Abstract
Periodontal diseases comprise mixed bacterial infections mainly caused by Gram‑negative anaerobic bacteria. Lipopolysaccharides (LPS) are important virulence factors and periodontal pathogens, which change local cytokine levels and promote osteoblast apoptosis, thereby leading to an imbalance in bone remodeling mechanisms and accelerating bone loss. Calcitonin gene‑related peptide (CGRP) is a vasoactive neuropeptide that is released from sensory nerves and has a positive effect on osteoblast proliferation and differentiation. In addition, this small molecule peptide is an important immune regulator in the inflammatory response. The aim of the present study was to assess the in vitro effects of CGRP on Porphyromonas gingivalis (Pg)LPS‑induced osteoblast apoptosis. Osteoblast cultures were stimulated either with various concentrations of PgLPS (0, 25, 50, 100, 500 and 1,000 ng/ml) for 48 h or with 500 ng/ml PgLPS for various lengths of time (0, 6, 12, 24, 48 and 72 h). The PgLPS‑stimulated cells were pretreated with different concentrations of CGRP (0, 1, 10, 100 and 1,000 nM) and cell viability and apoptotic rates were measured by Cell Counting kit‑8 assays and flow cytometry, respectively. CGRP, cleaved (c)‑Caspase‑8 and c‑Caspase‑3 protein expression levels were analyzed by western blotting. Changes in cytokine expression levels, which included tumor necrosis factor (TNF)‑α, interleukin (IL)‑1β, IL‑6, monocyte chemotactic protein (MCP)‑1 and MCP‑2, were measured by ELISA. PgLPS was demonstrated to inhibit osteoblast viability and promote apoptosis in a time‑ and concentration‑dependent manner. CGRP expression was revealed to reduce PgLPS‑induced cytostatic activity and apoptosis in osteoblasts. CGRP also suppressed the PgLPS‑induced release of TNF‑α and inhibited the activation of c‑Caspase‑3 and c‑Caspase‑8, thus preventing apoptosis in osteoblasts. CGRP may be an important neuropeptide in bone remodeling and may reduce osteoblast apoptosis in inflammatory conditions. These results may provide a solid foundation for CGRP to serve as a new target for the treatment of periodontitis.