基于肾虚血瘀理论探讨艾灸对实验性类风湿性关节炎家兔抗炎镇痛的作用机制
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摘要
目的:观察艾灸"肾俞""足三里"对实验性类风湿性关节炎(Rheumatoid arthritis,RA)家兔滑膜组织中TGF-β1、NF-κBp65、VEGF含量和TGF-β1/NF-κB通路的影响;基于肾虚血瘀理论探讨艾灸治疗RA的抗炎镇痛作用机制。方法:将30只日本大耳白兔(简称"家兔"),随机分为空白组、对照组、治疗组,每组10只。对照组、治疗组用0.5 mL/kg弗氏完全佐剂(FCA)法造模,空白组不作任何处理。在造模后第7天,用艾炷灸治疗组家兔双侧"肾俞""足三里"各5壮,1次/d,两侧交替进行,共治疗3个疗程18 d,两疗程之间休息1 d,空白组和对照组不进行艾灸治疗。治疗后第21天,取膝关节滑膜组织和滑膜液。采用免疫组化染色法进行滑膜组织染色并测定组织中TGF-β1、NF-κBp65、VEGF蛋白基因表达水平。结果:与空白组比较,对照组家兔滑膜组织中TGF-β1、NF-κBp65、VEGF蛋白含量均明显升高,具有显著的统计学意义(P<0.01)。治疗后,与对照组相比,治疗组RA家兔滑膜组织TGF-β1、NF-κBp65、VEGF蛋白含量均降低,具有显著的统计学意义(P<0.01)。结论:艾灸"肾俞""足三里"能降低RA家兔滑膜组织TGF-β1、NF-κBp65的蛋白含量,下调VEGF的表达,从而抑制血管新生和血管翳形成,缓解滑膜炎症,提示艾灸可能是通过调肾活血治法发挥抗炎镇痛之效。
Objective: To observe the influence of moxibustion at "Shenshu(BL23)","Zusanli(ST36)" on TGF-β1,NF-κBp65,VEGF and NF-κB/TGF-β1 pathway in the synovial tissue of rheumatoid arthritis(RA) rabbits based on the theory of kidney deficiency and blood stasis and discuss the mechanism of anti-inflammation and analgesic effect of moxibustion on RA. Methods: A total of 30 rabbits(Japanese Big-ear White species) were randomly divided into blank control group(hereinafter referred to as the "blank"), RA model group(hereinafter referred to as the "model"), moxibustion treatment group(hereinafter referred to as the "moxibustion") with 10 cases in each. RA model and moxibustion groups were established by injection of Freund's Complete Adjuvant(FCA, 0.5 mL/kg) into the animals' bilateral joint cavities. On the 7th day after modeling, moxibustion was applied to bilateral "Shenshu(BL23)" and "Zusanli(ST36)", 5 cones every time, once a day(except Sundays), 18 days altogether. The "blank" and "model" did not make any treatment. After treatment for 21 days,synovial was obtained and dyed immunohistochemically to measure the TGF-β1,NF-κBp65 and VEGF protein gene expression levels in the synovial tissue. Results: Compared with "blank",the TGF-β1,NF-κBp65 and VEGF protein levels were obviously enhanced in RA model group(P<0.01). After treatment,while compared with "model", those in "moxibustion" decreased considerably(P<0.01). Conclusion: Moxibustion at "Shenshu(BL23)" and "Zusanli(ST36)" can reduce the TGF-β1 and NF-κBp65 protein levels, regulate the VEGF expression and stop the vascular and pannus formation, and thus improving synovial inflammation. It is suggested that moxibustion may play an anti-inflammatory and analgesic effect by regulating the kidney and activating blood.
