穴位埋线对原发性痛经大鼠子宫组织核苷酸结合寡聚化结构域受体炎性小体的影响
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摘要
目的:观察穴位埋线对原发性痛经(PD)大鼠子宫组织核苷酸结合寡聚化结构域受体3(NLRP3炎性小体及其下游因子白细胞介素1β(IL-1β)、白细胞介素18(IL-18)表达的影响,探讨穴位埋线治疗PD的可能机制。方法:雌性SD大鼠随机分为正常组、模型组、埋线组、西药组,每组10只。苯甲酸雌二醇联合缩宫素皮下注射建立PD大鼠模型。埋线组于造模第1天及造模第5天对"关元""三阴交"穴进行埋线治疗;西药组采用芬必得灌胃治疗,每日1次,共10d。于第11天比较各组大鼠30min内扭体次数;HE染色法观察大鼠子宫病理形态学的变化,并进行病理损伤评分;Western blot法检测子宫组织NLRP3、半胱氨酰天冬氨酸特异性蛋白酶1(caspase-1)、IL-1β、IL-18的表达。结果:与正常组比较,模型组扭体次数增多(P<0.01);与模型组比较,埋线组、西药组30min内扭体次数均减少(P<0.01,P<0.05);与西药组比较,埋线组扭体次数明显减少(P<0.01)。模型组大鼠子宫内膜大范围剥脱并有较严重的水肿;埋线组及西药组子宫内膜损伤程度较模型组轻,子宫内膜剥脱少见且水肿程度较轻;与正常组比较,模型组病理评分明显上升(P<0.01);与模型组比较,埋线组和西药组病理评分显著降低(P<0.01)。与正常组比较,模型组大鼠子宫组织NLRP3、caspase-1、IL-1β、IL-18蛋白表达明显升高(P<0.01);与模型组比较,埋线组大鼠子宫组织NLRP3、caspase-1、IL-1β及IL-18蛋白表达显著降低(P<0.01,P<0.05),西药组大鼠子宫组织caspase-1、IL-1β、IL-18蛋白表达显著降低(P<0.01);与西药组比较,埋线组子宫组织NLRP3蛋白表达明显降低(P<0.01)。结论:穴位埋线可明显改善PD大鼠疼痛症状和病理损伤情况,其作用机制可能与抑制子宫组织中NLRP3炎性小体的活化有关。
Objective To observe the effect of acupoint catgut embedding on the expression of nucleotide-binding oligomerization domain(NOD)-like receptor protein 3(NLRP3)inflammasome,interleukin(IL)-1βand IL-18 in the uterine tissue of primary dysmenorrhea(PD)rats,so as to explore its underlying mechanisms in improving PD.Methods Forty female SD rats were randomly divided into control,model,acupoint catgut embedding and medication groups(n=10 in each group).The PD model was established by subcutaneous injection of Estradiol Benzoate(0.5 mg/rat on the 1 st and 10 thday,and 0.2 mg/rat from 2 nd to 9 th day)and Oxytocin(2 U/rat,i.p.).The catgut embedding was applied to bilateral"Sanyinjiao"(SP6)and"Guanyuan"(CV4)on the 1 st and 5 th day after modeling.Rats of the medication group were treated by intragastric perfusion of Fenbid(0.8 mL/rat,125 mg/100 mL)once daily for 10 days.The body writhing times in 30 min were recorded.The histopathological changes of the uterine were observed by H.E.staining.Western blot was used to detect the expression of NLRP 3,caspase-1,IL-1βand IL-18 in uterine tissues.Results The body writhing times were notably more in the model group than in the control group(P<0.01),and obviously fewer in both medication and catgut embedding groups than in the model group(P<0.05,P<0.01).After modeling,the rats' endometrium was extensively exfoliated and got swelling,the histopathological score and the expression levels of NLRP3,caspase-1,IL-1βand IL-18 proteins in the uterus tissue were evidently increased in the model group relevant to the control group(P<0.01).Following the treatment,the degree of endometrial exfoliation and edema of the uterus tissue was lightened,the pathological score was significantly reduced(P<0.01),and the expression levels of caspase-1,IL-1βand IL-18 protein in uterus tissue were markedly decreased in both acupoint catgut embedding and medication groups(P<0.01,P<0.05).The NLRP3 protein expression was significantly decreased in the acupoint catgut embedding group compared with that in the model group(P<0.01).The therapeutic effect of acupoint catgut embedding was significantly superior to that of medication in reducing writhing times and down-regulating expression of NLPR3 protein(P<0.01).No significant differences were found between catgut embedding and medication in histopathological score,and expression levels of caspase-1,IL-1β and IL-18 proteins(P>0.05).Conclusion The acupoint catgut embedding can significantly alleviate the symptoms and pathological damage in PD rats,which may be related to its effect in inhibiting the activation of NLRP3 inflammasome in the uterine tissue.
