HPA轴损伤对EAMG大鼠胸腺组织GR mRNA表达影响及补脾强力复方干预作用
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摘要
目的:观察HPA轴损伤对实验性自身免疫性重症肌无力模型(EAMG)大鼠胸腺组织GR mRNA表达影响及补脾强力复方干预作用。方法:将Lewis大鼠随机分为假手术组、模型组、强的松组及补脾强力复方大、中、小剂量组(以下称大、中、小剂量组),除假手术组外,余经双肾上腺摘除致HPA轴损伤,采用免疫诱导注射鼠源性AchR-α亚基97-116肽段复制EAMG模型,造模成功进行灌胃治疗4周后,ELISA法检测各组大鼠血清AchR-Ab、皮质醇水平,RT-q-PCR技术检测各组大鼠胸腺GR mRNA基因表达水平。结果:与假手术组比较,模型组AchR-Ab显著升高,皮质醇水平显著下降,均具有极显著统计学差异(P<0.01);与假手术组比较,模型组胸腺内GRmRNA相对表达量显著降低(P<0.05),与模型组比较,补脾强力复方大剂量组、中剂量组、强的松组胸腺GRmRNA相对表达量升高,具有显著性差异(P<0.05)。结论:HPA轴损伤EAMG大鼠胸腺GR mRNA相对表达量显著降低,而补脾强力复方可提高胸腺组织GR mRNA的表达,有效调节HPA轴功能,这可能是其治疗MG的作用机制之一。
Objective:To observe the effect of HPA-injury on expression of glucocorticoid receptor(GR) mRNA in thymus of experimental autoimmune myasthenia gravis(EAMG) model rats,meanwhile to illuminate the mechanism of Bupi Qiangli Compound in the treatment of MG through increasing GR mRNA expression.Methods:Female Lewis rats were randomly divided into normal group,sham-operation group,model group,prednisone group,and different doses of Bupi Qiangli Compound groups(hereinafter as large,medium and small dose srespectively).Bilateral adrenals of all rats were removed to cause pathological changes and dysfunction of HPA axis except the sham-operation group.R97-116 peptide immune-induced EAMG animal model was established.After the evaluation of successful EAMG models,the EAMG rats were treated once a day for 4 weeks.Then the AchR-Ab,cortisol levels in serum of all rats were detected by ELISA method and the glucocorticoid receptor(GR) mRNA expressions in thymus were determined by RT-q-PCR technique and immunohistochemical method as well.Results:Compared with sham-operation group,the content of AchR-Ab in the serum of model group increased,yet the cortisol level decreased,which both showed extremely significant difference(P<0.01).Compared with sham-operation group,the expression of GR mRNA in model group reduced remarkably,which showed significant difference(P<0.05).Compared with model group,the GR mRNA expression of large,middle dose of Bupi Qiangli Compound groups as well as prednisone group increased to different extents,which all showed significant difference(P<0.05).Conclusion:The expression of mRNA in thymus of HPA-injury EAMG rats decreased significantly and Bupi Qiangli Compound can improve the GR mRNA expression level in thymus,through which to regulate the function of HPA axis,which may be one of the therapeutic mechanisms of Bupi Qiangli Compound in treatment of MG.
Objective:To observe the effect of HPA-injury on expression of glucocorticoid receptor(GR) mRNA in thymus of experimental autoimmune myasthenia gravis(EAMG) model rats,meanwhile to illuminate the mechanism of Bupi Qiangli Compound in the treatment of MG through increasing GR mRNA expression.Methods:Female Lewis rats were randomly divided into normal group,sham-operation group,model group,prednisone group,and different doses of Bupi Qiangli Compound groups(hereinafter as large,medium and small dose srespectively).Bilateral adrenals of all rats were removed to cause pathological changes and dysfunction of HPA axis except the sham-operation group.R97-116 peptide immune-induced EAMG animal model was established.After the evaluation of successful EAMG models,the EAMG rats were treated once a day for 4 weeks.Then the AchR-Ab,cortisol levels in serum of all rats were detected by ELISA method and the glucocorticoid receptor(GR) mRNA expressions in thymus were determined by RT-q-PCR technique and immunohistochemical method as well.Results:Compared with sham-operation group,the content of AchR-Ab in the serum of model group increased,yet the cortisol level decreased,which both showed extremely significant difference(P<0.01).Compared with sham-operation group,the expression of GR mRNA in model group reduced remarkably,which showed significant difference(P<0.05).Compared with model group,the GR mRNA expression of large,middle dose of Bupi Qiangli Compound groups as well as prednisone group increased to different extents,which all showed significant difference(P<0.05).Conclusion:The expression of mRNA in thymus of HPA-injury EAMG rats decreased significantly and Bupi Qiangli Compound can improve the GR mRNA expression level in thymus,through which to regulate the function of HPA axis,which may be one of the therapeutic mechanisms of Bupi Qiangli Compound in treatment of MG.
