电针对佐剂性关节炎大鼠膝关节滑膜VEGF/Vav2/Rac1信号通路的影响

Effects of electroacupuncture on VEGF/Vav2/Rac1 signaling pathway in knee joint synovium of adjuvant arthritis rats

目的观察电针对佐剂性关节炎(adjuvant arthritis,AA)大鼠膝关节滑膜组织内血管内皮生长因子(vascular endothelial growth factor,VEGF)、鸟嘌呤核苷酸交换因子2(vav guanine nucleotide exchange factor 2,Vav2)和Ras相关C3肉毒素底物1(Ras-related C3 botulinum toxin substrate 1,Rac1)表达的影响,探究电针干预类风湿关节炎(rheumatoid arthritis,RA)血管新生的可能机制。方法将40只SPF级SD雄性大鼠随机分成4组(空白组、模型组、西药组和电针组),每组10只。除空白组,其他3组采用尾根部皮下注射完全弗氏佐剂制成AA大鼠模型。造模后第2天开始干预,电针组进行电针干预,以左侧足三里与关元穴配穴,右侧足三里与同侧阿是穴配穴,每次20 min,每天干预1次,共干预21次;西药组给予甲氨蝶呤灌胃给药,每周1次,共干预3次。观察各组体质量、关节红肿等一般情况,每3天测量1次大鼠后双侧足跖容积。干预结束后,通过大鼠一般情况、体质量、足跖容积分析各组大鼠足跖肿胀程度,HE染色观察AA大鼠膝关节滑膜组织的病理形态学,免疫组织化学法检测膝关节滑膜组织内CD31表达变化,Western blot法检测大鼠膝关节滑膜组织VEGF、Vav2和Rac1蛋白表达量。结果与空白组相比,模型组大鼠第21天足跖容积均明显增大(P<0.01),膝关节滑膜组织出现显著性病理改变,CD31表达量明显升高(P<0.05),VEGF、Vav2、Rac1蛋白表达量显著升高(P<0.01)。与模型组相比,西药组足跖容积在第21天显著减小(P<0.01),膝关节滑膜组织病理改变明显改善,VEGF、Vav2表达量明显下降(P<0.05),Rac1蛋白表达量显著下降(P<0.01);电针组足跖容积在第21天显著减小(P<0.01),膝关节滑膜组织病理改变明显改善,VEGF、Rac1表达量明显下降(P<0.05),Vav2蛋白表达量显著下降(P<0.01)。结论电针改善RA的作用机制可能与抑制VEGF/Vav2/Rac1信号通路,进而抑制血管新生有关。

Objective To observe the effects of electroacupuncture on the expressions of vascular endothelial growth factor(VEGF),Vav guanine nucleotide exchange factor 2(Vav2),and Ras-related C3 botulinum toxin substrate 1(Rac1)in knee joint synovial tissue of adjuvant arthritis(AA)rats,so as to investigate the possible mechanism of electroacupuncture intervention on angiogenesis in rheumatoid arthritis(RA).Methods A total of 40 SPF SD male rats were randomly divided into blank group,model group,western medicine group,and electroacupuncture group,with 10 rats in each group.Except for the blank group,the other three groups were treated with subcutaneous injection of complete Freund's adjuvant at the base of the tail to establish AA rats model.Two days after model establishment,the electroacupuncture group were given electroacupuncture at left"Zusanli"(ST36)with"Guanyuan"(BL26),right"Zusanli"(ST36)with ipsilateral"Ashi"(experience acupoint)for 20 min each time,once a day for 21 consecutive days.The rats in western medicine group were given methotrexate by gavage once a week,for a total of 3 times.The body weight,redness and swelling of joints,and other general conditions in each group were observed and the plantar volume was measured every 3 days.After the intervention,the degree of plantar swelling in each group was analyzed by the general conditions,body weight,redness and swelling of joints,and plantar volume of rats.Hematoxylin-eosin staining was used to observe the pathological morphology of knee joint synovial tissue of AA rats,Immunohistochemistry was used to measure the expression change of CD31 in knee joint synovial tissue,and the protein expression levels of VEGF,Vav2,and Rac1 were determined by Western blot.Results Compared with the blank group,the rats'plantar volume in the model group significantly increased on the 21st day(P<0.01),the knee joint synovial tissue showed significant pathological changes,the expression level of CD31 was significantly higher(P<0.05),and the protein expression levels of VEGF,Vav2,and Rac1 were significantly higher(P<0.01).Compared with the model group,the rats'plantar volume in the western medicine group significantly decreased on the 21st day(P<0.01),the pathological changes of knee joint synovial tissue were significantly improved,the expression levels of VEGF and Vav2 were significantly lower(P<0.05),and the protein expression level of Rac1 was significantly lower(P<0.01);the rats'plantar volume in electroacupuncture group significantly decreased on the 21st day(P<0.01),the pathological changes of knee joint synovial tissue were significantly improved,the expression levels of VEGF and Rac1 were significantly lower(P<0.05),and the protein expression level of Vav 2 was significantly lower(P<0.01).Conclusion The mechanism of electroacupuncture in improving RA may be related to the inhibition of VEGF/Vav2/Rac1 signaling pathway,thereby inhibiting angiogenesis.