Calcitonin promotes collagen synthesis and osteogenesis in human periodontal ligament stem cells / 第二军医大学学报

ABSTRACT

Objective: To investigate the effect of calcitonin (CT) on promoting collagen synthesis and osteogenesis of human periodontal ligament stem cells (hPDLSCs). Methods: Fifty adult participants were divided into chronic periodontitis (CP) group (n=25) and control group (n=25). In the CP group, the anterior maxilla with probing depth ≥5 mm and the sites with imaging evidence of bone loss were selected. The gingival crevicular fluid (GCF) samples were collected from 6 maxillary sites in each patient. In the control group, multiple sites without inflammation (10 to 12 per subject) were sampled to ensure that a sufficient amount of GCF was collected. The expression of CT, transforming growth factor β1 (TGF-β1) and bone morphogenetic protein (BMP) 2/4/7 in GCF was detected by enzyme linked immunosorbent assay (ELISA), and the correlation between CT expression and clinical parameters such as periodontal pocket probing depth (PD), clinical attachment loss (CAL) and gingival index (GI), and the above-mentioned indicators was investigated with Spearman correlation analysis. hPDLSCs were infected with the adenoviruses carrying CT gene (Ad.CT) and the expression of mRNA and protein of TGF-β1, BMP2/4/7, alkaline phosphatase (ALP), osteocalcin (OCN) and collagen type I/III (Col I/III) were detected by quantitative real-time PCR and Western blotting. Results: The expression level of CT in GCF of the CP group was significantly higher than that of the control group ([32.62±1.46] ng/mL vs [17.70 ± 0.76] ng/mL, P<0.01). The expression of CT was positively correlated with clinical parameters such as PD, CAL and GI (P<0.01, P<0.05). The expression levels of BMP2/4/7 and TGF-β1 in GCF of the CP group were significantly higher than those of the control group (BMP2 [138.67 ± 4.04] ng/mL vs [103.96 ± 2.78] ng/mL, BMP4 [155.53 ± 3.55] ng/mL vs [133.15 ± 2.92] ng/mL; BMP7 [106.59 ± 2.85] ng/mL vs [90.22±1.56] ng/mL; TGF-β1 [105.92 ± 3.40] ng/mL vs [89.85 ± 2.42] ng/mL; all P<0.01). The expression of BMP2/4/7 and TGF-β1 was negatively correlated with CT expression (P<0.01, P<0.05). The overexpression of CT significantly increased the expression of TGF-β1, Col I/III and osteoblast markers BMP2/4, ALP and OCN in GCF (all P<0.01). Compared with the cells co-infected with Ad.CT and Ad.Null, the cells co-infected with Ad.CT and small interfering RNA specifically blocking TGF-β1 (Ad.TGF-β1 siRNA) had significantly lower collagen expression (Col I 0.16 ± 0.02 vs 0.22 ± 0.03; Col III 0.11±0.01 vs 0.15 ± 0.02; both P<0.01). Compared with Ad.CT infected cells, the protein expression levels of ALP and OCN were significantly decreased in Ad.CT and noggin co-treated cells (ALP 0.19 ± 0.02 vs 0.25 ± 0.03; OCN 0.13 ± 0.01 vs 0.19 ± 0.02; both P<0.01). Conclusion: CT can promote collagen synthesis and osteogenesis in hPDLSCs through TGF-β1 and BMP signaling transduction pathways.