目的 观察温阳补肾方对兔激素性股骨头坏死(SANFH)组织中破骨细胞抑制因子(OPG)、核因子-κB受体活化因子(RANK)和核因子-κB受体活化因子配体(RANKL)的mRNA表达的影响。方法 普通级健康新西兰大白兔46只，分为正常组(n = 10)和造模组(n = 36)。应用改进马血清联合甲基强的松龙法制作SANFH模型。造模后随机选取正常组2只、造模组4只处死，用于模型鉴定。再将造模组剩余动物随机分为模型组(n = 8)，中药低(n = 8)、中(n = 8)和高(n = 8)剂量组。正常组和模型组生理盐水10 ml/d灌胃。中药低、中、高剂量组分别为以6.44 g/(kg?d)、9.66 g/(kg?d)、12.88 g/(kg?d)灌胃温阳补肾方汤剂，连续8周。采用逆转录聚合酶链反应检测OPG、RANK和RANKL的mRNA表达。结果 模型组空骨陷窝率显著高于正常组(t = 17.085, P < 0.001)。与模型组比较，中药各剂量组OPG mRNA表达均明显升高(P < 0.01)，RANK和RANKL的mRNA表达明显降低(P < 0.01)；与中药低剂量组比较，中药中、高剂量组OPG mRNA表达明显升高(P < 0.01)，RANK和RANKL的mRNA表达明显降低(P < 0.01)；中药中、高剂量组比较均无显著性差异(P > 0.05)。结论 温阳补肾方能提高兔SANFH股骨头组织中OPG的mRNA表达，抑制RANK和RANKL的mRNA表达。
Objective To observe the effect of Wenyangbushen formula on mRNA expression of osteoprotegerin (OPG), receptor activator of nuclear factor- kappa B (RANK) and receptor activator of nuclear factor- kappa B ligand (RANKL) in rabbits with steroid-induced avascular necrosis of femoral head (SANFH). Methods A total of 46 healthy conventional New Zealand white rabbits were randomly divided into normal group (n = 10) and model building group (n = 36). The modified method of horse serum plus methylprednisolone was used to establish the SANFH model. Two rabbits from the normal group and four from the model building group were used for HE staining. Then the other models were randomly divided into model group, and low-dose, medium-dose and high-dose treatment groups, with eight rabbits in each group. The normal group was given normal saline 10 ml/d, and the treatment groups were given Wenyangbushen formula 6.44 g/(kg?d), 9.66 g/(kg?d) and 12.88 g/(kg?d), respectively, for eight weeks. The mRNA expression of OPG, RANK and RANKL was detected by reverse transcription- polymerase chain reaction. Results The empty lacuna rate was significantly higher in the model group than in the normal group (t = 17.085, P < 0.001). Compared with the model group, the mRNA expression of OPG increased (P < 0.01), and the mRNA expression of RANK and RANKL decreased (P < 0.01), in the treatment groups. Compared with the low-dose treatment group, the mRNA expression of OPG increased (P < 0.01), and the mRNA expression of RANK and RANKL decreased (P < 0.01), in the medium-dose and high-dose treatment groups. There was no significant difference in the mRNA expression of OPG, RANK and RANKL between the low-dose treatment group and the high-dose treatment group (P > 0.05). Conclusion Wenyangbushen formula could increase the mRNA expression of OPG and inhibit the mRNA expression of RANK and RANKL in the femoral head tissue of the rabbits with SANFH.
[1] 胥少汀,葛宝丰,徐印坎. 实用骨科学[M]. 4版. 北京:人民军医出版社, 2012: 1857-1879.
[2] 梁川,康庆林,贾亚超,等. 骨髓间充质干细胞：激素性股骨头坏死早期治疗的新希望[J]. 中国矫形外科杂志, 2018, 26(9): 821-825.
[3] 陶静,柳维林,黄佳,等. 基于Notch1信号通路观察电针对脑缺血再灌注损伤大鼠缺血周边皮质与SVZ区神经干细胞增殖的影响[J]. 康复学报, 2015, 25(3): 23-34.
[4] 陈宏伟,包丽亚,黄月,等. 清眩降压汤控制自发性高血压大鼠血压升高与抑制心肾大血管损害的药效学研究[J]. 康复学报, 2017, 27(2): 28-32.
[5] 王伟,刘利英,王坤正,等. 激素性股骨头坏死模型的建立及其发病机理的探讨[J]. 西安交通大学学报(医学版), 2007, 28(5): 544-547.
[6] 施新猷. 医用实验动物学[M]. 西安:陕西科学技术出版社, 1989: 418.
