Influence of rs2228570 on Transcriptional Activation by the Vitamin D Receptor in Human Gingival Fibroblasts and Periodontal Ligament Cells

Background: rs2228570 is the only known single nucleotide polymorphism of the vitamin D receptor (VDR) that alters the protein structure. VDRs can be distinguished using the restriction endonuclease FokI and accordingly divided into three genotypes: FF, Ff, and ff. Influence of rs2228570 on transcriptional activation by VDRs in human gingival fibroblasts (hGFs) and periodontal ligament cells (hPDLCs) is investigated in this study. Methods: From 15 donors, hGFs and hPDLCs were cultured, genomic DNA was extracted, and genotypes were determined using the polymerase chain reaction (PCR)-restriction fragment length polymorphism method. Cells were stimulated with calcitriol with or without VDR antagonist ZK159222 or osteogenic induction. Alkaline phosphatase, osteocalcin, and VDR messenger RNA (mRNA) expression were detected using real-time PCR. Alkaline phosphatase and osteocalcin protein expression were detected by enzyme activity assays with p-nitrophenyl phosphate substrate and enzyme-linked immunosorbent assay, respectively. Results: Among the 15 donor cell cultures, the number of FF, ff, and Ff genotypes were 5, 3, and 7, respectively. There were no significant differences in expression of alkaline phosphatase or osteocalcin among the three genotypes in hGFs. However, after stimulation with calcitriol, alkaline phosphatase and osteocalcin mRNA levels in FF-hPDLCs were significantly higher than in other hPDLCs genotypes, as was osteocalcin protein expression. Furthermore, when ZK159222 was included, this difference disappeared, and when osteogenic induction was performed, alkaline phosphatase and osteocalcin mRNA and protein levels were higher in FF-hPDLCs than in the other hPDLCs genotypes. Conclusion: The FF-VDR genotype is associated with the most remarkable upregulation of alkaline phosphatase and osteocalcin in hPDLCs.