Biochanin A Attenuates Renal Fibrosis Via Selective Inhibition of Smad3-mediated Epithelial-Mesenchymal Transition in a Murine Unilateral Ureteral Obstruction Model.

Abstract

Renal fibrosis driven by TGF-β/Smad3 signaling is a critical factor in the advancement of chronic kidney disease, and partial epithelial-to-mesenchymal transition (EMT) in tubular cells plays a significant role in this process. In this study, we explored the reno-protective role of biochanin A (BCA), a natural isoflavone, in mice with unilateral ureter obstruction (UUO) and in cultured renal tubular TCMK1 cells stimulated with the profibrotic cytokine TGF-β1. Our results demonstrated that BCA (20 and 50 mg/kg) dose-dependently ameliorated structural damage in UUO kidneys with reduced extracellular matrix deposition and downregulated levels of fibrotic genes Col1a1, fibronectin, and α-SMA. In vitro, BCA (5 and 10 μg/mL) also exhibited a dose-dependent anti-fibrosis effect in TCMK1 cells induced by TGF-β1. Moreover, BCA prevented tubular partial EMT by mitigating the upregulation of mesenchymal marker genes N-cadherin and vimentin in UUO kidneys and TGF-β1-induced TCMK1 cells. BCA treatment also reversed the reduced staining of Lotus Tetragonolobus Lectin and peanut agglutinin, two markers of mature renal tubular epithelia, in UUO kidneys. Mechanistically, BCA inhibited Smad2 and Smad3 phosphorylation and specifically downregulated Smad3 protein expression, without affecting Smad2, in TGF-β1-stimulated TCMK1 cells. Most importantly, in TGF-β1-treated TCMK1 cells, Smad3 overexpression abolished the anti-fibrosis and anti-EMT effects of BCA. In conclusion, our findings demonstrate the potential of BCA as a therapeutic agent against renal fibrosis by inhibiting TGF-β/Smad3 signaling-mediated tubular partial EMT.