时辰艾灸干预类风湿性关节炎模型大鼠核心钟基因Clock、Bmal1表达与细胞焦亡的关系  

Effects of Chrono-moxibustion on core clock genes Clock,Bmal1 and pyroptosis in rats with rheumatoid arthritis

作　　者：余明芳 陈白露 何欣玲 王嫒洋 吴晓 Yu Mingfang;Chen Bailu;He Xinling;Wang Aiyang;Wu Xiao(College of Integrated Traditional Chinese and Western Medicine,Traditional Chinese Medicine Hospital Affiliated to Southwest Medical University,Luzhou 640000,Sichuan Province,China;Department of Acupuncture,Luzhou T.C.M.Hospital,Luzhou 646000,Sichuan Province,China)

机构地区：[1]西南医科大学中西医结合学院·附属中医医院,四川省泸州市640000 [2]泸州市中医医院针灸科,四川省泸州市646000

出　　处：《中国组织工程研究》2022年第35期5700-5706,共7页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金项目(81804208);项目负责人:吴晓,项目名称:不同时辰艾灸治疗RA的抗炎节律及核心钟基因的调控机制研究。

摘　　要：背景:类风湿关节炎是症状特点具有昼夜节律性的炎症性疾病,艾灸治疗类风湿关节炎疗效显著,但是其具体机制是否与钟基因调控及细胞焦亡的抑制有关尚不清楚。目的:探究不同时辰艾灸治疗类风湿关节炎模型大鼠对核心钟基因和细胞焦亡的影响。方法:随机将40只成年SD大鼠分为空白组、类风湿关节炎模型组(模型组)、辰时艾灸组(辰时组)、酉时艾灸组(酉时组),雌雄各半,每组10只。空白组大鼠于右后足垫皮内注射无菌生理盐水,其余以弗氏完全佐剂制备实验性类风湿关节炎大鼠模型,辰时组(7:00-9:00)和酉时组(17:00-19:00)于造模后第7天在相应时辰取足三里、肾俞进行艾灸治疗,1次/d,每周治疗6 d后休息1 d,共治疗18 d。空白组及模型组行同法固定,不予艾灸干预。采用排水法分别测量大鼠模型制备前、制备后及每周艾灸干预后右足足容积;治疗结束后采集大鼠下丘脑和右膝关节滑膜,应用实时荧光定量PCR和Western blot观察下丘脑或膝关节滑膜组织中Clock、Bmal1基因、NOD样受体热蛋白结构域相关蛋白3(Nod-like receptor protein 3,NLRP3)、半胱氨酸蛋白酶1(Caspase-1)、消皮素D的表达情况。结果与结论:(1)各组大鼠下丘脑中Clock、Bmal1 mRNA表达水平差异无显著性意义(P>0.05);在Clock、Bmal1 mRNA相对表达量中,下丘脑各组大鼠的表达丰度均比关节滑膜高(P<0.01);关节滑膜中,与空白组比较,模型组Clock、Bmal1 m RNA和蛋白表达均显著降低(P<0.01);与模型组相比,艾灸可显著升高滑膜Clock、Bmal1 mRNA和蛋白表达(P<0.05,P<0.01);与辰时组比较,酉时组中Clock、Bmal1 mRNA和蛋白表达稍上调,但差异无显著性意义(P>0.05);(2)与空白组比较,模型组大鼠关节滑膜中NLRP3、Caspase-1、消皮素D mRNA表达量明显升高(P<0.01);与模型组比较,酉时艾灸可明显降低类风湿关节炎关节滑膜中NLRP3、Caspase-1、消皮素D mRNA表达量(P<0.01);与辰�BACKGROUND:Rheumatoid arthritis is an inflammatory disease with a circadian variation of symptoms.Moxibustion has achieved remarkable efficacy in the treatment of rheumatoid arthritis,but it is unclear whether the specific mechanism is related to the regulation of the clock gene and the inhibition of pyro ptosls.OBJECTIVE;To explo re the effects of moxibustion at different time points on core clock genes Clock,Bmal1 and pyroptosis in rats with rheumatoid arthritis.METHODS:Forty adult Sprague-Dawley rats of both sexes were randomized into a control group,a model group,a moxibustion at 7:00-9:00 A.M.group and a moxibustion at 5:00-7:00 P.M.group,with 10 rats in each group.To establish the rheumatoid arthritis model,the rats were injected with Freund s Complete Adjuvant at the right foot.Rats in the control group were given the normal saline at the right foot.At the 7;after modeling,the two moxibustion groups began to be treated with moxibustion at Zusanli and Shenshu acupoints at 7:00-9:00 A.M.or 5:00-7:00 P.M.,once a day,for total 18 treatments with a 1-day break every 6 days.We measured the right foot volume of rats using drainage method before model establishment,after model establishment and after treatment.The hypothalamus and the synovium of the right knee joint were collected from the rats after treatment.The expressions of Clock,Bma11,Nodlike receptor family pyrin domain-containing 3(NLRP3),Caspase-1,and Gasdermin D(GSDMD)in the hypothalamus or knee synovium were tested by real-time fluorescence qua ntitative PCR and western blot.RESULTS AND CONCLUSION:The mRNA expreession of Clock and BMAL1 in the hypothalamus had no obvious change among the four groups(P>0.05).The relative mRNA expression of Clock and Bmal in the hypothalamus was higher than that in the synovium(P<0.01).The expression of Clock and Bmoll in the synovium was significantly down-regulated in the model group compared with the control group(P<0.01).Moxibustion significantly increased the mRNA and protein expression of Clock and Bmali in the syn

关 键 词：类风湿关节炎 艾灸 CLOCK基因 Bmal1基因 细胞焦亡 大鼠 

分 类 号：R459.9[医药卫生—治疗学] R318[医药卫生—临床医学]