引文
[1] Renton AE,Pliner HA,Provenzano C.A Genome-wide Association Study of Myasthenia Gravis[J].JAMA Neurol,2015,72(4):396-404.
[2] BalázsC.A New Direction:euro-endocrino-immunology[M]//Neuro-Ophthalmology.Heidelberg:Springer International Publishing,2016:15-17.
[3] Sternberg EM.Neuroendocrine regulation of autoimmune/inflammatory disease[J].Endocrinol,2001,169(3):429-435.
[4] Baggi F,Annoni A,Ubiali F,et al.Breakdown of tolerance to a self-peptide of acetylcholine receptorα-subunit induces experimentalmyasthenia gravis in rats[J].J of immunology,2004,172:2697-2703.
[5] 许文华,韩莹莹,汪思应,等.鼠源乙酰胆碱受体α亚基97-116肽段免疫Lewis鼠诱导实验性重症肌无力模型[J].中国神经精神疾病杂志,2006,32(2):179-180.
[6] Gupta BBP,Lalchhandama K.Molecular mechanisms of glucocorticoid action[J].Curt Sci,2002,83(9):1103-1111.
[7] 孙华,郑金瓯.影响重症肌无力治疗想的相关因素评价[J].中国临床康复,2004,8(34):7801-7802.
[8] 隋黎明,徐仁宝.大鼠胸腺细胞糖皮质激素受体结合与糖皮质激素效应的定量关系[J].生理学报,1991,43(3):243-248.
[9] Rossini G P,Malaguti C.Nanomolar concentrations of untransformed glucocorticoid receptor in nuclei of intact cells[J].The Journal of steroid biochemistry and molecular biology,1994,51(5):291-298.
[10] 黄莹,赵重波,朱雯华,等.重症肌无力患者外周血白细胞糖皮质激素受体α、βmRNA表达与激素疗效的关系[J].复旦学报:医学版,2012,39(1):31-35.
[11] 姜雄,何前松,况时祥.补脾强力复方对自身免疫性重症肌无力大鼠胸腺组织GRmRNA表达的影响[J].辽宁中医杂志,2017,44(2):398-402.
[12] Shimizu H,Ichikawa Y,Yoshida M,et al.Lymphocyte Subsets of the Peripheral Blood in Myasthenia Gravis Determined by Two-Color Flowcytometry[J].Autoimmunity,1990,6(3):173-182.
[13] 罗东林,周继红,刘宝华,等.GR和HsP70在大鼠严重多发伤后肝组织中的变化及其意义[J].第三军医大学学报,2003,25(3):255-257.
[14] Bamberger C M,Bamberger A M,de Castro M,et al.Glucocorticoid receptor beta,a potential endogenous inhibitor of glucocorticoid action in humans[J].Journal of Clinical Investigation,1995,95(6):2435.
[15] 严灿,徐志伟,史亚飞,等.加减四逆散对慢性心理应激大鼠胸腺糖皮质激素受体作用的研究[J].中国中西医结合杂志,2002,22(9):691-693.
[16] 张星虎.重症肌无力患者外周血单个核细胞糖皮质激素受体的研究[D].北京:中国协和医科大学,2000.
[17] 于新娟,侯桂华.中药对糖皮质激素受体影响的研究进展[J].国外医学:中医中药分册,2004,26(4):216-219,223.
[18] 包照日格图,吴敦序,陈淑俊,等.补肾健脾三方对哮喘大鼠糖皮质激素受体及皮质酮的影响[J].中国中西医结合杂志,1999,19(8):487-489.
[19] 詹秀琴.补肾方药对老年大鼠胸腺细胞糖皮质激素受体的作用研究[J].南京中医药大学学报:自然科学版,2000,16(3):152-153.
[20]况时祥,张树森,赵芝兰,等.补脾强力胶囊治疗重症肌无力Ⅰ、Ⅱa型临床研究[J].新中医,2012,44(7):53-54.
[21]马宗超,唐智宏,张海,等.谈附子的药理及临床应用[J].时珍国医国药,2004,15(11):790-792.
[22]马子密,傅延龄.历代本草药性会解[M].北京:中国医药科技出版社,2002:811,482,183.
[23]常波,杨剑.序贯法和单纯进补法对慢性运动应激大鼠脑组织和胸腺组织糖皮质激素受体的影响[J].体育科学,2006,26(2):48-54.