Objective To observe the effect of acupoint catgut embedding on the expression of nucleotide-binding oligomerization domain(NOD)-like receptor protein 3(NLRP3)inflammasome,interleukin(IL)-1βand IL-18 in the uterine tissue of primary dysmenorrhea(PD)rats,so as to explore its underlying mechanisms in improving PD.Methods Forty female SD rats were randomly divided into control,model,acupoint catgut embedding and medication groups(n=10 in each group).The PD model was established by subcutaneous injection of Estradiol Benzoate(0.5 mg/rat on the 1 st and 10 thday,and 0.2 mg/rat from 2 nd to 9 th day)and Oxytocin(2 U/rat,i.p.).The catgut embedding was applied to bilateral"Sanyinjiao"(SP6)and"Guanyuan"(CV4)on the 1 st and 5 th day after modeling.Rats of the medication group were treated by intragastric perfusion of Fenbid(0.8 mL/rat,125 mg/100 mL)once daily for 10 days.The body writhing times in 30 min were recorded.The histopathological changes of the uterine were observed by H.E.staining.Western blot was used to detect the expression of NLRP 3,caspase-1,IL-1βand IL-18 in uterine tissues.Results The body writhing times were notably more in the model group than in the control group(P<0.01),and obviously fewer in both medication and catgut embedding groups than in the model group(P<0.05,P<0.01).After modeling,the rats' endometrium was extensively exfoliated and got swelling,the histopathological score and the expression levels of NLRP3,caspase-1,IL-1βand IL-18 proteins in the uterus tissue were evidently increased in the model group relevant to the control group(P<0.01).Following the treatment,the degree of endometrial exfoliation and edema of the uterus tissue was lightened,the pathological score was significantly reduced(P<0.01),and the expression levels of caspase-1,IL-1βand IL-18 protein in uterus tissue were markedly decreased in both acupoint catgut embedding and medication groups(P<0.01,P<0.05).The NLRP3 protein expression was significantly decreased in the acupoint catgut embedding group compared with that in the model group(P<0.01).The therapeutic effect of acupoint catgut embedding was significantly superior to that of medication in reducing writhing times and down-regulating expression of NLPR3 protein(P<0.01).No significant differences were found between catgut embedding and medication in histopathological score,and expression levels of caspase-1,IL-1β and IL-18 proteins(P>0.05).Conclusion The acupoint catgut embedding can significantly alleviate the symptoms and pathological damage in PD rats,which may be related to its effect in inhibiting the activation of NLRP3 inflammasome in the uterine tissue.
引文
[1]IACOVIDES S,AVIDON I,BAKER F C.What we know about primary dysmenorrhea today:a critical review[J].Hum Reprod Update,2015,21(6):762-778.
[2]CHEN C X,KWEKKEBOOM K L,WARD S E.Self-report pain and symptom measures for primary dysmenorrhoea:a critical review[J].Eur J Pain,2015,19(3):377-391.
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[4]郑仁省,刘金里.芍药甘草汤对痛经患者血清IL-1β、TNF-α及疼痛的影响[J].长春中医药大学学报,2017,33(4):614-616.
[5]范郁山,苗芙蕊,廖爱妮,等.隔药灸“命门”穴对痛经模型大鼠血清雌激素、孕激素水平及其受体表达的影响[J].针刺研究,2013,38(5):352-357.
[6]谢丹,居明乔,曹鹏,等.炎症因子和原发性痛经相关性的研究进展[J].中国临床药理学与治疗学,2014,19(3):346-350.
[7]徐崇权.电针对原发性痛经模型大鼠镇痛效应及免疫机制的研究[D].广州:广州中医药大学,2009.
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[9]JO E K,JIN K K,SHIN D M,et al.Molecular mechanisms regulating NLRP3inflammasome activation[J].Cell Mol Immunol,2016,13(2):148-159.
[10]LU A,WU H.Structural mechanisms of inflammasome assembly[J].FEBS J,2015,282(3):435-444.
[11]陈自喜.1.NLRP3炎性小体与分娩启动的研究.2.临产前后硫化氢生成酶的表达与收缩相关蛋白相关性研究[D].上海:第二军医大学,2017.
[12]HEIDARIFAR R,MEHRAN N,HEIDARI A,et al.Effect of Dill(Anethum graveolens)on the severity of primary dysmenorrhea in compared with mefenamic acid:a randomized,double-blind trial[J].J Res Med Sci,2014,19(4):326-330.
[13]PETRAGLIA F,PARKE S,SERRANI M,et al.Estradiol valerate plus dienogest versus ethinylestradiol plus levonorgestrel for the treatment of primary dysmenorrhea[J].Int J Gynaecol Obstet,2014,125(3):270-274.
[14]陈盼碧,陈静,崔瑾,等.穴位埋线法对原发性痛经大鼠神经-内分泌-免疫网络的调控与影响[J].针刺研究,2018,43(1):29-33.
[15]于蓝,邰浩清.穴位埋线治疗原发性痛经实验研究[J].河南中医,2015,35(1):56-57.
