文献类型：期刊文献

中文题名：热补针法对类风湿性关节炎寒证家兔滑膜组织中长链非编码RNA表达谱的影响

英文题名：Effect of heat-reinforcing needling on expression profile of LncRNAs in synovial tissues of rabbits with cold-syndrome type rheumatoid arthritis

作者：杜小正[1];刘莉梅[2];刘翠[1];井维尧[1];袁博[2];张星华[3];张枫帆[3];李向军[3];刘强[1];王海东[3]

第一作者：杜小正

机构：[1]甘肃中医药大学针灸推拿学院,兰州730000;[2]甘肃中医药大学附属医院,兰州730000;[3]甘肃省中医院,兰州730050

第一机构：甘肃中医药大学针灸推拿学院

年份：2025

卷号：50

期号：1

起止页码：65

中文期刊名：针刺研究

外文期刊名：Acupuncture Research

收录：;北大核心:【北大核心2023】；

基金：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);甘肃省自然科学基金项目(No.22JR11RA198、22JR11RA201);甘肃省青年科技基金计划项目(No.23JRRA1686)。

语种：中文

中文关键词：类风湿性关节炎;热补针法;长链非编码RNA

外文关键词：Rheumatoid arthritis;Heat-reinforcing needling;Long noncoding RNA

摘要：目的:观察热补针法对类风湿性关节炎(RA)寒证模型家兔滑膜组织中长链非编码RNA(LncRNA)表达谱的影响,探讨热补针法治疗RA的作用机制。方法:将18只家兔随机分为正常组、模型组、热补针法组,每组6只。采用卵蛋白联合弗氏完全佐剂混合溶液注射,配合低温冷冻法构建RA寒证模型。热补针法组予以热补针法针刺双侧“足三里”,留针30 min/次,1次/d,共14 d。干预结束后观察家兔一般情况,测定膝关节周径、痛阈值;以膝关节彩色多普勒超声观察关节腔积液、滑膜厚度及内部血流信号;以HE染色法观察滑膜病理形态学变化;以ELISA法检测各组家兔血清肿瘤坏死因子α(TNF-α)、白细胞介素(IL)1β及IL-6含量;以高通量测序技术检测并分析家兔滑膜组织中差异LncRNA及生物学功能;以实时荧光定量PCR法验证5个显著差异LncRNA的表达水平。结果:与正常组比较,模型组家兔膝关节周径增加(P<0.05),痛阈值降低(P<0.05),滑膜弥漫性增生、关节腔积液及异常血流信号明显,滑膜衬里增生、变厚,衬里下可见毛细血管生成,大量炎性细胞簇集,血清中TNF-α、IL-1β、IL-6含量升高(P<0.05)。与模型组比较,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),滑膜弥漫性增生、关节腔积液及异常血流信号均有不同程度减轻,滑膜衬里增生明显减少,可见少量炎性细胞簇集,血清TNF-α、IL-1β、IL-6含量降低(P<0.05)。高通量测序和生物信息分析结果显示,热补针法组和模型组共有142个LncRNAs有相反的表达趋势,其中有59个LncRNA被逆向上调,主要表现为代谢途径、过氧化物酶体增殖物激活受体(PPAR)信号通路和AMP依赖的蛋白激酶(AMPK)信号通路为核心的功能聚类;有83个LncRNA被逆向下调,主要表现为细胞周期、丝裂原活化蛋白激酶(MAPK)信号通路和细胞衰老为核心的功能聚类。PCR验证结果显示,与正常组比较,模型组家兔膝关节滑膜组织中MSTRG.15448.1和MSTRG.17571.1表达水平降低(P<0.05),MSTRG.5134.1、MSTRG.13294.1和MSTRG.15972.1表达水平升高(P<0.05);与模型组比较,热补针法组家兔膝关节滑膜组织中MSTRG.15448.1和MSTRG.17571.1表达水平升高(P<0.05),MSTRG.5134.1、MSTRG.13294.1和MSTRG.15972.1表达水平降低(P<0.05)。结论:热补针法能够调节RA滑膜组织中LncRNA表达谱的变化,这可能是其抑制滑膜炎性反应、改善症状的机制之一。
Objective To observe the effect of heat-reinforcing needling on the expression profile of long non-coding RNA(LncRNA)in the synovial tissue of rabbits with cold-syndrome type rheumatoid arthritis(RA),so as to explore its mechanisms underlying improvement of RA.Methods A total of 18 New Zeland rabbits were randomly divided into normal,model and heat-reinforcing needling(acupuncture)groups,with 6 rabbits in each group.The RA model was established by injection of mixed solution(emulsifier)of ovalbumin powder(80 g)+NaCl(20 mL)+Freund’s complete adjuvant(20 mL),combined with cold compress(ice cube+crystal calcium chloride)of the knee-joint.In addition,rabbits of the normal and model groups received regular grasping and fixation in the same way as those of the acupuncture group.For rabbits of the acupuncture group,the heat-reinforcing needling stimulation was applied to bilateral"Zusanli"(ST36)for about 1 min,followed by retaining the needle for 30 min,once a day,for a total of 7 d.The rabbits’general condition was observed,and the knee joint circumference and pain threshold after intervention