Potential function of TGF-β isoforms in maturation-stage ameloblasts

Abstract Objectives To investigate potential functions of transforming growth factor-beta (TGF-β) isoforms in maturation-stage ameloblasts during amelogenesis. Methods In vivo activation of TGF-β was characterized by using matrix metalloproteinase 20 null ( Mmp20 -/- ) and wild-type ( Mmp20 +/+ ) mice. Using mHAT9d cells cultured in the presence of each TGF-β isoform, (1) cell proliferation was determined by MTS assay, (2) immunostaining with anti-cleaved caspase-3 monoclonal antibody was performed and apoptotic indices were measured, (3) gene expression was analyzed by RT-qPCR, and (4) the uptake of amelogenin into mHAT9d cells was directly observed using a fluorescence microscope. Results TGF-β1 and TGF-β3 were present in the enamel matrix of developing teeth which were activated by MMP20 in vivo . A genetic study revealed that the three TGF-β isoforms upregulate kallikrein 4 (KLK4) mRNA levels but downregulate carbonic anhydrase II. Moreover, TGF-β1 and TGF-β2 significantly upregulated the mRNA level of amelotin, whereas TGF-β3 dramatically downregulated the mRNA levels of odontogenic ameloblast-associated protein (ODAM), family with sequence similarity 83 member H (FAM83H), and alkaline phosphatase (ALP). Immunostaining analysis showed that the apoptosis of mHAT9d cells is induced by three TGF-β isoforms, with TGF-β3 being most effective. Both TGF-β1 and TGF-β3 induced endocytosis of amelogenin. Conclusions We propose that TGF-β is regulated in an isoform-specific manner to perform multiple biological functions such as gene expression related to the structure of basal lamina/ameloblasts, mineral ion transport, apoptosis, and endocytosis in maturation-stage ameloblasts.