[16]许卫国.穴位埋线治疗原发性痛经98例[J].江西中医药,2014,45(12):57-58.
[17]孙萍萍,蒙珊.分期穴位埋线治疗原发性痛经的临床疗效观察[J].针灸临床杂志,2015,31(9):46-49.
[18]毕颖,邵晓梅,宣丽华.分期穴位埋线治疗原发性痛经:随机对照研究[J].中国针灸,2014,34(2):115-119.
[19]郭爱松,李爱红,陈鑫,等.穴位埋线对急性脑梗死患者运动功能及血清高敏C-反应蛋白、白介素-6的影响[J].针刺研究,2013,38(3):224-228.
[20]苗艳换,赵吉平,云洁,等.痛经患者三阴交穴压痛反应研究[J].针刺研究,2014,39(5):401-405.
[21]王霞灵,曹大农,单志群.艾附暖宫丸治疗痛经的实验研究[J].湖北中医学院学报,2003,5(2):18-19.
[22]陈盼碧,杨孝芳,王兴桂,等.艾灸对原发性痛经大鼠血液流变学及子宫组织PGF2α含量的影响[J].中国疼痛医学杂志,2015,21(11):826-829.
[2]CHEN C X,KWEKKEBOOM K L,WARD S E.Self-report pain and symptom measures for primary dysmenorrhoea:a critical review[J].Eur J Pain,2015,19(3):377-391.
[3]YANG L,CAO Z Y,YU B Y,et al.An in vivo mouse model of primary dysmenorrhea[J].Exp Anim,2015,64(3):295-303.
[4]郑仁省,刘金里.芍药甘草汤对痛经患者血清IL-1β、TNF-α及疼痛的影响[J].长春中医药大学学报,2017,33(4):614-616.
[5]范郁山,苗芙蕊,廖爱妮,等.隔药灸“命门”穴对痛经模型大鼠血清雌激素、孕激素水平及其受体表达的影响[J].针刺研究,2013,38(5):352-357.
[6]谢丹,居明乔,曹鹏,等.炎症因子和原发性痛经相关性的研究进展[J].中国临床药理学与治疗学,2014,19(3):346-350.
[7]徐崇权.电针对原发性痛经模型大鼠镇痛效应及免疫机制的研究[D].广州:广州中医药大学,2009.
[8]RATHINAM V A K,FITZGERALD K A.Inflammasome complexes:emerging mechanisms and effector functions[J].Cell,2016,165(4):792-800.
[9]JO E K,JIN K K,SHIN D M,et al.Molecular mechanisms regulating NLRP3inflammasome activation[J].Cell Mol Immunol,2016,13(2):148-159.
[10]LU A,WU H.Structural mechanisms of inflammasome assembly[J].FEBS J,2015,282(3):435-444.
[11]陈自喜.1.NLRP3炎性小体与分娩启动的研究.2.临产前后硫化氢生成酶的表达与收缩相关蛋白相关性研究[D].上海:第二军医大学,2017.
[12]HEIDARIFAR R,MEHRAN N,HEIDARI A,et al.Effect of Dill(Anethum graveolens)on the severity of primary dysmenorrhea in compared with mefenamic acid:a randomized,double-blind trial[J].J Res Med Sci,2014,19(4):326-330.
[13]PETRAGLIA F,PARKE S,SERRANI M,et al.Estradiol valerate plus dienogest versus ethinylestradiol plus levonorgestrel for the treatment of primary dysmenorrhea[J].Int J Gynaecol Obstet,2014,125(3):270-274.
[14]陈盼碧,陈静,崔瑾,等.穴位埋线法对原发性痛经大鼠神经-内分泌-免疫网络的调控与影响[J].针刺研究,2018,43(1):29-33.
[15]于蓝,邰浩清.穴位埋线治疗原发性痛经实验研究[J].河南中医,2015,35(1):56-57.
[16]许卫国.穴位埋线治疗原发性痛经98例[J].江西中医药,2014,45(12):57-58.
[17]孙萍萍,蒙珊.分期穴位埋线治疗原发性痛经的临床疗效观察[J].针灸临床杂志,2015,31(9):46-49.
[18]毕颖,邵晓梅,宣丽华.分期穴位埋线治疗原发性痛经:随机对照研究[J].中国针灸,2014,34(2):115-119.
[19]郭爱松,李爱红,陈鑫,等.穴位埋线对急性脑梗死患者运动功能及血清高敏C-反应蛋白、白介素-6的影响[J].针刺研究,2013,38(3):224-228.
[20]苗艳换,赵吉平,云洁,等.痛经患者三阴交穴压痛反应研究[J].针刺研究,2014,39(5):401-405.
[21]王霞灵,曹大农,单志群.艾附暖宫丸治疗痛经的实验研究[J].湖北中医学院学报,2003,5(2):18-19.
[22]陈盼碧,杨孝芳,王兴桂,等.艾灸对原发性痛经大鼠血液流变学及子宫组织PGF2α含量的影响[J].中国疼痛医学杂志,2015,21(11):826-829.