Abstract
Aim：Transforming growth factor β1 (TGF-β1) plays an important role in cell proliferation, matrix formation, and osteogenesis / odontogenesis. The purpose of this study is to investigate the effects of TGF-β1 on human apical papilla cells. We hypothesize that TGF-β1 can stimulate the two signaling pathways, MEK/ERK and Smad 2/3 to mediate alkaline phosphatase (ALP) expression, collagen matrix deposition and calcium deposition in human apical papilla cells. Materials and Methods：Primary-cultured human apical papilla cells were treated with TGF-β1. In some experiments, apical papilla cells were pretreated with SB431542 ( an ALK5 / Smad 2/3 inhibitor) or U0126 ( a MEK/ERK inhibitor) 30 minutes before adding TGF-β1. Cell viability and cell growth were examined by MTT assay or direct counting of viable cell numbers. Collagen content was determined by Sircol Collagen assay. Cell differentiation and mineralization were evaluated by alkaline phosphatase（ALP）staining, ALP activity quantitative assay, Alizarin Red staining, and calcium quantitative analysis. Changes in mRNA expression were determined by reverse-transcription Polymerase Chain Reaction（RT-PCR）. Results：In human apical papilla cells, TGF-β1 increased cell numbers and cell viability. Cells under the treatment of TGF-β1 (>0.1 ng/ml) would induce collagen formation. Pretreatment of U0126（a MEK/ERK inhibitior）was not effective to reverse the effects of TGF-β1 on cell growth and matrix formation; but SB431542（an ALK5/ Smad 2/3 inhibitor）could prevent those effects. In the differentiation, TGF-β1 down-regulates ALP activity in the lower concentration （0.5-1 ng/ml) and up-regulates it in the higher concentration（5-10 ng/ml）. SB431542 can reverse the effect of TGF-β1 at lower or higer concentration; but U0126 only can reverse the effect of TGF-β1 at lower concentration. Besides, TGF-β1 inhibits the mineralization on apical papilla cells. Conclusion：Signal transduction of TGF-β1 in human apical papilla cells is complex. Different cell sources, different concentrations of growth factor, presence of absence of serum or culture conditions can affect the effects of TGF-β1. Besides, the well-known Smad 2/3 pathway, non Smad pathway（ex. ERK）may also take part in this complex system. These results highlight our future use of growth factors in pulpal repair and dentinogenesis.
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