目的 研究淫羊藿苷聯(lián)合GM6001治療酒精性股骨頭壞死大鼠的作用機制，同時探究其對骨保護素（OPG）/核因子-κB受體活化因子（RANK）/核因子-κB受體活化因子配體（RANKL）通路的調(diào)節(jié)作用。方法 大鼠隨機分為對照組、模型組、淫羊藿苷組、GM6001組及聯(lián)合組（淫羊藿苷＋GM6001），每組12只。采用ig給予大鼠白酒建立酒精性股骨頭壞死大鼠模型，淫羊藿苷組按照60 mg/kg ig給予大鼠淫羊藿苷，GM6001組按照100 mg/kg尾iv GM6001，聯(lián)合組同時給予淫羊藿苷（60 mg/kg）及GM6001（100 mg/kg），1次/d，連續(xù)8周，對照組及模型組給予生理鹽水。曠場實驗測定大鼠活動次數(shù)、活動總距離，抓力測定儀測定大鼠最大抓力；顯微CT儀測定股骨頭骨體積分?jǐn)?shù)、骨小梁間距、骨小梁數(shù)目、骨小梁厚度；酶聯(lián)免疫吸附法（ELISA）測定血清磷、鈣水平，血清及股骨頭骨形成蛋白-2（BMP-2）、轉(zhuǎn)化生長因子-β（TGF-β）、堿性成纖維細胞生長因子（bFGF）水平。蘇木精–伊紅（HE）染色法測定股骨頭組織病理學(xué)變化；實時定量聚合酶鏈?zhǔn)椒磻?yīng)（PCR）測定股骨頭OPG、RANK、RANKL mRNA水平；免疫印跡法測定股骨頭OPG、RANK及RANKL蛋白水平。結(jié)果 與模型組比較，淫羊藿苷組、GM6001組、聯(lián)合組活動次數(shù)、活動總距離、最大抓力、骨體積分?jǐn)?shù)、骨小梁數(shù)目、骨小梁厚度，血清磷、鈣、BMP-2、TGF-β、bFGF水平，股骨頭BMP-2、TGF-β、bFGF水平，股骨頭OPG mRNA及蛋白水平顯著升高（P＜0.05）；骨小梁間距、股骨頭RANK和RANKL的mRNA及蛋白水平顯著降低（P＜0.05），且聯(lián)合組降低更為顯著（P＜0.05）。結(jié)論 淫羊藿苷聯(lián)合GM6001能夠顯著修復(fù)大鼠酒精性股骨頭壞死，緩解股骨頭組織病理學(xué)改變，其機制可能與調(diào)節(jié)OPG/RANK/RANKL有關(guān)。

Objective To investigate the mechanism of icariin combined with GM6001 in treatment of alcohol-induced osteonecrosis of the femoral head in rats, and to explore its regulatory effect on OPG/RANK/RANKL system. Methods Rats were randomly divided into control group, model group, icariin group, GM6001 group, and combined group (icariin + GM6001), with 12 rats in each group. Rats in icariin group were given icariin 60 mg/kg by gavage, GM6001 group were given GM6001 100 mg/kg by tail vein, rats in combination group were given icariin 60 mg/kg and GM6001 100 mg/kg by gavage, once daily for 8 weeks. The control group and model group were given normal saline. The open field experiment was used to measure the number of activities and the total distance of activities of the rats. The maximum grasping force of the rats was measured by the grip tester. The volume fraction, trabecular spacing, trabecular number, and trabecular thickness of femoral head were measured by microCT. Serum phosphorus and calcium levels, BMP-2, TGF-β, bFGF levels in serum and femoral head were determined by enzyme-linked immunosorbent assay (ELISA).Hematoxylin and eosin (HE) staining was used to determine the histopathological changes of femoral head. Real-time quantitative polymerase chain reaction (PCR) was used to determine the mRNA levels of OPG, RANK, and RANKL in femoral head. The protein levels of OPG, RANK, and RANKL of femoral head were determined by Western blotting. Results Compared with the model group, the number of activities, total distance of activities, maximum grip, bone volume fraction, trabecular bone number, trabecular bone thickness, serum phosphorus, calcium, BMP-2, TGF-β, bFGF levels, and BMP-2, TGF-β, bFGF of serum and femoral head levels in icariin group, GM6001 group, and combined group were significantly increased. The mRNA and protein levels of OPG in femoral head were significantly increased (P < 0.05), the mRNA and protein levels of trabecular spacing, femoral head RANK and RANKL were significantly decreased (P < 0.05). And the combined group was more significant (P < 0.05). Conclusion Icariin combined with GM6001 can significantly repair alcohol-induced femoral head necrosis, and alleviate histopathological changes of femoral head in rats, and the mechanism may be related to the regulation of OPG/RANK/RANKL.

R285.5

海南省衛(wèi)生健康行業(yè)科研項目(20A200025)