Objective: To observe the influence of moxibustion at "Shenshu(BL23)","Zusanli(ST36)" on TGF-β1,NF-κBp65,VEGF and NF-κB/TGF-β1 pathway in the synovial tissue of rheumatoid arthritis(RA) rabbits based on the theory of kidney deficiency and blood stasis and discuss the mechanism of anti-inflammation and analgesic effect of moxibustion on RA. Methods: A total of 30 rabbits(Japanese Big-ear White species) were randomly divided into blank control group(hereinafter referred to as the "blank"), RA model group(hereinafter referred to as the "model"), moxibustion treatment group(hereinafter referred to as the "moxibustion") with 10 cases in each. RA model and moxibustion groups were established by injection of Freund's Complete Adjuvant(FCA, 0.5 mL/kg) into the animals' bilateral joint cavities. On the 7th day after modeling, moxibustion was applied to bilateral "Shenshu(BL23)" and "Zusanli(ST36)", 5 cones every time, once a day(except Sundays), 18 days altogether. The "blank" and "model" did not make any treatment. After treatment for 21 days,synovial was obtained and dyed immunohistochemically to measure the TGF-β1,NF-κBp65 and VEGF protein gene expression levels in the synovial tissue. Results: Compared with "blank",the TGF-β1,NF-κBp65 and VEGF protein levels were obviously enhanced in RA model group(P<0.01). After treatment,while compared with "model", those in "moxibustion" decreased considerably(P<0.01). Conclusion: Moxibustion at "Shenshu(BL23)" and "Zusanli(ST36)" can reduce the TGF-β1 and NF-κBp65 protein levels, regulate the VEGF expression and stop the vascular and pannus formation, and thus improving synovial inflammation. It is suggested that moxibustion may play an anti-inflammatory and analgesic effect by regulating the kidney and activating blood.
引文
[1] Kormendy D,Hoff H,Hoff P, et al. Impact of the CTLA-4/cd28 axis on the processes of joint inflammation in rheumatoid arthritis[J]. Arthritis Rheum, 2013, 65(1):81-87.
[2] Hoy D, March L, Brooks P, et al. The global burden of hip and knee osteoarthritis: estimates from the global burden of disease 2010 study[J]. Annals of the Rheumatic Diseases, 2014, 73(7):1323-1330.
[3] Gao J, Liu XG, Huang DJ, et al. Involvement of the hypothalamus-pituitary-adrenal axis in moxibustion-induced changes of NF-kappaB signaling in the synovial tissue in rheumatic arthritic rats[J]. Acupuncture research, 2010, 35(3):198.
[4] 张阔, 徐媛, 丁沙沙,等. 基于文献研究针刺治疗类风湿关节炎选穴规律[J]. 中国针灸, 2017, 37(2): 221-224.
[5] 高梓珊, 甘君学, 郭江燕, 等. 麦粒灸调控IL-17对实验性RA大鼠血清TNF-a、IL-1b含量影响的研究[J]. 针灸临床杂志, 2017, 33(1): 37-41.
[6] 殷媛, 陈天帷, 张瑞, 等. 艾灸对胶原诱导性关节炎大鼠血清白介素-17和肿瘤坏死因子α含量的影响[J]. 针刺研究, 2017, 42(2): 149-152.
[7] 章呜.羟甲基几丁糖的兔佐剂性关节炎滑膜细胞体外培养影响的试验研究[J].浙江临床医学,2002,4(6):405-406.
[8] 余曙光,徐斌.实验针灸学[M].北京:人民卫生出版社,2012:269-272.
[9] 李蓉, 宁乔怡, 姚血明,等. 从肾虚血瘀理论探讨类风湿关节炎的发病机制[J]. 中华中医药学刊, 2017,35(5):1206-1208.
[10] 陈红淑,杨元宵,周小杰,等.肾气丸通过TGF-β1/Smads/ILK信号途径干预肾小管上皮细胞间充质转分化的分子机制[J].中华中医药杂志,2016,31(6):2102-2105.
[11] 温红辉,霍燕微,刘捷裕,等.NF-κB和TGF-β1在阿霉素肾病模型中的检测与病理意义[J].现代医院,2013,13(5):28-30.
[12] 王金越.转化生长因子β信号与肌肉调控[J].中国组织工程研究,2016,20(33):4979.
[13] 王连雷.重组人TGF-β1和NF-κB在腰椎黄韧带细胞中的作用研究[D],济南:山东大学,2016:30-31.
[14] Wang HZ, Wang HH, Huang SS, et al. Inhibitory effect of baicalin on collagerr induced arthritis in rats through the nuclear factor-Κb pathway[J].J Pharmacol Exp Ther,2014,350(2):435-443.
[15] 陈白露,彭麒,熊燕,等.艾灸对实验性RA家兔滑膜HIF-1α/VEGF的调节作用[J].时珍国医国药,2016,27(12):3042-3044.
