Effects of aging on ossification of periodontal ligament cells

Objectives: Epidemiological studies have indicated positive correlations between senescence and disease severity of periodontitis. However, the molecular mechanism(s) of an age-dependent periodontal tissue destruction are not fully elucidated. Periodontal ligament (PDL) cells play critical roles in the wound healing and regeneration process of periodontal tissues. PDL cells are characterized by unique stemness-like property, such as self-renewal and differentiation into osteoblastic or cementblastic cells. Balancing the proper abilities of cell growth and multipotency of PDL cells and mesenchymal stem cells is crucial for periodontal tissue homeostasis. Therefore, accumulated cellular damage in the process of aging could affect the disease onset and healing ability of periodontal tissues. In this study, we investigated at molecular level whether cellular aging can affect the hard tissue-forming ability of PDL cells.

Methods: We used MPDL22 (murine periodontal ligament cells, passage #6~10)　which were derived from BALB/c mice. MPDL22-S (Senescence prototype, passage #70~) was  utilized for the analysis of aged PDL cells. MPDL22 were cultured in the calcification-inducing medium including L-ascorbic acid and b-glycelophosphate in the presence or absence of BMP-2, TGF-b w/wo SB431542. Calcified nodule formation and mRNA expression of calcification-related genes were examined by Alizarin staining and real-time PCR, respectively. Collagen type I and CTGF productions from PDL cells in culture supernatants were quantified by Western blotting.

Results: Both of MPDL22 and MPDL22-S showed similar cell shape and growth rate.MPDL22-S showed retarded ossification paralleled with the mature collagen type I and CTGF production in long term calcification culture. SB431542 treatment restored the low responsiveness of MPDL22-S to BMP-2-induced ossification.

Conclusions: These results suggest that regulation of TGF-b signaling may ameliorate low ECM protein production and cytodifferentiation of aged PDL cells.