慢性间歇低氧对小鼠骨密度和骨代谢的影响

摘要

目的：探讨慢性间歇低氧（CIH）对骨密度（BMD）和骨代谢的影响。方法将16只12周龄C57BL/6雄性小鼠按随机数字表法分为慢性间歇低氧组（CIH组）和正常对照组（CON组），每组8只。CON组小鼠在正常环境下饲养，CIH组小鼠在间歇低氧环境下饲养，低氧暴露时间为每日9：00～17：00，6d/周，共8周。于8周末取血，用ELISA法测定血清抗酒石酸酸性磷酸5b （TRACP-5b）、骨钙素（OC）、白介素-6（IL-6）和肿瘤坏死因子-α（TNF-α）水平；取右侧股骨检测其BMD；取左侧股骨观察骨组织形态学改变，并分别采用免疫组化和real- time qPCR法检测骨保护蛋白（OPG）和核因子-κβ受体活化因子配体（RANKL）和其mRNA表达。结果与CON组相比，CIH组小鼠BMD、血清OC水平及OPG/RANKL mRNA比值均明显下降（P＜0.05或0.01），而血清TRACP-5b、IL-6和TNF-α水平、骨组织内RANKL和mRNA表达均明显升高（P＜0.05或0.01）；但骨组织内OPG和mR-NA表达差异均无统计学意义（均P＞0.05）；小鼠股骨干骺端骨小梁分布稀疏，变细及断裂。血清IL-6水平与BMD呈负相关（P＜0.05），与骨组织RANKL mRNA呈正相关（P＜0.01）；血清TNF-α水平与BMD呈负相关（P＜0.01），与骨组织RANKL mRNA呈正相关（P＜0.05）。结论 CIH能导致骨吸收增加、骨形成下降和BMD下降，与低氧以及CIH引起的IL-6、TNF-α水平升高导致RANKL升高有关。%Objective To investigate the effects of chronic intermittent hypoxia (CIH) on bone mineral density (BMD) and bone metabolism in mice. Methods Sixteen male C57BL/6 mice aged 12 weeks were randomly assigned into 2 groups:CON group (n=8) and CIH group (n=8). Mice in CIH group were exposed to hypoxic environment 8h ×6d/week, while mice in CON group were exposed to normal environment. At the end of 8 weeks al animals were sacrificed. Serum levels of OC,TRACP- 5b, IL- 6 and TNF- αwere measured by ELISA;histopathology of bone tissue was observed by H.E staining and BMD examination was performed.The expression of OPG and RANKL protein and mRNA in bone tissue was detected by immunohistochemical staining and Real time- qPCR, respectively. Results Histopathology showed that in the metaphysis of distal femurs of CIH mice the bone trabecula became thinner and broken resulting in widening the intertrabecula space. Compared to CON group the BMD, serum OC level and ratio of OPG/RANKL were decreased significantly in the distal femurs of CIH group (P<0.05 or 0.01), while the serum TRACP- 5b,IL- 6 and TNF- αlevel and the expression of RANKL protein and RANKL mRNA in bone were increased significantly (P<0.05 or 0.01). But there were no significant differences in expression of OPG protein and OPG mRNA in bone be-tween two groups(P>0.05). The serum IL- 6 and TNF- αlevels were negatively correlated with BMD(P<0.05 or 0.01), and posi-tively correlated with the expression of RANKL mRNA in bone tissue (P<0.05 or 0.01). Conclusion The results suggest that chronic intermittent hypoxia can increase bone resorption, decrease bone formation and bone mineral density, which are associ-ated with up- regulation of IL- 6 and TNF- αand higher RANKL level.