The roles of calcium-sensing receptor and calcium channel in osteogenic differentiation of undifferentiated periodontal ligament cells

Abstract

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 CaCl2 significantly induced proliferation, bone-related gene expression, and calcification in cell line 1-17. During culture with 5 mM CaCl2, 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 CaCl2 exposure alone. The L-VDCC inhibitor, nifedipine, significantly suppressed osteogenic differentiation of cell line 1-17 treated with 5 mM CaCl2 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.