Influence of exogenous TGFβ₁ on the expression of Smad2 and Smad3 in rat bone marrow-derived mesenchymal stem cells

Abstract

The expression of Smad2 and Smad3 and the influence of exogenous transforming growth factorβ₁ (TGFβ₁) on them in rat bone marrow-derived mesenchymal stem cells (MSCs) cultured in vitro were investigated. The effects of different concentrations of TGFβ₁ on cell proliferation and ALP activity were detected by MTT and PNPP in, MSCs respectively. The expression of Smad2 and Smad3 and the influence of exogenous TGFβ₁ on them were also examined by immunocytochemistry and Western blot assays. The exogenous TGFβ₁ induced a dose-dependent decrease in cell proliferation and a dose-dependent increase, in ALP activity, which plateaued at 5 ng/ml. Smad2 and Smad3 proteins were detected only in the cytoplasm in the absence of TGFβ₁ and TGFβ₁ could stimulate the translocation of them from the cytoplasm to the nucleus. The total amount of Smad2 protein remained unchanged before and after TGFβ₁ treatment (P>0.05). The expression levels of Smad3 remained unchanged after 3 h and 6 h treatment (P>0.05), but decreased markedly after 24 h treatment (P<0.05). It was concluded that TGFβ₁ is a latent osteoinductive factor involved in osteoblastic differentiation. Both Samd2 and Samd3 mediate TGFβ₁ signaling as downstream mediators in MSCs. The biological output of TGFβ₁ triggering the osteoblastic differentiation could be entirely determined by Smad3 in MSCs.

Keywords

transforming growth factorβ₁ / mesenchymal stem cells / Smad2 / Smad3

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