Sang-Bai-Pi extract and its constituent regiafuran C ameliorate renal fibrosis through TGF-β/Smad and Wnt/β-catenin signaling pathways

Background: Renal fibrosis is a major pathological feature of many chronic kidney diseases, and traditional Chinese medicines (TCM) have shown promising therapeutic potential for treating renal fibrosis. Although the extracts or fractions of Morus alba leaves and twigs have been reported to ameliorate renal fibrosis, the beneficial effects of M. alba root bark (commonly known as Sang-Bai-Pi), a well-known TCM, on this disorder have not been investigated.

Purpose: This study aims to investigate the effects and mechanisms of Sang-Bai-Pi extract/fractions and their constituents on renal fibrosis.

Methods: Immunoblotting was first used to assess the effects of different Sang-Bai-Pi fractions on key fibrotic markers. The most potent fraction, EA-3, was further evaluated using a unilateral ureteral obstruction (UUO) rat model to assess its antifibrotic effects. The efficacy of EA-3 was evaluated by analyzing the pathomorphological changes in the kidney tissues of rats using histological staining and by detecting the expression of relevant proteins via Western blotting. The transforming growth factor-β1 (TGF-β1)-stimulated human renal proximal tubular cell line HK-2 was used to elucidate the likely modes of action of EA-3 and its constituent regiafuran C (RFC). Network pharmacology and molecular docking analyses were utilized to explore the detailed molecular mechanism of RFC.

Results: Fraction EA3 effectively alleviated UUO-induced renal fibrosis in rats. Mechanistically, EA-3 suppressed the epithelial-mesenchymal transition, accumulation of extracellular matrix, and activation of the TGF-β/Smad and Wnt/β-catenin signaling pathways in vitro and in vivo. RFC also demonstrated antifibrotic potential by inhibiting TGF-β/Smad and Wnt/β-catenin signaling in HK-2 cells. Further investigations revealed that RFC inhibited TGF-β/Smad pathway by blocking the interaction of Smad3 with TGF-βRII and TGF-βRI might be a potential direct target of RFC.

Conclusion: The antirenal fibrotic effects of Sang-Bai-Pi extract/fractions and the constituent RFC were evaluated for the first time. Fraction EA-3 and RFC alleviated renal fibrosis by targeting the TGF-β/Smad and Wnt/β-catenin pathways. These findings provide valuable insights into the development of Sang-Bai-Pi-based phytotherapy and new drug molecules for the safe treatment of renal diseases.