Differentiation of human embryonic stem cells on periodontal ligament fibroblasts in vitro

Human embryonic stem cells (hESCs) are pluripotent cells with unlimited proliferation potential and differentiation capacity to all types of somatic cells. Periodontal tissue engineering based on in vitro expanded cells holds the promise to overcome the limitations associated with contemporary regenerative techniques. The aim of this study was to investigate the differentiation patterns of hESCs under the influence of periodontal ligament cells in vitro. hESCs (HUES-9) were expanded and characterized for their pluripotency. Then they were transfected with green fluorescent protein-carrying plasmid, and cocultured with human periodontal ligament fibroblastic cells for 21 days. Two experimental groups were established with different medium constituents. Specimens were fixed at days 7, 14, and 21 and were analyzed morphologically under inverted light microscope, and by immunohistochemistry using antibodies against collagen types I and III, fibronectin, fibroblast surface protein, vimentin, and pancytokeratin. Our results demonstrate different patterns of cell differentiation between groups, with about one-fifth of cells in colonies acquiring characteristics similar to periodontal ligament fibroblastic progenitors while others proceed toward distinctive lineages. This indicates the feasibility to direct the differentiation of hESCs toward the periodontal ligament fibroblastic progenitors to some extent. These findings support the notion that hESCs may become a cell source with unlimited supply for periodontal tissue engineering applications.