditions. NRK-52E cells cultured under high-glucose conditions, and streptozotocin (STZ)-induced diabetic rats, were treated with different doses of 1,25(OH)₂D₃ and used as in vitro and in vivo models, respectively. Renal biochemical indicators were then measured to evaluate the influence of 1,25(OH)₂D₃ treatment on DKD in diabetic rats. Histological analysis was also performed to determine the extent of infiltration by inflammatory cells and tubulointerstitial fibrosis. Using RT-qPCR, western blotting, immunohistochemistry and immunofluorescence, we determined the expression levels of TLR4, MyD88, NF-κB p65, MCP-1 and α-SMA to investigate whether 1,25(OH)₂D₃ could reduce the development of tubulointerstitial fibrosis. Knocking down TLR4 abolished the tubulointerstitial fibrosis caused by high-glucose conditions. High doses of 1,25(OH)₂D₃ consistently reduced the expression of TLR4-MyD88-NF-κB in NRK-52E cells. Moreover, high doses of 1,25(OH)₂D₃ had an obvious protective effect on kidney injury and inhibited the infiltration of inflammatory cells and tubulointerstitial fibrosis in diabetic rats. In conclusion, high doses of 1,25(OH)₂D₃ protected against tubulointerstitial fibrosis both in vitro and in vivo by downregulating the expression of TLR4-MyD88-NF-κB.

Keywords: 1,25(OH)(2)D(3); Diabetic kidney disease; TLR4-MyD88-NF-κB; Tubulointerstitial fibrosis.