Role of tumor necrosis factor-α in epithelial-to-mesenchymal transition in transplanted kidney cells in recipients with chronic allograft dysfunction

Background: Chronic allograft dysfunction (CAD) is characterized by allograft kidney interstitial fibrosis, the underlying mechanism of which is unclear. Our aim was to elucidate the role and mechanism of TNF-alpha-induced epithelial-to-mesenchymal transition (EMT) in transplant kidney tubular interstitial fibrosis. Methods: Human kidney tissues from normal volunteers and CAD patients were assessed using periodic acid Schiff, Masson trichrome and immunohistochemical staining. mRNA and protein expression of E-cadherin, a smooth muscle actin (SMA) and fibronectin(FN) in renal proximal tubule epithelial (HK-2) cells after treatment with TNF-alpha under different conditions were assessed using western blot and qRT-PCR analysis. Cell motility and migration were assessed using wound healing and transwell assays. Expression of Smurf2 and TNF-alpha-signaling pathway -related proteins in HK-2 cells treated with TNF-alpha was detected by western blotting. E-cadherin and alpha-SMA expression was also assessed in Smurf2 plasmid-transfected or Smurf2 siRNA-treated HK-2 cells. Results: The expression of TNF-alpha, Smurf2, alpha-SMA, and fibronectin was significantly upregulated, while the expression of E-cad was downregulated in the CAD group compared with the normal group. The in vitro results showed that TNF-alpha remarkably upregulated the expression of Smurf2, a-SMA and fibronectin and down regulated the expression of E-cadherin in HK-2 cells and enhanced motility and migration in HK-2 cells. Overexpression of Smurf2 could promote the expression of alpha-SMA and inhibit the expression of E-cad, whereas knockdown of Smurf2 expression reversed TNF-alpha-induced upregulation of alpha-SMA and prohibited the reduction of E-cad expression. Furthermore, TNF-alpha-induced Smurf2 expression promoted EMT through the Akt signaling pathway. Conclusions: TNF-alpha induced EMT via the TNF-alpha/Akt/Smurf2 signaling pathways, and it may play a role in aggravating allograft kidney interstitial fibrosis in CAD patients.