[24]许建华,范忠泽,吴敦序,等.补肾定喘汤对哮喘大鼠肺组织糖皮质激素受体及血浆皮质酮、ACTH的影响[J].中国中医基础医学杂志,2003,9(1):27-29.
[2] BalázsC.A New Direction:euro-endocrino-immunology[M]//Neuro-Ophthalmology.Heidelberg:Springer International Publishing,2016:15-17.
[3] Sternberg EM.Neuroendocrine regulation of autoimmune/inflammatory disease[J].Endocrinol,2001,169(3):429-435.
[4] Baggi F,Annoni A,Ubiali F,et al.Breakdown of tolerance to a self-peptide of acetylcholine receptorα-subunit induces experimentalmyasthenia gravis in rats[J].J of immunology,2004,172:2697-2703.
[5] 许文华,韩莹莹,汪思应,等.鼠源乙酰胆碱受体α亚基97-116肽段免疫Lewis鼠诱导实验性重症肌无力模型[J].中国神经精神疾病杂志,2006,32(2):179-180.
[6] Gupta BBP,Lalchhandama K.Molecular mechanisms of glucocorticoid action[J].Curt Sci,2002,83(9):1103-1111.
[7] 孙华,郑金瓯.影响重症肌无力治疗想的相关因素评价[J].中国临床康复,2004,8(34):7801-7802.
[8] 隋黎明,徐仁宝.大鼠胸腺细胞糖皮质激素受体结合与糖皮质激素效应的定量关系[J].生理学报,1991,43(3):243-248.
[9] Rossini G P,Malaguti C.Nanomolar concentrations of untransformed glucocorticoid receptor in nuclei of intact cells[J].The Journal of steroid biochemistry and molecular biology,1994,51(5):291-298.
[10] 黄莹,赵重波,朱雯华,等.重症肌无力患者外周血白细胞糖皮质激素受体α、βmRNA表达与激素疗效的关系[J].复旦学报:医学版,2012,39(1):31-35.
[11] 姜雄,何前松,况时祥.补脾强力复方对自身免疫性重症肌无力大鼠胸腺组织GRmRNA表达的影响[J].辽宁中医杂志,2017,44(2):398-402.
[12] Shimizu H,Ichikawa Y,Yoshida M,et al.Lymphocyte Subsets of the Peripheral Blood in Myasthenia Gravis Determined by Two-Color Flowcytometry[J].Autoimmunity,1990,6(3):173-182.
[13] 罗东林,周继红,刘宝华,等.GR和HsP70在大鼠严重多发伤后肝组织中的变化及其意义[J].第三军医大学学报,2003,25(3):255-257.
[14] Bamberger C M,Bamberger A M,de Castro M,et al.Glucocorticoid receptor beta,a potential endogenous inhibitor of glucocorticoid action in humans[J].Journal of Clinical Investigation,1995,95(6):2435.
[15] 严灿,徐志伟,史亚飞,等.加减四逆散对慢性心理应激大鼠胸腺糖皮质激素受体作用的研究[J].中国中西医结合杂志,2002,22(9):691-693.
[16] 张星虎.重症肌无力患者外周血单个核细胞糖皮质激素受体的研究[D].北京:中国协和医科大学,2000.
[17] 于新娟,侯桂华.中药对糖皮质激素受体影响的研究进展[J].国外医学:中医中药分册,2004,26(4):216-219,223.
[18] 包照日格图,吴敦序,陈淑俊,等.补肾健脾三方对哮喘大鼠糖皮质激素受体及皮质酮的影响[J].中国中西医结合杂志,1999,19(8):487-489.
[19] 詹秀琴.补肾方药对老年大鼠胸腺细胞糖皮质激素受体的作用研究[J].南京中医药大学学报:自然科学版,2000,16(3):152-153.
[20]况时祥,张树森,赵芝兰,等.补脾强力胶囊治疗重症肌无力Ⅰ、Ⅱa型临床研究[J].新中医,2012,44(7):53-54.
[21]马宗超,唐智宏,张海,等.谈附子的药理及临床应用[J].时珍国医国药,2004,15(11):790-792.
[22]马子密,傅延龄.历代本草药性会解[M].北京:中国医药科技出版社,2002:811,482,183.
[23]常波,杨剑.序贯法和单纯进补法对慢性运动应激大鼠脑组织和胸腺组织糖皮质激素受体的影响[J].体育科学,2006,26(2):48-54.
[24]许建华,范忠泽,吴敦序,等.补肾定喘汤对哮喘大鼠肺组织糖皮质激素受体及血浆皮质酮、ACTH的影响[J].中国中医基础医学杂志,2003,9(1):27-29.
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