Yishen-Qingli-Huoxue formula attenuates renal fibrosis by inhibiting indoxyl sulfate via AhR/snai1 signaling

Chronic kidney disease (CKD) is challenging to reverse and its treatment options are limited. Yishen-Qingli-Huoxue Formula (YQHF) is an effective treatment Chinese formula for CKD, as verified by clinical randomized controlled trial. However, the correlative YQHF therapeutic mechanisms are still unknown.The current study aimed to investigate the potential anti-renal fibrosis effects of YQHF as well as the underlying mechanism.After affirming the curative effects of YQHF on adenine-induced CKD rats, Masson staining, immunohistochemistry, and ELISA were used to assess the effects of YQHF on renal fibrosis. Subsequently, metabolomics and transcriptomics analyses were conducted to clarify the potential mechanisms. Furthermore, high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), molecular docking analysis and in vitro experiments were used to verify final mechanism of anti-fibrosis.Our results demonstrated that YQHF could improve renal morphology, decrease blood urea nitrogen (BUN), serum creatinine (Scr), and increase body weight gain of model rats. Masson staining, immunohistochemistry of collagen I, fibronectin (FN), α-smooth muscle actin (α-SMA), vimentin and E-cadherin showed that YQHF delayed CKD progression by alleviating renal fibrosis, and the expression of fibrotic factors smoc2 and cdh11 were obviously suppressed by YQHF. Metabolomic and transcriptomic measures discovered that indoxyl sulfate might be a crucial factor inducing renal fibrosis, and the antagonistic effect of YQHF on renal fibrosis may be exerted via AhR/snai1 signaling. Subsequently, western blot and immunohistochemical experiments revealed YQHF indeed inhibited AhR/snai1 signaling in adenine-induced renal fibrosis of CKD rat, which confirmed previous results. In addition, molecular docking and in vitro experiments further supported this conclusion, in which astilbin, the main compound identified YQHF, was certified to exert a significant effect on AhR.Our findings showed that YQHF can effectively treat CKD by antagonizing renal fibrosis, the potential mechanisms were relating with the regulation on AhR/snai1 signaling.