Norcantharidin protects against renal interstitial fibrosis by suppressing TWEAK-mediated Smad3 phosphorylation

This study investigated the role and mechanism of action of tumor necrosis factor-like weak inducer of apoptosis (TWEAK) in the pathogenesis of renal interstitial fibrosis (RIF), and its involvement in the anti-RIF effect of norcantharidin (NCTD).

Mice with unilateral ureteral obstruction and BUMPT mouse proximal tubular cells exposed to transforming growth factor (TGF)-β1 were used as in vivo and in vitro models of RIF, respectively. NCTD was administered to mice by intraperitoneal injection (0.075 mg kg^-1·day^-1). Hematoxylin-eosin and Masson's trichrome staining were performed to assess pathologic changes in the kidney. Immunohistochemistry, western blotting, and real-time PCR were performed to evaluate the expression of TWEAK and the fibrotic factors fibronectin (FN) and collagen type I (Col-I). The role of TWEAK in RIF and in the anti-RIF effect of NCTD was evaluated by TWEAK overexpression and neutralization with a specific antibody, and specific inhibitor of Mothers against decapentaplegic homolog (Smad)3 (SIS3) was used to examine the involvement of TGF-β1/Smad3 signaling.

TWEAK was mainly expressed in renal tubules in mice; the level was markedly elevated in both in vivo and in vitro RIF models. TWEAK overexpression in BUMPT cells increased the levels of phosphorylated Smad3, FN, and Col-I, which were reduced by treatment with SIS3. NCTD suppressed FN and Col-I expression by blocking TWEAK-mediated Smad3 phosphorylation.

Upregulation of TWEAK contributes to RIF by promoting Smad3 phosphorylation, while NCTD inhibits this process.