金刚丸对骨质疏松大鼠骨代谢和血清骨钙素的影响

Effects of Jingang Pill on bone metabolism and serum osteocalcin in osteoporosis rats

目的探讨金刚丸对骨质疏松大鼠骨代谢和血清骨钙素的影响。方法将60只SPF级大鼠随机分为正常对照组、模型组及高、中、低剂量组和己烯雌酚组,每组10只。高、中、低剂量金刚丸组每天分别以3.24、1.62、0.81 g/kg的剂量灌胃1次;己烯雌酚组大鼠每天给予1.0 mg/kg灌胃1次;正常对照组及模型组给予等体积的生理盐水。给药6周后取股骨,测量股骨最大载荷和断裂载荷;HE染色法观察骨小梁结构;ELISA法测定股骨组织中血清骨源性碱性磷酸酶(bone alkaline phosphatase, BALP)、雌二醇(estradiol E2)和血清骨钙素(bone gla-containing protein, BGP)、骨保护蛋白(osteoprotegerin, OPG)、核因子-κB受体活化因子(receptor activator of NF-κB, RANK)及核因子-κB受体活化因子配体(receptor activator of NF-κB ligand, RANKL)的水平;采用光子骨密度(bone mineral density, BMD)测试仪测定BMD;采用Western blot法检测瞬时受体电位阳离子通道亚族V成员6 (transient receptor potential cation channel subfamily V member 6, TRPV6)蛋白相对表达量。结果模型组骨小梁形态模糊、广泛断裂,骨髓腔增宽,骨细胞排列散乱;高、中、低剂量组可见骨小梁形态较模型组改善、结构较为紧密,骨髓腔减小,骨外膜排列改善。与正常对照组相比,模型组体质量、BALP、BGP、RANK、RANKL水平明显升高,E2、BMD、最大负荷、断裂负荷、TRPV6、OPG水平均明显降低(P<0.05)。与模型组相比,高、中、低剂量组体质量、BALP、BGP、RANK、RANKL水平降低,E2、BMD、最大负荷、断裂负荷、OPG、TRPV6水平显著升高(P<0.05)。结论金刚丸通过提高绝经后骨质疏松大鼠雌激素水平,调节骨代谢,提高生物力学性能和BMD,起到抗骨质疏松的疗效和骨保护作用,其机制可能是通过影响TRPV6表达上调OPG表达,下调RANKL表达。

Objective To investigate the effects of Jingang Pill on bone metabolism and serum osteocalcin in osteoporosis rats. Methods 60 SPF rats were randomly divided into control group, model group, high, medium and low dose Jingang Pill groups and diethylstilbestrol group, with 10 rats in each group. The high, medium and low dose Jingang Pill groups were given intragastric administration of 3.24, 1.62, 0.81 g/kg per day respectively;the rats in the diethylstilbestrol group were given diethylstilbestrol1.0 mg/kg every day;rats in the control group and model group were given equal volume of normal saline. After 6 weeks of administration,the femur was taken to detect the maximum load and fracture load;bone trabecular structure was observed by HE staining;the levels of serum bone alkaline phosphatase(BALP), estradiol(E2) and serum bone gla-containing protein(BGP), osteoprotegerin(OPG),receptor activator of NF-κB(RANK) and receptor activator of NF-κB ligand(RANKL) were measured by ELISA;bone mineral density(BMD) was measured by BMD tester;the relative expression of transient receptor potential cation channel subfamily V member 6(TRPV6) protein was detected by Western blot. Results In the model group, bone trabecular morphology was blurred and extensively fractured, bone marrow cavity was widened, and bone cells were scattered. Compared with the model group, bone trabecular morphology and structure were improved in high, medium and low dose groups, bone marrow cavity was reduced, and bone membrane arrangement was improved. Compared with the control group, the levels of body weight, BALP, BGP, RANK and RANKL in model group were significantly increased, while the levels of E2, BMD, maximum load, breaking load, OPG and TRPV6 were significantly decreased(P <0.05). Compared with the model group, the levels of body weight, BALP, BGP, RANK and RANKL in medium and high dose groups were significantly increased, while the levels of E2, BMD, maximum load, breaking load, OPG and TRPV6 were significantly decreased(P<0.05). Conclusion Jingang Pill has the effect of anti-osteoporosis and bone protection by increasing the estrogen level, regulating bone metabolism, improving biomechanical properties and BMD in postmenopausal osteoporosis rats, and mechanism may be through affecting the expression of TRPV6, so as to up-regulate the expression of bone protective protein and down-regulate the expression of RANKL.