ABSTRACT
BACKGROUND:The mycobacterium tuberculosis heat shock protein 10 exerts effects on the osteoclasts by in vitro mouse cranium experiment, OBJECTIVE:To investigate the effect and mechanism of recombinant mycobacterium tuberculosis heat shock protein 10 (CPN10) on the differentiation of osteoclasts in the in vitro culture system that induces osteoclast differentiation. METHODHuman macrophage colony-stimulating factor-dependent adhesive blood mononuclear cells were divided into four groupreceptor activator for nuclear factor-κB ligand (RANKL)+CPN10 (1 mg/L), RANKL, CPN10 (1 mg/L), and negative control (complete culture medium). Monocytes were resuspended in a-MEM medium containing macrophage colony-stimulating factor, and were cultured in each group for 7, 14, 21 days. The morphology, quantity and bone resorption area of osteoclasts were examined by tartrate-resistant acid phosphatase (TRAP) staining. The expressions of NFATc1 and c-Fos gene and protein were also detected. RESULTS AND CONCLUSION:In negative control group, no TRAP-positive multinucleated osteoclasts generated, while in the other groups, TRAP-positive multinucleated osteoclasts differentiated and formed the lacunae in the smal bone grinding. The number of osteoclasts formation and resorption in CPN10 group were significantly lower than that in RANKL+CPN10 group. The expression of NFATc1 and c-Fos in the negative control group C was significantly lower than that of RANKL+CPN10 group and CPN10 group. However, CPN10 expressed NFATc1 and c-Fos protein, which was significantly lower than RANKL+CPN10 group. CPN10 is involved in the formation of osteoclasts, and the mechanism is related with the upregulation of NFATc1, c-Fos expression.
ABSTRACT
BACKGROUND:Mycobacterium tuberculosis heat shock protein 10 (r-Mt cpn10) is one of the main factors that cause bone tuberculosis dissolution and absorption as wel as inhibits the proliferation of osteoblasts. Receptor activator of nuclear factor kappa B ligand and osteoprotegerin are the important factors influencing bone metabolism. OBJECTIVE:To observe the effect of r-Mt cpn10 on human osteoblast proliferation, alkaline phosphatase secretion, expression of receptor activator of nuclear factor-kappa B ligand mRNA and osteoprotegerin mRNA. METHODS:Human bone marrow stromal cel s were induced to differentiate into osteoblasts, and osteoblasts at passage 3 were cultured with various concentrations of r-Mt cpn10 (0.1, 1, 10 mg/L). Osteoblasts cultured without r-Mt CPN10 were assigned as controls. RESULTS AND CONCLUSION:MTT assay results showed that, compared with control group, r-Mt cpn10 at different concentrations inhibited osteoblast proliferation and alkaline phosphatase secretion (P<0.05). RT-PCR analysis showed that, r-Mt cpn10 at different concentrations increased receptor activator of nuclear factor-kappa B ligand mRNA expression (P<0.01), and inhibited osteoprotegerin mRNA expression in a concentration-dependent manner (P<0.01). 10 mg/L r-Mt cpn10 exhibited the strongest effect (P<0.01). The r-Mt cpn10 can inhibit osteoblast proliferation and alkaline phosphatase activity, and it may influence bone metabolism by regulating the expression of receptor activator of nuclear factor-kappa B ligand mRNA and osteoprotegerin mRNA.