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Elevated extracellular calcium has been shown to promote the differentiation of osteoblasts. However, the way that calcium affects the osteogenic differentiation of human periodontal ligament stem/progenitor cells (PDLSCs) remains unclear. Our aim has been to investigate the proliferation and osteogenic differentiation of a calcium-exposed human PDLSC line (cell line 1-17) that we have recently established and to elucidate the roles of the calcium-sensing receptor (CaSR) and L-type voltage-dependent calcium channel ( L-VDCC) in this process. Proliferation activity was investigated by WST-1 assay, and gene and protein expression was examined by quantitative reverse transcriptase plus the polymerase chain reaction and immunostaining, respectively. Calcification assay was performed by von Kossa and Alizarin red staining. Treatment with 5 mM CaCl significantly induced proliferation, bone-related gene expression, and calcification in cell line 1-17. During culture with 5 mM CaCl, this cell line up-regulated the gene expression of CaSR, which was reduced after 7 days. Simultaneous treatment with NPS2143, a CaSR inhibitor, and calcium significantly further increased bone-related gene expression and calcification as compared with CaCl exposure alone. The L-VDCC inhibitor, nifedipine, significantly suppressed osteogenic differentiation of cell line 1-17 treated with 5 mM CaCl and promoted the expression of CaSR, as compared with calcium treatment alone. Thus, elevated extracellular calcium promotes the proliferation and osteogenic differentiation of a PDLSC line. Antagonizing CaSR further enhances the effect of calcium on osteogenic differentiation, with CaSR expression being regulated by L-VDCC under extracellular calcium. Extracellular calcium might therefore modulate the osteogenic differentiation of PDLSCs through reciprocal adjustments of CaSR and L-VDCC. [ABSTRACT FROM AUTHOR]

Repair of damaged periodontal ligament (PDL) tissue is an essential challenge in tooth preservation. Various researchers have attempted to develop efficient therapies for healing and regenerating PDL tissue based on tissue engineering methods focused on targeting signaling molecules in PDL stem cells and other mesenchymal stem cells. In this context, we investigated the expression of epidermal growth factor (EGF) in normal and surgically wounded PDL tissues and its effect on chemotaxis and expression of osteoinductive and angiogenic factors in human PDL cells (HPDLCs). EGF as well as EGF receptor (EGFR) expression was observed in HPDLCs and entire PDL tissue. In a PDL tissue-injured model of rat, EGF and IL-1β were found to be upregulated in a perilesional pattern. Interleukin-1β induced EGF expression in HPDLCs but not EGFR. It also increased transforming growth factor-α ( TGF-α) and heparin-binding EGF-like growth factor ( HB-EGF) expression. Transwell assays demonstrated the chemotactic activity of EGF on HPDLCs. In addition, EGF treatment significantly induced secretion of bone morphogenetic protein 2 and vascular endothelial growth factor, and gene expression of interleukin-8 ( IL-8), and early growth response-1 and -2 ( EGR-1/ 2). Human umbilical vein endothelial cells developed well-formed tube networks when cultured with the supernatant of EGF-treated HPDLCs. These results indicated that EGF upregulated under inflammatory conditions plays roles in the repair of wounded PDL tissue, suggesting its function as a prospective agent to allow the healing and regeneration of this tissue. [ABSTRACT FROM AUTHOR]

Basic fibroblast growth factor (bFGF) is a cytokine that promotes the regeneration of the periodontium, the specialized tissues supporting the teeth. bFGF, does not, however, induce the synthesis of smooth muscle actin alpha 2 (ACTA2), type I collagen (COL1), or COL3, which are principal molecules in periodontal ligament (PDL) tissue, a component of the periodontium. We have suggested the feasibility of using transforming growth factor-β1 (TGFβ1) to induce fibroblastic differentiation of PDL stem/progenitor cells (PDLSCs). Here, we investigated the effect of the subsequent application of TGFβ1 after bFGF (bFGF/TGFβ1) on the differentiation of PDLSCs into fibroblastic cells. We first confirmed the expression of bFGF and TGFβ1 in rat PDL tissue and primary human PDL cells. Receptors for both bFGF and TGFβ1 were expressed in the human PDLSC lines 1-11 and 1-17. Exposure to bFGF for 2 days promoted vascular endothelial growth factor gene and protein expression in both cell lines and down-regulated the expression of ACTA2, COL1, and COL3 mRNA in both cell lines and the gene fibrillin 1 ( FBN1) in cell line 1-11 alone. Furthermore, bFGF stimulated cell proliferation of these cell lines and significantly increased the number of cells in phase G2/M in the cell lines. Exposure to TGFβ1 for 2 days induced gene expression of ACTA2 and COL1 in both cell lines and FBN1 in cell line 1-11 alone. BFGF/TGFβ1 treatment significantly up-regulated ACTA2, COL1, and FBN1 expression as compared with the group treated with bFGF alone or the untreated control. This method might thus be useful for accelerating the generation and regeneration of functional periodontium. [ABSTRACT FROM AUTHOR]