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.

Involvement of the calcium-sensing receptor in mineral trioxide aggregate-induced osteogenic gene expression in murine MC3T3-E1 cells

“…Also, NPS2143 decreased the protein level of CaSR in the presence of MTA. It has been demonstrated previously that exogenous calcium stimuli increased the expression of CaSR in mammary gland epithelial cells 30) and periodontal ligament cells 17) . These findings were consistent with our results.…”

“…Comparable effects have been observed in bone marrow-derived progenitor cells 21) , preadipocytes 22) , and periodontal ligament cells 17) . These findings suggest the presence of a mechanism involving CaSR expression on the cell membrane responding to local changes in calcium ion concentrations.…”

“…Many in vitro and in vivo studies have demonstrated that this receptor is a key mediator of the direct actions of calcium ions on the parathyroid, and it regulates homeostatic responses that restore calcium ions in the blood to normal level 16) . Other cell types, including bone and periodontal ligament cells, have been shown to express CaSR 17,18) . Especially in osteoblasts, local changes in the concentrations of extracellular calcium ions can be sensed by CaSR, resulting in regulation of their proliferation, differentiation, and mineralization 19) .…”

Calcium-Sensing Receptor-Mediated Osteogenic and Early-Stage Neurogenic Differentiation in Umbilical Cord Matrix Mesenchymal Stem Cells from a Large Animal Model

“…In the same study, the authors reported evidences demonstrating that the signaling pathway between extracellular Ca 2+ and bone morphogenetic protein 2 (BMP-2), a protein essential to maintain bone homeostasis and having a prominent role in fracture healing, involves type L voltage-dependent Ca 2+ channels (L-VDCC) rather/more than CaSR. A subsequent study strengthen the role of CaSR in supporting osteogenic differentiation, as it demonstrated that extracellular calcium promotes osteogenic differentiation in human periodontal ligament stem/progenitor cells and that CaSR and L-VDCC appear to act reciprocally in mediating this process [63]. In ovine amniotic fluid MSCs, increasing [Ca 2+ ] o was reported to increase ALP activity [49].…”

Clinical and Molecular Perspectives of Reparative Dentin Formation

“…However, the presence of CaSR in bone-forming cells is controversial 108 , and osteoblasts derived from CaSR-null mice remained to possess osteogenic differentiation potential 109,110 . Further, another study demonstrated that inhibition of CaSR further induced, rather than suppressed, Ca 2+ -mediated osteogenic differentiation 111 , suggesting that CaSR is dispensable in osteogenic differentiation and mineralization.…”