were measured.The knee-joint cavity effusion,synovial thickness,and internal blood flow signals were observed using a color Doppler ultrasound apparatus.Histopathological changes of synovium were observed using H.E.staining.The contents of serum tumor necrosis factorα(TNF-α),interleukin(IL)1βand IL-6 were detected using enzyme linked immunosorbent assay(ELISA),and the differential LncRNAs and biological functions in the synovial tissues were detected and analyzed using high-throughput sequencing technology.The expression levels of 5 significantly differentially expressed LncRNAs were verified using qPCR.Results Compared with the normal group,the model group of rabbits showed a significant increase in the knee joint circumference(P<0.05)and the content of serum TNF-α,IL-1β,IL-6(P<0.05),and a significant decrease in the pain threshold(P<0.05),and diffuse synovial hyperplasia of the synovial membrane,joint effusion and abnormal blood flow signals,proliferated and thickened synovial lining,with visible capillary formation beneath the lining and a large number of inflammatory cell clustering.In comparison with the model group,the knee-joint circumference,and the serum contents of TNF-α,IL-1βand IL-6 were obviously decreased(P<0.05),while the pain threshold was notably increased(P<0.05),and H.E.staining showed a reduction in the hyperplasia of synovial lining,with a milder inflammatory cell infiltration.High-throughput sequencing and bioinformatics analysis displayed that a total of 142 LncRNAs were found to have opposite expression trends between the acupuncture group and the model group,of which 83 LncRNAs were up-regulated in the model group vs the normal group and downregulated in the acupuncture group vs the model group,mainly involving the core functional clusters of cell cycle,mitogen-activated protein kinase(MAPK)signaling and cell senescence,and 59 LncRNAs were down-regulated in the model group vs the normal group,and up-regulated in the acupuncture group vs the model group,mainly involving the metabolic pathway,peroxisome proliferator-activated receptor(PPAR)signaling and AMP-dependent protein kinase(AMPK)signaling.The results of qPCR showed that compared with the normal group,the expression levels of MSTRG.15448.1 and MSTRG.17571.1 in the synovial tissue of knee joint were significantly decreased(P<0.05),and those of MSTRG.5134.1,MSTRG.13294.1 and MSTRG.15972.1 were significantly increased in the model group(P<0.05).Compared with the model group,the expression levels of MSTRG.15448.1 and MSTRG.17571.1 were significantly increased(P<0.05),and those of MSTRG.5134.1,MSTRG.13294.1 and MSTRG.15972.1 were significantly decreased in the acupuncture group(P<0.05).Conclusion The heat-reinforcing needling can regulate the expression profile of LncRNAs in the RA synovial tissue,which may be one of the mechanisms underlying inhibition of synovial inflammatory response and improvement of symptoms.