Overexpression of smad7 inhibits the TGF-β/Smad signaling pathway and EMT in NPHP1-defective MDCK cells

作者：Wu, Xiaohong; Wang, Haiyan; Chen, Huamu; Lin, Hongrong; Li, Min; Yue, Zhihui; Sun, Liangzhong^*

来源：Biochemical and Biophysical Research Communications, 2021, 582: 57-63.

DOI：10.1016/j.bbrc.2021.10.037

• Nephronophthisis
• TGF-beta
• Fibrosis
• NPHP1
• EMT

Background: Nephronophthisis (NPHP) is a kind of ciliopathy. Interstitial fibrosis occurs at the early stage of the disease. TGF-beta/Smad is a key signaling pathway in regulating interstitial fibrosis and epithelial-mesenchymal transition (EMT). In this study, we explored the activation of the TGF-beta/Smad signaling pathway and EMT in NPHP1-defective MDCK cells to further understand the pathogenesis of NPHP. @@@ Methods: NPHP1-knockdown (NPHP1(KD)) MDCK cells were constructed by recombinant lentiviral short hairpin RNA, and NPHP1-knockout (NPHP1(KO)) MDCK cells were constructed by using the CRISPR/Cas9 technique. The morphology and migration ability were observed under a microscope. Western blotting was used to detect the expression of E-cadherin, beta-catenin, alpha-smooth muscle actin (alpha-SMA), fibroblast-specific protein-1(FSP1), TGF-beta 1, Smad2, Smad3, p-Smad3, Smad4 and Smad7. The localization of Smad3 was determined by immunofluorescence assay. @@@ Results: NPHP1(KD) and NPHP1(KO) MDCK cells were spindle-shaped and presented EMT-like changes. E-cadherin and beta-catenin expression decreased, while alpha-SMA and FSP1 expression increased; the TGF-beta/Smad signaling pathway was activated, Smad2, Smad3, p-Smad3 and Smad4 expression increased, Smad3 translocated to nuclear and Smad7 expression decreased compared with those in wild type MDCK cells. Overexpression of Smad7 reversed these changes to different degrees. @@@ Conclusions: Our results indicate that NPHP1 defects induce the activation of the TGF-beta/Smad signaling pathway and EMT in MDCK cells. These factors may be implicated in the pathogenesis of interstitial fibrosis in NPHP.

• 单位
  中山大学; 1; 南方医科大学