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首页|期刊导航|针刺研究|热补针法对类风湿关节炎寒证家兔膝关节滑膜组织缺氧诱导因子1α和糖酵解活性的影响

热补针法对类风湿关节炎寒证家兔膝关节滑膜组织缺氧诱导因子1α和糖酵解活性的影响OA北大核心CSTPCDMEDLINE

Effect of heat-reinforcing needling on hypoxia-inducible factor 1α and glycolysis activity in rabbits with cold syndrome of rheumatoid arthritis

中文摘要英文摘要

目的:观察热补针法对类风湿关节炎(RA)寒证家兔膝关节滑膜炎性反应及滑膜组织、血清中缺氧诱导因子1α(HIF-1α)、糖酵解活性的影响,探讨热补针法治疗RA的机制.方法:选取32只新西兰兔随机分为正常组、模型组、抑制剂组、热补针法组,每组8只.采用卵蛋白联合弗氏完全佐剂诱导及低温冷冻法复制RA寒证模型.抑制剂组给予2-甲氧基雌二醇腹腔注射,热补针法组给予热补针法针刺"足三里",留针30 min.各组干预均1次/d,连续14 d.观察家兔膝关节周径及痛阈变化;采用ELISA法检测家兔血清中还原型辅酶Ⅱ(NADPH)、己糖激酶(HK2)、6-磷酸果糖激酶2/2,6-二磷酸果糖激酶3(PFKFB3)含量;HE染色法观察家兔膝关节滑膜组织形态变化;免疫组织化学法检测家兔膝关节滑膜组织中肿瘤坏死因子α(TNF-α)、白细胞介素(IL)-1β、IL-6、IL-17的阳性表达;分光光度法检测家兔膝关节滑膜组织中乳酸含量;Western blot法检测膝关节滑膜组织中HIF-1α、乳酸脱氢酶(LDHA)、丙酮酸激酶 2(PKM2)蛋白表达水平.结果:干预后,与正常组比较,模型组家兔膝关节周径增大(P<0.05),痛阈值降低(P<0.05),可见滑膜细胞中度增生及炎性浸润,滑膜组织病理评分升高(P<0.05),滑膜组织中TNF-α、IL-1β、IL-6、IL-17阳性表达及乳酸含量均升高(P<0.05),血清NADPH、HK2、PFKFB3含量及滑膜组织中HIF-1α、LDHA、PKM2蛋白表达水平均升高(P<0.05).与模型组比较,抑制剂组及热补针法组家兔膝关节周径缩小(P<0.05),痛阈值升高(P<0.05),滑膜增生、炎性细胞浸润均改善,滑膜组织病理评分降低(P<0.05),滑膜组织中TNF-α、IL-1β、IL-6、IL-17阳性表达及乳酸含量均下降(P<0.05),血清NADPH、HK2、PFKFB3含量及滑膜组织中HIF-1α、LDHA、PKM2蛋白表达水平均下降(P<0.05).与抑制剂组比较,热补针法组家兔滑膜组织病理评分升高(P<0.05),滑膜组织中TNF-α、IL-1β、IL-6、IL-17阳性表达及乳酸含量均升高(P<0.05),血清NADPH、HK2、PFKFB3含量及滑膜组织中HIF-1α、LDHA、PKM2蛋白表达水平均升高(P<0.05).结论:热补针法干预RA寒证家兔可提高痛阈,减小膝关节周径,抑制炎性反应,其作用机制与下调HIF-1α及糖酵解活性相关.

Objective To observe the effect of heat-reinforcing needling(HRN)on synovial inflammation,hypoxia-inducible factor-1α(HIF-1α)and glycolytic activity in serum and synovial tissue in rabbits with cold syndrome of rheumatoid arthritis(RA),so as to explore its mechanisms underlying improvement of RA.Methods A total of 32 rabbits were randomly divided into normal,model,inhibitor and HRN groups,with 8 rabbits in each group.The RA with cold syndrome model was induced by injecting ovalbumin dry powder and Freund's complete adjuvant combined with cold freezing.Rabbits in the inhibitor group were intraperitoneally injected with 2-methoxyestradiol(2.5 mg/kg),rabbits in the HRN group were received HRN at bilateral"Zusanli"(ST36)for 30 min.The treatments were conducted once daily for 14 consecutive days.After the interventions,the knee circumference and pain threshold were measured.The contents of nicotinamide adenine dinucleotide phosphoric(NADPH),Hexokinase II(HK2)and 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3(PFKFB3)in serum of rabbits were detected by ELISA.The pathological morphology of synovial tissue of the knee joints were observed by HE staining.The positive expressions of tumor necrosis factor(TNF-α),interleukin(IL)-1β,IL-6 and IL-17 in synovial tissue of knee joint were detected by immunohistochemistry.The content of lactic acid in synovial tissue of rabbit knee joint was detected by spectrophotometry.The expression levels of HIF-1α,pyruvate kinase 2(PKM2)and lactate dehydrogenase(LDHA)in synovial tissue of knee joint were detected by Western blot.Results After intervention,compared with the normal group,the knee circumference was significantly enlarged(P<0.05),the pain threshold was significantly decreased(P<0.05);the synovial tissue of knee joints showed significant cell proliferation and inflammatory infiltration,the pathological score was significantly increased(P<0.05);positive expressions of TNF-α,IL-1β,IL-6 and IL-17,the content of lactic acid in synovial tissue,the contents of NADPH,HK2 and PFKFB3 in serum,and the protein expression levels of HIF-1α,PKM2 and LDHA in synovial tissue were increased(all P<0.05)in the model group.Compared with model group,the circumference of knee joint was significantly decreased(P<0.05),the pain threshold was significantly increased(P<0.05);in synovial tissue,the pathological score was decreased(P<0.05);the positive expressions of TNF-α,IL-1β,IL-6 and IL-17 in synovial tissue were decreased(P<0.05),the lactic acid content in synovial tissue was decreased(P<0.05);the contents of NADPH,HK2 and PFKFB3 in serum and the protein expression levels of HIF-1α,PKM2 and LDHA in synovial tissue were decreased(P<0.05)in inhibitor group and HRN group.Compared with the inhibitor group,the synovial pathological score was significantly increased(P<0.05),positive expressions of TNF-α,IL-1β,IL-6 and IL-17,the content of lactic acid in synovial tissue,the contents of NADPH,HK2 and PFKFB3 in serum,and the protein expression levels of HIF-1α,PKM2 and LDHA in synovial tissue were increased(all P<0.05)in HRN group.Conclusion HRN can increase the pain threshold,reduce the knee circumference and inhibit the inflammatory response in rabbits with cold syndrome of RA.The possible mechanism is related to the down-regulation of HIF-1α and glycolysis activity.

李向军;李福欣;井维尧;刘翠;刘莉梅;张星华;张枫帆;陈平;杜小正

甘肃省中医院风湿骨病科,兰州 730050甘肃中医药大学针灸推拿学院,兰州 730000甘肃省中医院针灸科,兰州 730050

类风湿关节炎热补针法糖酵解缺氧诱导因子1α炎性反应

Rheumatoid arthritisHeat-reinforcing needlingGlycolysisHypoxia-inducible factor-1αInflammatory response

《针刺研究》 2024 (006)

585-593 / 9

国家中医药管理局甘肃郑氏针法学术流派传承工作室项目(No.2305135901);国家自然科学基金项目(No.82060891);甘肃省科技计划项目(No.22JR11RA201);甘肃省针灸推拿临床医学中心开放基金项目(No.ZJKF2203)

10.13702/j.1000-0607.20230487

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