[7] 颜冰. 活血补肾汤对激素性股骨头坏死VEGF及OPG/RANKL表达的影响[D]. 福州:福建中医药大学, 2010: 7-14.
[8] 余航. 髓芯减压联合红花提取物对兔股骨头缺血坏死的治疗作用[D]. 福州:福建中医药大学, 2012: 8.
[9] Jiang Y, Liu D, Kong X, et al. Huogu I formula prevents steroid-induced osteonecrosis in rats by down-downregulating PPAR gamma expression and activating wnt/LRP5/beta-catenin signaling [J]. J Tradit Chin Med, 2014, 34(3): 342-350.
[10] Boquete-castro A, Gómez-moreno G, Calvo-guirado J L, et al. Denosumab and osteonecrosis of the jaw.A systematic analysis of events reported in clinical trials [J]. Clin Oral Implants Res, 2016, 27(3): 367-375.
[11] Rupp R E, Rupp S N. Femoral head avascular necrosis is not caused by arthroscopic posterolateral femoroplasty [J]. Orthopedics, 2016, 39(3): 177-180.
[12] Liu G, Luo G, Bo Z, et al. Impaired osteogenic differentiation associated with connexin43/ microRNA-206 in steroid-induced avascular necrosis of the femoral head [J]. Exp Mol Pathol, 2016, 101(1): 89-99.
[13] Wang Y, Cao Y, LI Y, et al. Genetic association of the ApoB and ApoA1 gene polymorphisms with the risk for alcohol-induced osteonecrosis of femoral head [J]. Int J Clin Exp Pathol, 2015, 8(9): 11332-11339.
[14] Yu T, Xie L, Chu F. A sclerotic rim provides mechanical support for the femoral head in osteonecrosis [J]. Orthopedics, 2015, 38(5): e374-e379.
[15] 王亚军,浪万英,宋亚文,等. 针刺血清对成骨细胞OPG、RANKL mRNA表达的影响[J]. 中国骨质疏松杂志, 2017, 23(10): 1334-1339.
[16] 吴佳莹,李岳泽,刘红,等. “肾主骨”机理研究——左归丸对Hepcidin、Fpn1及OPG/RANKL mRNA表达的影响[J]. 中国中医基础医学杂志, 2017, 23(11): 1548-1551.
[17] Park E K, Kim M S, Lee S H, et al. Furosin, an ellagitannin, suppresses RANKL-induced osteoclast differentiation and fumetion through inhibition of MAP kinase activation and actin ring formation [J]. Biochem Biophys Res Commun, 2004, 325(4): 1472-1480.
[18] 张文信,秦亚梅,党少平. 补肾活血法治疗激素性股骨头坏死理论探析[J]. 新中医, 2012, 44(8): 193-194.
[19] 宋红梅,吴斌,魏迎辰,等. 温阳补肾方对激素性股骨头坏死模型兔股骨头形态学的影响[J]. 中国中医骨伤科杂志, 2014, 22(3): 1-3.
[20] 陈卫衡,王和鸣. 温阳补肾中药促进骨髓基质细胞分化的实验研究进展[J]. 中国骨伤, 2011, 24(4): 352-356.
[21] 黄胜杰,李媚,王和鸣. 温阳补肾药对骨髓间充质干细胞促增殖的实验研究[J]. 中国中医骨伤科杂志, 2012, 20(10): 1-8.
[22] 杨黎丽,黄胜杰,李媚,等. 温阳补肾药对抗骨髓间充质干细胞凋亡的实验研究[J]. 中医正骨, 2013, 25(2): 3-7.
[23] 宋红梅,吴斌,魏迎辰,等. 温阳补肾方对兔激素性股骨头坏死血清OPG、RANK、RANKL的影响[J]. 中国中医骨伤科杂志, 2013, 21(12): 1-7.
[24] 魏迎辰. 温阳补肾方治疗兔激素性股骨头坏死的实验研究[D]. 福州:福建中医药大学, 2014: 8.
[25] 宋红梅. 温阳补肾方对兔激素性股骨头坏死VEGF及RANKL/RANK/OPG通路的影响[D]. 福州:福建中医药大学, 2015: 8.
[26] 宋红梅,魏迎辰,吴斌,等. 温阳补肾方对兔激素性股骨头坏死组织OPG-RANK-RANKL通路的影响[J]. 中华中医药杂志, 2016, 31(1): 302-305.
[27] Song H, Wei Y, Li N, et al. Wenyangbushen-induced expression of VEGF, OPG, RANK, and RANKL in rabbits with steroid-induced femoral head avascular necrosis [J]. Genetika, 2015, 47(2): 477-488.
