补肾活血汤干预骨-血耦连改善大鼠激素性股骨头坏死的作用机制研究

Mechanism of Bushen Huoxue Decoction in Improving Steroid-Induced ONFH in Rats by Intervening‘Bone-Blood Coupling’

目的:探讨补肾活血汤改善激素性股骨头坏死大鼠的作用机制。方法:64只雄性SD大鼠随机分成对照组、模型组、阳性药物组和补肾活血汤低、中、高药物组(简称低、中、高剂量组)。除对照组外,其余组别均建立激素性股骨头坏死疾病模型,药物组分别给予对应的药物及剂量干预8周,干预后收集大鼠股骨组织,采用micro-CT检测股骨形态;HE染色观察股骨头形态;Western blot、RT-qPCR检测BMP2、VEGFA、DLL4蛋白表达;免疫荧光检测股骨髓组织Osterix、vWF表达情况。结果:与对照组相比,模型组大鼠股骨内骨小梁紊乱,骨小梁面积比明显减小(P<0.05),空骨陷窝率明显增加(P<0.05),BMP2、VEGFα蛋白表达明显下调(P<0.05),DLL4蛋白表达明显上升(P<0.05),免疫荧光下可见骨髓中Osterix、vWF分泌减少;各治疗组骨小梁紊乱改善,除低剂量组外骨小梁面积比明显增大(P<0.05),空骨陷窝率明显增加(P<0.05),BMP2、VEGFα蛋白表达明显上调(P<0.05),DLL4蛋白表达明显下调(P<0.05);高剂量组免疫荧光中可见Osterix、vWF分泌较模型组明显增加。结论:补肾活血汤能促进激素性股骨头坏死大鼠股骨头内成骨、成血管分化,从而改善股骨头坏死病程。

Objective:To investigate the mechanism of Bushen Huoxue Decoction in improving steroid-induced osteonecrosis of the femoral head(ONFH)in rats.Methods:64 SD male rats were randomly divided into the control group,the model group,the positive drug group,and the Bushen Huoxue Decoction groups of low-dose,medium-dose and high-dose.Except the rats in the control group,the rest were established the model of steroid-induced ONFH in the other groups.Corresponding interventions had been carried out for 8 weeks.After 8 weeks of intervention,femoral heads were collected,and micro-CT was performed to detect the femoral head;HE staining was performed to observe the morphology of femoral head;Western blot and RT-qPCR were used to detect the protein expressions of BMP2,VEGFA and DLL4;immunofluorescence was used to detect the expressions of Osterix and vWF in femoral bone marrow tissues.Results:Compared with those in the control group,the trabeculae in the femur were disordered and the trabecular area was significantly reduced(P<0.05);the number of empty bone traps was significantly increased(P<0.05);the protein expressions of BMP2 and VEGFαwere significantly down-regulated(P<0.05);the protein expression of DLL4 was significantly increased(P<0.05);the secretions of Osterix and vWF in the bone marrow were decreased under immunofluorescence observation in the model group.The bone trabecular disorder was improved in all intervention groups;the trabecular area was increased significantly except the low-dose Bushen Huoxue Decoction group(P<0.05);the number of empty bone traps was increased significantly(P<0.05);the protein expressions of BMP2 and VEGFαwere up-regulated significantly(P<0.05);the protein expression of DLL4 was down-regulated significantly(P<0.05);the secretions of Osterix and vWF were increased under immunofluorescence observation in the high-dose Bushen Huoxue Decoction group.Conclusion:Bushen Huoxue Decoction can promote osteogenic and vascular differentiation within the femoral head of the rats with steroid-induced ONFH,thus to improve the course of femoral head necrosis.