Establishment of a Primary Culture System of Human Periodontal Ligament Cells that Differentiate into Cementum Protein 1-expressing Cementoblast-like Cells.

Abstract

A better understanding of cementogenesis and cementoblast differentiation would be useful for periodontal therapy. The aim of this study was to establish a cell culture system that reflects cementum formation in periodontal tissue and determine whether or not isolated and cultured primary human periodontal ligament (PDL) cells could be used for the study of the differentiation of cementoblast. PDL cells were isolated from the outgrowths of tissue fragments of human PDL. PDL cells were incubated for up to 21 days in differentiation medium containing β-glycerophosphate and ascorbic acid. The changes in the cells were detected by alkaline phosphatase (ALP) and von Kossa staining. Real-time polymerase chain reaction was also performed for cementum protein 1 (CEMP1), which is a specific marker of cementoblasts and their progenitors. On day 5, a small number of PDL cells, which were fibrous, were positive for ALP. On day 7, almost all cells were positive for ALP. On day 14, mineralization nodules appeared, as seen by positive von Kossa staining; the nodules increased in number and size by day 21. The expression of CEMP1 was detected on day 5, and its expression level increased gradually by day 7, reached a peak on day 14, and decreased by day 21. Human PDL cells were used to establish a culture system that reflects cementum formation. Our results suggested that this culture method is convenient and useful for the study of cementogenesis and cementoblast differentiation.