Effect of long-term cultivation on morphological and biological characteristics of human periodontal ligament stem cells.

OBJECTIVES: Recently, it was demonstrated that human periodontal ligament stem cells have great potential for tissue engineering and regenerative medicine not limited to oro-maxillofacial region. They are easily accessible and they may be expanded under in vitro conditions. In this study we assessed the effect of long-term cultivation on the selected biological and morphological properties of human periodontal ligament stem cells.

METHODS: Periodontal tissues were obtained from normal impacted third molars of healthy donors (n=5; aged 18-27 years), after obtaining informed consent. The explant technique was used to initialize cell culture and further expansion in vitro was carried out in complete culture medium (D-MEM + 10% foetal bovine serum + gentamicin) with passaging in 80% of confluence using trypsine up to 25th passage. Cells were continually analyzed for morphology changes by inverted and transmission electron microscope. The analysis of selected biological characteristics (expression of surface antigens and selected genes involved in cell regulation and apoptosis, cell cycle analysis and cell senescence) were performed, as well.

RESULTS AND CONCLUSIONS: Obtained results showed that long-term cultivation lead in to considerable changes in morphology and affect the proliferation and cell cycle of human periodontal ligament stem cells. On the other hand, it did not affect their immunophenotype as well as function of cell cycle, apoptosis regulators and telomerase activity also in high passages. However, further studies considering stem cells bio-safety have to be carried out prior their clinical application.