Enhancement of periodontal tissue regeneration by transplantation of osteoprotegerin-engineered periodontal ligament stem cells.

INTRODUCTION: The objective of the present study was to evaluate the capacity of a tissue-engineered complex of human osteoprotegerin (hOPG)-transfected periodontal ligament stem cells (PDLSCs) seeding on beta-tricalcium phosphate (ß-TCP) to regenerate alveolar bone defects in New Zealand rabbits. METHODS: PDLSCs were isolated from rabbit periodontal ligament tissues and expanded in vitro to enrich PDLSC numbers, and their proliferative activities and differentiation capability were evaluated under specific induction conditions. Lentiviral vector containing hOPG and enhanced green fluorescent protein (EGFP) was constructed by using Gateway technology and transfected into rabbit PDLSCs. The expression of hOPG was determined with quantitative real-time reverse transcription-polymerase chain reaction and Western blot. The PDLSCs with or without engineered hOPG were seeded on ß-TCP scaffolds prior to transplantation. Morphological characterization of cells and materials was done by scanning electron microscope. Twenty rabbits with alveolar bone defects were randomly allocated into four groups and transplanted with ß-TCP, PDLSCs/ß-TCP, and hOPG-transfected PDLSCs/ß-TCP or were left untreated as a control. Animals were sacrificed 12 weeks after operation for histological observation and histomorphometric analysis. RESULTS: PDLSCs expressed STRO-1 and vementin and favored osteogenesis and adipogenesis in conditioned media. Expressions of hOPG were significantly upregulated after transfection of the lentiviral vector into PDLSCs. PDLSCs attached and spread well on ß-TCP, and there was no significant difference in growth of PDLSCs on ß-TCP between the hOPG transfection group and the non-transfection group. The histological observation and histomorphometric analysis showed that the hOPG-transfected PDLSCs/ß-TCP complex exhibited an earlier mineralization and more bone formation inside the scaffold than control, ß-TCP, and PDLSCs/ß-TCP complexes. Implantation of hOPG-transfected PDLSCs contributed to new bone formation as determined by EGFP gene expression under circularly polarized light microscopy. CONCLUSIONS: The present study demonstrated the feasibility of ß-TCP scaffolds for primary PDLSC culture and expression of hOPG gene in vitro and in vivo, and hOPG-transfected PDLSCs could serve as a potential cell source for periodontal bone regeneration, which may shed light on the potential of systemic hOPG gene therapy in combination with PDLSC tissue engineering as a good candidate in periodontal tissue engineering for alveolar bone regeneration.

[Protective effect of resveratrol on apoptosis of human periodontal ligament cells in vitro].

OBJECTIVE: To investigate the effects of resveratrol (RES) on apoptosis of human periodontal ligament cells (HPLC). METHODS: HPLC were subjected to oxidative injury induced by H2O2 for 24 h after pretreatment with different concentration of RES. HPLC were then divided into the control, model, vector, RES 1, 10, 30, 50 micromol/L treatment group. The viability of the HPLC was determined by methyl thiazolyl tetrazolium (MTT) method. Lactate dehydrogenase (LDH) rate and malondialdehyde (MDA) in the culture medium, superoxide dismutase (SOD) in the HPLC homogenate were evaluated by spectrophotometry. The apoptotic HPLC was detected by flow cytometry (FCM) and calculated by relative apoptosis rate. Bax and Bcl-2 protein levels were detected by Western blotting. RESULTS: RES increased the cell survival rate after H2O2 injury. The survival rate of RES 30 micromol/L group was (86.1 +/- 4.1)% and the model group was (54.6 +/- 4.0)%, which was significantly different between the two groups (P < 0.01). The LDH leakage rate and MDA content of the RES 30 micromol/L group were (32.6 +/- 2.0)% and (1.70 +/- 0.21) micromol/L, which were significantly different with that in the model group (P < 0.01). At the same time RES could remarkably restore the vitality of SOD in the HPLC. RES increased Bcl-2 and reduced the expression of Bax protein. The apoptosis rate of the RES 30 micromol/L group and model group was (14.84 +/- 1.36)% and (64.37 +/- 2.34)%, respectively (P < 0.01). The protective effect of RES on the cell apoptosis was in a dose-dependent manner, reaching peak at a concentration of 30 micromol/L (P < 0.01). CONCLUSIONS: RES reduced oxidative stress and apoptosis in an experimental HPLC injury model induced by H2O2. RES plays a key role in the HPLC protection against oxidative injury.