运动性骨疲劳大鼠0PG／RANKL／RANK系统和雌激素变化的实验研究

The Experiment on the changes of OPG/RANKL/RANK system and E2 in exersize bone fatigue

目的研究骨疲劳大鼠OPG／RANKL／RANK系统和雌激素的变化，为寻找骨疲劳早期预警指标提供参考。方法雌性SD大鼠适应性训练1周后随机分为对照组（n=10）和训练组（n=lO）。用Instron5800双立柱万能材料试验机测定胫骨骨刚度判定骨疲劳是否建模成功。用HOLOGICDiscoveryW（S／N87901）双能骨密度检测仪检测骨密度，用荧光实时定量RT—PCR和WesternBla进行骨组织RANKL、OPG和ERa的mRNA测定和蛋白相对定量，ELISA检测血清RANKL、OPG和E2。结果6周训练后，训练组比对照组的骨刚度下降27．43％，提示大鼠骨疲劳建模成功。训练组骨密度【（0．247±0．011）g／cm。1低于对照组【（O．266±0．004）g，cm。1（t=4．244，P=0．001）。骨疲劳大鼠骨组织OPGmRNA和蛋白，ERa蛋白低于对照组，RANKLmRNA和蛋白高于对照组，差异均有统计学意义。训练组大鼠的血清OPG、E2及OPG／RANKL比值低于对照组，RANKL水平高于对照组，差异均有统计学意义。血清OPG与血清E2、骨组织OPG呈正相关；血清E2与骨组织OPG、ERa呈正相关关系。结论骨疲劳大鼠血清E2降低，骨吸收增强，血清OPG、RANKL和E2可作为运动性骨疲劳发生的早期预警指标。

Objects To study the changes of OPG/RANKL/RANK system in rats bone fatigue,and to provide the reference for early alrming indexes of rats bone fatigue. Methods The female SD rats were randomly divided into 2 groups. Instron 5800 three-point bending manchine was used for tibia bmechanical testing. Tibia bone density was measured by HOLOGIC Discovery W （S/N 87901 ）dual-energy X-rays.The RANKL,OPG and ERot mRNA expression was detected by real-time PCR the relative protein amount was detectd by Western Blot assay,and the serum OPG, RANKL and E2 levels were detected by ELISA. Resuhs After heavy treadmill running of six weeks, the bones＇ stiffness of rat tibia decreases 27.43%. The tibia bone density of training group was （0.247 ±0.011 ） g/cm2, it was signicantly lower than the control group＇ s（0.266±0.004） g/cm2.The amount of OPG mRNA and protein, ERoL protein in tibia ,and the level of serum OPG, E2 and OPG/RANKL of the bone fatigue rats were signiticantly lower the contol groups. However, the amout of RANKL mRNA and protein, and the serum RANKL level showed the adverse relation. Correlation analysis showed a significant positive correlation between the level of serum OPG and the serum E2 level and the tibia OPG respectively, and the significant positive Correlation were also fiound between the serum E2 and the amout of tibia OPG and ERa separately. Conclusions The serum E2 level would decreased in bone fatigue rats,and their tibia bone resorption was enhanced too.The level of serum OPG, RANKL and E2 can be used for early alarm biomarkers of exersize bone fatigue.