短刺加电针法对膝骨关节炎兔膝关节软骨修复的影响

目的:观察短刺加电针法对膝骨关节炎兔软骨γ-谷氨酰基羧化酶(GGCX)、基质金属蛋白酶(MMP)13及血清非羧化基质GLA蛋白(ucMGP)的影响,探讨短刺加电针法对软骨组织的修复作用。方法:新西兰大白兔随机分为正常组10只和造模组30只,造模组动物采用Hulth-Telhag法手术复制膝关节炎模型,将造模成功的动物随机分为模型组、短刺组、普通针刺组,每组10只。短刺组取左侧"内膝眼""外膝眼""阴陵泉""足三里""梁丘",采用短刺法加用电针,普通针刺组常规针刺以上穴位加用电针,均每日治疗1次,每次20min,5d为一疗程,共4个疗程。采用蛋白免疫印迹法检测软骨细胞中GGCX、MMP 13的蛋白表达;免疫组化法检测GGCX的活性;酶联免疫吸附测定法检测血清ucMGP的含量。结果:与正常组相比,模型组膝关节软骨中MMP 13、血清ucMGP表达明显提高(P0.01,P0.05),而GGCX表达量下降(P0.01);与模型组相比,短刺组和普通针刺组MMP 13、ucMGP表达明显降低(P0.01,P0.05),而GGCX表达升高(P0.01);与普通针刺组相比,短刺组MMP 13、ucMGP表达明显降低(P0.01,P0.05),而GGCX表达升高(P0.01)。结论:普通电针法与短刺加电针法均可促进膝关节软骨细胞修复,且短刺法有一定的优势,其治疗作用可能与上调GGCX表达,促进MGP羧基化,抑制MMP 13表达有关。

Effects of Short Thrust Needing plus Electroacupuncture Intervention on Cartilage Tissue in Rabbits with Knee Osteoarthritis

Objective To observe the effectiveness of short thrust needling(STN,close-to-bone needing)plus electroacupuncture(EA)in healing knee cartilage tissue and in regulating expressions of cartilage vitamin K dependent gamma-glutamyl carboxylase(GGCX),matrix metalloproteinase-13(MMP 13)and serum uncarboxylated matrix gla protein(ucMGP)in rabbits with knee osteoarthritis(KOA),so as to reveal its mechanism underlying improvement of KOA.Methods Forty New Zealand rabbits were randomly divided into normal,model,EA and STN+EA groups(n=10in each group).The KOA model was created by cutting the medial lateral ligament and medial parapatellar arthrotomy of rabbits as described by Hulth and colleagues.For rabbits in the STN+EA group,"Neixiyan"(EX-LE 4)and"Waixiyan"(ST 35)were punctured with filiform needles by controlling the needle-tip obliquely to advance till the bone surface of the knee joint cavity,and"Yinlingquan"(SP 9)and"Zusanli"(ST 36)punctured by holding the filiform needles vertically along the tibia,and"Liangqiu"(ST 34)was punctured by controlling the filiform needle to advance till the thigh-bone,followed by EA stimulation.EA(2Hz/100 Hz,1-3mA)was applied to unilateral EX-LE 4and ST 35,and ST 36 and SP 9,separately for 20 min,once daily for 20 days except weekends.The pathological changes of the knee cartilage cells were observed using H.E.staining,Toluidine blue staining and electron transmission microscope,respectively.The immunoactivity of GGCX of the knee cartilage was determined by immunohistochemistry and the expression levels of GGCX and MMP 13 proteins in the cartilage were detected by Western blot,and the content of serum ucMGP was assayed by ELISA.Results H.E.staining,Toluidine blue staining and electron transmission microscope results showed that pathological changes of knee cartilage cells in structure after modeling were improved in both the STN+EA and EA groups,particularly the former group.In comparison with the normal group,the expression levels of GGCX protein in the cartilage tissue showed by both Western blot and immunohistochemistry were notably down-regulated(P<0.01),and the cartilage MMP 13 protein expression and serum ucMGP content were considerably up-regulated in the model group(P<0.01,P<0.05).After STN+EA and simple EA,the decreased GGCX and the increased MMP 13 expression and serum ucMGP content were reversed(P<0.01,P<0.05).The effects of STN+EA were significantly superior to those of simple EA in down-regulating MMP13 and ucGLA levels,and upre-gulating GGCX expression.Conclusion Both STN+EA and simple EA can effectively improve pathological changes of cartilage cells in KOA rabbits,which may be associated with their actions in up-regulating the expression of cartilage GGCX protein and lowering the levels of serum ucMGP content and cartilage MMP 13 protein expression,and the effects of STN+EA are better.