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Estrogen attenuates TGF-β1 induced elastogenesis in rat urethral smooth muscle cells by inhibiting Smad response
Guiting Lin1, Amjad Alwaal1, Fionna Sun1
1Knuppe Molecular Urology Laboratory, Department of Urology, School of Medicine, University of California-San Francisco, San Francisco, California.
Purpose:
We investigated the effect and mechanism of estrogen on elastogenesis in urethral smooth muscle cells in vitro.
Materials And Methods:
Urethral smooth muscle cells were isolated from normal adult female rats. For elastogenesis assay cells were treated with TGF-β1, the potent TGF-β1 receptor inhibitor SB431542 and estrogen for 2 weeks. Real-time polymerase chain reaction was performed to assay gene expression during this process. Activity of the TGF-β1 responsive elements CAGA(12)-Luc and GCCG(12)-Luc were also assayed. Estrogen receptor and Smad2/3 interaction was evaluated by immunoprecipitation and Western blot.
Results:
TGF-β1 induced elastogenesis in rat urethral smooth muscle cells. This effect was partially blocked by estrogen and completely abrogated by SB431542. SB431542 completely inhibited activation of the Smad2/3 response element CAGA(12)-Luc and estrogen significantly inhibited activation. The Smad1/4 response element GCCG(12)-Luc was not affected by SB431542 treatment but estrogen partially inhibited the activation of GCCG(12)-Luc induced by TGF-β1. Estrogen receptor bound to Smad 2 and 3 in vitro.
Conclusions:
Estrogen attenuated TGF-β1 induced elastogenesis via binding of its activated receptor to Smad2/3 to inhibit the TGF-β1 response element in rat urethral smooth muscle cells.
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