[Hypoxia regulates osteogenic differentiation of human periodontal ligament stem cells]

OBJECTIVE: To investigate the effect of hypoxia on isolated human periodontal ligament stem cells (hPDLSC) in vitro.

METHODS: hPDLSC were exposed to normoxia (20% O(2)) and hypoxia (1.5% - 2% O(2)). Total cellular RNA and protein were collected on day 1, 2, 4 and 6 after culture. Western analysis and semi-quantitative RT-PCR were used to analyze osteogenic differentiation of hPDLSC, including alkaline phosphatase (ALP), osteocalcin (OCN), secreted protein acidic and rich in cysteine (SPARC) and bone morphogenetic protein-2 (BMP-2).

RESULTS: In the first two days hypoxia slightly increased the growth of hPDLSC [A value was 0.697(hypoxia) vs 0.617 (nomoxia)] and after the third day hypoxia dramatically decreased the growth of the hPDLSC [A value was 0.870(hypoxia) vs 1.242 (nomoxia)]. Up to 90% reduction of ALP activity was observed in hPDLSC after 5 days of hypoxia [A value was 0.004(hypoxia) vs 0.049(nomoxia)]. Hypoxia decreased SPARC expression at protein level and down-regulated ALP, OCN and BMP-2 expression at mRNA level in comparison with nomoxia.

CONCLUSIONS: Hypoxia inhibited proliferation of hPDLSC and down-regulated BMP-2 mRNA expression, then down-regulated their target genes such as ALP, OCN, and SPARC, thus inhibiting critical steps in osteogenic differentiation of hPDLSC.