降钙素对小鼠成骨细胞增殖及OPG/RANKL表达的影响

目的探讨降钙素对小鼠成骨细胞增殖及骨保护素（OPG）／细胞核因子KB受体活化因子配体（RANKL）表达的影响。方法取出生24h内的小鼠30只,无菌的条件下取出颅骨,应用酶消化法进行成骨细胞的培养,在培养成骨细胞的培养液中加入不同浓度的降钙素,首先应用MTT法检测这种药物对成骨细胞增殖的影响,然后应用RT-PCR法检测其对OPG／RANKL mRNA的表达及ELISA法检测OPG蛋白分泌的影响。结果降钙素能促进成骨细胞的增殖。降钙素能增强成骨细胞中OPG mRNA的表达同时抑制成骨细胞中RANKL mRNA的表达,并能促进OPG蛋白的分泌。结论降钙素能促进成骨细胞的增殖及成骨细胞中OPG mRNA的表达,同时抑制RANKL的mRNA的表达。

Effects of calcitonin on osteoblast cell proliferation and OPG/RANKL expression: experiment with mouse osteoblasts

OBJECTIVE: To investigate the effects of calcitonin on the proliferation of osteoblasts and the OPG/RANKL expression therein. METHODS: Osteoblasts were separated by enzyme digestion methods from the cranial bones of 30 newborn mice, cultured, and inoculated in 96-well plate. Calcitonin of the concentrations of 10, 40, and 80 IU/L was administrated into the culture medium for 24, 48, and 72 hours. The proliferation of osteoblasts was detected by MTT method. Flow cytometry was used to analyze the cell cycle. The mRNA expression of osteoprotegerin (OPG) and RANKL and protein expression of OPG were examined by RT-PCR and ELISA respectively. RESULTS: The proliferation rates of the osteoblasts exposed to the calcitonin of the concentrations of 40 and 80 IU/L for 24. 48, and 72 hours cell were all significantly higher than those of the blank control group (P < 0.05, P < 0.01). The index of osteoblasts at the G1 phase was significantly lower and the indexes of the osteoblasts at the S phase and G2-M phase were significantly increased after exposure to calcitonin (all P < 0.01). The OPG mRNA expression of the osteoblasts exposed to 80 IU/L calcitonin was significantly higher and the RANKL mRNA expression was significantly lower compared with the control group (P < 0.05). The OPG protein expression of the osteoblasts exposed to 80 LU/L calcitonin was significantly higher than that of the control group at different time points (P < 0.05, P < 0.01). CONCLUSION: Calcitonin increases the osteoblast proliferation and the OPG mRNA expression in the osteoblasts while RANKL mRNA expression in the osteoblasts is inhibited.