补肾活血汤对激素性股骨头坏死模型大鼠神经肽Y系统的影响

Effect of Bushen Huoxue Decoction(补肾活血汤)on Neuropeptide Y System in Rats Model with Steroidinduced Osteonecrosis of Femoral Head

目的探讨补肾活血汤治疗激素性股骨头坏死(SONFH)的可能作用机制。方法32只SD大鼠随机分为对照组、模型组、补肾活血汤组和拮抗剂组,每组8只。除对照组外,其余各组依次尾静脉注射内毒素(20μg/kg)2次和臀部肌肉注射强的松(40mg/kg)3次,每次间隔24h,制备SONFH大鼠模型。末次肌肉注射24h后,补肾活血汤组大鼠给予补肾活血汤6.3ml/(kg·d)灌胃,对照组和模型组用等量生理盐水灌胃,拮抗组大鼠给予JNJ-31020028(10mg/kg)皮下注射。灌胃和皮下注射均每周3次,持续8周。采用HE染色观察股骨头病理学改变,计算空骨陷窝率及骨小梁面积占比;ELISA法检测下丘脑神经肽Y(NPY)、Y2受体(Y2R)和股骨头NPY、Y1受体(Y1R)含量;Western Blot法检测下丘脑和股骨头NPY以及股骨头B淋巴细胞瘤2(Bcl-2)、Bcl-2相关x蛋白(Bax)表达。结果HE染色可见模型组骨小梁稀疏、破坏,多见空骨陷窝和脂肪空泡,补肾活血方组和拮抗组病理损伤明显减轻;与模型组比较,补肾活血汤组和拮抗组大鼠空骨陷窝率下降,骨小梁面积占比上升(P<0.01)。与对照组比较,模型组大鼠下丘脑NPY、Y2R和股骨头NPY含量及蛋白表达升高,Bcl-2下降、Bax上升(P<0.05或P<0.01);与模型组比较,补肾活血汤组和拮抗剂组大鼠下丘脑NPY、Y2R含量及蛋白表达降低,股骨头NPY含量及蛋白表达升高,Bcl-2升高、Bax降低(P<0.01)。结论补肾活血汤可改善SONFH股骨头病理损伤,其机制可能与抑制下丘脑NPY、Y2R和增加骨组织中NPY表达,提高骨组织抗凋亡能力有关。

Objective To explore the possible mechanism of Bushen Huoxue Decoction(补肾活血汤)in the treatment of steroid-induced osteonecrosis of the femoral head(SONFH).Methods Thirty-two SD rats were randomised into control group,model group,Bushen Huoxue Decoction group and antagonist group,with eight rats in each group.The rats in all the groups except for control group were injected with twice of endotoxin(20μg/kg)in the tail vein and three times of prednisone(40 mg/kg)in the hip muscles,with an interval of 24 hours each time to prepare the SONFH rat model.Twenty-four hours after the last intramuscular injection,rats in Bushen Huoxue Decoction group were administered with 6.3 ml/(kg·d)of Bushen Huoxue Decoction by gavage;same amount of normal saline was given to the control group and the model group by gavage;and JNJ-31020028(10 mg/kg)was subcutaneously injected to the antagonist group;all the interventions were administered three times per week,for eight weeks.The pathological changes of the femoral head were assessed by HE staining,and percentage of empty lacuna and bone trabecular area were calculated;ELISA method was used to assess the content of hypothalamic neuropeptide Y(NPY),Y2 receptor(Y2 R),femoral head NPY and Y1 receptor(Y1 R);Western Blot method was used to detect the expression of hypothalamic and femoral head NPY,femoral head B-cell lymphoma 2(Bcl-2),and Bcl-2 related x protein(Bax).Results The HE staining results showed that the bone trabecula was sparse and damaged in the model group,with empty bone lacuna and fat vacuoles commonly seen.The pathological damages were significantly reduced in Bushen Huoxue Decoction group and antagonist group.Compared to the model group,Bushen Huoxue Decoction group and antagonist group had lower percentage of empty bone lacuna and larger bone trabecular area(P<0.01).Compared to the control group,the model group had increased levels of hypothalamic NPY,Y2 R,femoral head NPY and protein expression,as well as decreased Bcl-2 and increased Bax(P<0.05 or P<0.01).Compared to model group,Bushen Huoxue Decoction group and antagonist group had lower hypothalamic NPY,Y2 R content and protein expression,higher femoral head NPY and protein expression,as well as higher Bcl-2 and lower Bax(P<0.01).Conclusion Bushen Huoxue Decoction can reduce the pathological damages of femoral head in SONFH rats,which may be related to its function of inhibiting hypothalamic NPY and Y2 R,promoting NPY expression in bone tissues,and increasing anti-apoptotic ability of bone tissues.