Hydrogen sulfide suppresses mineralized nodule formation by osteoblastic ROS17/2.8 cells

Hydrogen sulfide (H₂S), which is the main component of the volatile sulfur compounds (VSCs) produced by putrefactive bacteria, plays a role in not only oral malodor, but also the initiation and progress of periodontitis. The loss of alveolar bone associated with periodontitis appears to be related to local factors that change the balance between bone formation and resorption. Previous studies have indicated that VSCs, including H₂S, induce bone resorption by stimulating the differentiation and activation of osteoclasts; however, there is little information about the effect of VSCs on bone formation by osteoblasts. Therefore, we examined the effect of H₂S on cell proliferation, alkaline phosphatase (ALPase) activity, non-collagenous extracellular matrix protein (ECMP) expression, and mineralized nodule formation using ROS17/2.8 cells as osteoblasts. Cells were cultured with 0 (control), 10^-4, 10^-3, or 10^-2 M sodium hydrogen sulfide (NaHS; H₂S donor). Mineralized nodule formation was detected by alizarin red staining. The expression of non-collagenous ECMP, including bone sialoprotein (BSP) and osteopontin (OPN), was examined at the mRNA and protein levels using real-time PCR and Western blotting, respectively. Cell proliferation was suppressed by the addition of 10^-2 M NaHS, but was unaffected by 10-3 and 10^-4 M NaHS. ALPase activity and the expression of BSP and OPN at the mRNA and protein levels were decreased when cells were cultured with 10^-4 and/or 10^-3 M NaHS. In addition, mineralized nodule formation was strongly inhibited by 10^-4 and 10^-3 M NaHS. These results suggest that H₂S suppresses mineralized nodule formation by decreasing ALPase activity and the production of BSP and OPN by osteoblasts.