[16] Yao J, Ling HU, Song XG, et al. Influence of moxibustion at “Shenshu” ( BL 23) and “Zusanli” ( ST 36) on Ras-MAPK signal pathways in synovial tissues of rats with experimental rheumatoid arthritis[J]. World Journal of Acupuncture-Moxibustion, 2013, 23(2):29-33.
[17] 郑保主,胡玲,宋小鸽,等. 艾灸对类风湿关节炎大鼠下丘脑POMCmRNA 和 PDYNmRNA 表达的影响[J]. 中国针灸,2013,33(5):433-437.
[18] 吴菲,杨涵棋,张耀,等. 隔姜灸对 RA 家兔不同组织 HSP70表达的影响[J]. 中华中医药学刊,2015,33(4):801-803.
[2] Hoy D, March L, Brooks P, et al. The global burden of hip and knee osteoarthritis: estimates from the global burden of disease 2010 study[J]. Annals of the Rheumatic Diseases, 2014, 73(7):1323-1330.
[3] Gao J, Liu XG, Huang DJ, et al. Involvement of the hypothalamus-pituitary-adrenal axis in moxibustion-induced changes of NF-kappaB signaling in the synovial tissue in rheumatic arthritic rats[J]. Acupuncture research, 2010, 35(3):198.
[4] 张阔, 徐媛, 丁沙沙,等. 基于文献研究针刺治疗类风湿关节炎选穴规律[J]. 中国针灸, 2017, 37(2): 221-224.
[5] 高梓珊, 甘君学, 郭江燕, 等. 麦粒灸调控IL-17对实验性RA大鼠血清TNF-a、IL-1b含量影响的研究[J]. 针灸临床杂志, 2017, 33(1): 37-41.
[6] 殷媛, 陈天帷, 张瑞, 等. 艾灸对胶原诱导性关节炎大鼠血清白介素-17和肿瘤坏死因子α含量的影响[J]. 针刺研究, 2017, 42(2): 149-152.
[7] 章呜.羟甲基几丁糖的兔佐剂性关节炎滑膜细胞体外培养影响的试验研究[J].浙江临床医学,2002,4(6):405-406.
[8] 余曙光,徐斌.实验针灸学[M].北京:人民卫生出版社,2012:269-272.
[9] 李蓉, 宁乔怡, 姚血明,等. 从肾虚血瘀理论探讨类风湿关节炎的发病机制[J]. 中华中医药学刊, 2017,35(5):1206-1208.
[10] 陈红淑,杨元宵,周小杰,等.肾气丸通过TGF-β1/Smads/ILK信号途径干预肾小管上皮细胞间充质转分化的分子机制[J].中华中医药杂志,2016,31(6):2102-2105.
[11] 温红辉,霍燕微,刘捷裕,等.NF-κB和TGF-β1在阿霉素肾病模型中的检测与病理意义[J].现代医院,2013,13(5):28-30.
[12] 王金越.转化生长因子β信号与肌肉调控[J].中国组织工程研究,2016,20(33):4979.
[13] 王连雷.重组人TGF-β1和NF-κB在腰椎黄韧带细胞中的作用研究[D],济南:山东大学,2016:30-31.
[14] Wang HZ, Wang HH, Huang SS, et al. Inhibitory effect of baicalin on collagerr induced arthritis in rats through the nuclear factor-Κb pathway[J].J Pharmacol Exp Ther,2014,350(2):435-443.
[15] 陈白露,彭麒,熊燕,等.艾灸对实验性RA家兔滑膜HIF-1α/VEGF的调节作用[J].时珍国医国药,2016,27(12):3042-3044.
[16] Yao J, Ling HU, Song XG, et al. Influence of moxibustion at “Shenshu” ( BL 23) and “Zusanli” ( ST 36) on Ras-MAPK signal pathways in synovial tissues of rats with experimental rheumatoid arthritis[J]. World Journal of Acupuncture-Moxibustion, 2013, 23(2):29-33.
[17] 郑保主,胡玲,宋小鸽,等. 艾灸对类风湿关节炎大鼠下丘脑POMCmRNA 和 PDYNmRNA 表达的影响[J]. 中国针灸,2013,33(5):433-437.
[18] 吴菲,杨涵棋,张耀,等. 隔姜灸对 RA 家兔不同组织 HSP70表达的影响[J]. 中华中医药学刊,2015,33(4):801-803.