In chronic renal diseases, progressive loss of renal function correlates with advancing tubulo-interstitial fibrosis. TGFβ1-Smad (transforming growth factor-β1–Sma and Mad protein) signalling plays an important role in the development of renal tubulo-interstitial fibrosis. Secretion of CTGF (connective-tissue growth factor; CCN2) by PTECs (proximal-tubule epithelial cells) and EMT (epithelial–mesenchymal transdifferentiation) of PTECs to myofibroblasts in response to TGFβ are critical Smad-dependent events in the development of tubulo-interstitial fibrosis. In the present study we have investigated the distinct contributions of Smad2 and Smad3 to expression of CTGF, E-cadherin, α-SMA (α-smooth-muscle actin) and MMP-2 (matrix-metalloproteinase-2) in response to TGFβ1 treatment in an in vitro culture model of HKC-8 (transformed human PTECs). RNA interference was used to achieve selective and specific knockdown of Smad2 and Smad3. Cellular E-cadherin, α-SMA as well as secreted CTGF and MMP-2 were assessed by Western immunoblotting. TGFβ1 treatment induced a fibrotic phenotype with increased expression of CTGF, MMP-2 and α-SMA, and decreased expression of E-cadherin. TGFβ1-induced increases in CTGF and decreases in E-cadherin expression were Smad3-dependent, whereas increases in MMP-2 expression were Smad2-dependent. Increases in α-SMA expression were dependent on both Smad2 and Smad3 and were abolished by combined knockdown of both Smad2 and Smad3. In conclusion, we have demonstrated distinct roles for Smad2 and Smad3 in TGFβ1-induced CTGF expression and markers of EMT in human PTECs. This can be of therapeutic value in designing targeted anti-fibrotic therapies for tubulo-interstitial fibrosis.
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January 2006
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Research Article|
December 23 2005
The differential role of Smad2 and Smad3 in the regulation of pro-fibrotic TGFβ1 responses in human proximal-tubule epithelial cells
Mysore K. Phanish;
Mysore K. Phanish
1
*South West Thames Institute For Renal Research, St. Helier Hospital, Carshalton, Surrey SM5 1AA, U.K.
1To whom correspondence should be addressed (email m.phanish@btinternet.com).
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Nadia A. Wahab;
Nadia A. Wahab
†Renal Section, Division of Medicine, Imperial College London, Hammersmith Hospital, Ducane Road, London W12 ONN, U.K.
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Paul Colville-Nash;
Paul Colville-Nash
*South West Thames Institute For Renal Research, St. Helier Hospital, Carshalton, Surrey SM5 1AA, U.K.
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Bruce M. Hendry;
Bruce M. Hendry
‡Department of Renal Medicine, King's College Hospital, King's College London, Bessemer Road, London SE5 9PJ, U.K.
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Mark E. C. Dockrell
Mark E. C. Dockrell
*South West Thames Institute For Renal Research, St. Helier Hospital, Carshalton, Surrey SM5 1AA, U.K.
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Biochem J (2006) 393 (2): 601–607.
Article history
Received:
July 11 2005
Revision Received:
September 13 2005
Accepted:
October 27 2005
Accepted Manuscript online:
October 27 2005
Citation
Mysore K. Phanish, Nadia A. Wahab, Paul Colville-Nash, Bruce M. Hendry, Mark E. C. Dockrell; The differential role of Smad2 and Smad3 in the regulation of pro-fibrotic TGFβ1 responses in human proximal-tubule epithelial cells. Biochem J 15 January 2006; 393 (2): 601–607. doi: https://doi.org/10.1042/BJ20051106
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