牛磺鹅去氧胆酸对糖皮质激素受体激活效应
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摘要
探讨牛磺鹅去氧胆酸(TCDCA)对糖皮质激素受体(GR)的激活效应,采用免疫荧光术检测NR8383细胞内GR的表达及不同浓度TCDCA对NR8383细胞核移位的影响,采用实时荧光定量PCR法分析了TCDCA对AA大鼠FLS细胞中由GR产生的VCAM-1和COX-2mRNA表达的影响,采用酶联免疫吸附法(ELISA)分析了TCDCA对AA大鼠FLS细胞中由GR介导的VCAM-1蛋白表达的影响。结果表明,TCDCA可以极显著性(P<0.01)的激活NR8383细胞内的GR受体,且在加入GR的特异性抑制剂RU486之后,GR受体的荧光值显著下降,表明RU486阻断了TCDCA对GR受体的激活作用。而TCDCA组和DEX组间无显著性差异(P>0.05);TCDCA可以抑制VCAM-1及COX-2的表达量。由此进一步证明TCDCA是通过GR受体发挥抗炎和免疫调节作用。
The activating effect of taurochenodeoxycholic acid(TCDCA)on glucocorticoid receptors(GRs)has been explored.The expression of GRs and the nuclear translocation in NR8383 cells effected by different concentration of TCDCA were detected by immunofluorescence technique.Using real-time PCR evaluated the gene expression levels of VCAM-1 and COX-2 induced by GRs in FLS cells from AA rats.The protein expression of VCAM-1 was also measured by ELISA.The results demonstrated that TCDCA could dramatically improve the activation of GRs in NR8383 cells(P<0.01).The fluorescence value was significantly decreased after treated with RU486,a specific inhibitor of GRs,which indicated that the TCDCA-induced activation of GRs could be blocked by RU486.Moreover,there was no significant difference between TCDCA group and DEX group(P>0.05).In addition,the expression of VCAM-1 and COX-2 can be reduced by TCDCA treatment.It was further proved that the anti-inflammatory and immuuoregulatory effects of TCDCA were performed via GRs.
The activating effect of taurochenodeoxycholic acid(TCDCA)on glucocorticoid receptors(GRs)has been explored.The expression of GRs and the nuclear translocation in NR8383 cells effected by different concentration of TCDCA were detected by immunofluorescence technique.Using real-time PCR evaluated the gene expression levels of VCAM-1 and COX-2 induced by GRs in FLS cells from AA rats.The protein expression of VCAM-1 was also measured by ELISA.The results demonstrated that TCDCA could dramatically improve the activation of GRs in NR8383 cells(P<0.01).The fluorescence value was significantly decreased after treated with RU486,a specific inhibitor of GRs,which indicated that the TCDCA-induced activation of GRs could be blocked by RU486.Moreover,there was no significant difference between TCDCA group and DEX group(P>0.05).In addition,the expression of VCAM-1 and COX-2 can be reduced by TCDCA treatment.It was further proved that the anti-inflammatory and immuuoregulatory effects of TCDCA were performed via GRs.
引文
[1]李培锋,关红,哈斯苏荣,等.鸡胆汁有效成分CD-CA和TCDCA的抗炎作用研究[J].中兽医医药杂志,2002,21(112):7-10.
[2]胡霞敏,石朝周.牛磺结合及其游离胆汁酸在小鼠镇咳祛痰抗炎模型上的作用比较[J].中国临床药学杂志,2001,10(2):85-88.
[3]何秀玲,李培锋,关红,等.牛磺酸鹅去氧胆酸对小鼠免疫功能的影响[J].中药材,2005,28(12):1089-1092.
[4]李培锋,刘明强,关红.牛磺鹅去氧胆酸对小鼠脾脏中白介素-2基因表达的影响[J].中国畜牧兽医,2009,36(3):66-71.
[5]李欣,牛春宇,毛伟,等.TCDCA对大鼠脂肪代谢及其相关受体FXR表达量的影响[J].中国兽医学报,2018,38(7):1430-1434.
[6]王旭,张子英,李培锋.牛磺鹅去氧胆酸对TGR5介导的NR8383细胞TNF-ɑ的影响[J].中国兽医学报,2016,36(1):108-111.
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[9]SOLA S.Nuclear translocation of UDCA by the glucocorticoid receptor is required to reduce TGF-β1-induced apoptosis in rat hepatocytes[J].Hepatology,2005,42(4):925-934.
[10]SHARMA R,LONG A,GILMER J F.Advances in bile acid medicinal chemistry[J].Curr Med Chem,2011,18(26):4029-4052.
[11]TANAK A,MAKIN O.Ligand-independent activation of the glucocorticoid receptor by ursodeoxycholic acid:repression of IFN-γ-induced MHC class II gene expression viaa glucocorticoid receptor-dependent pathway[J].J Immunol,1996,156(4):1601-1608.
[12]SUSANA S,CASTRO R E,KREN B T,et al.Modulation of nuclear steroid receptors byursodeoxycholic acid inhibits TGF-beta1-induced E2F-1/p53-mediated apoptosis of rat hepatocytes[J].Biochemistry,2004,43(26):8429-38.
[13]WEITZELCHRISTOF A,STARKDAGMAR B,KULLMANNFRANK C,et al.Ursodeoxycholic acid induced activation of the glucocorticoid receptor in primary rat hepatocytes[J].Eur J Gastroen Hepat,2005,17(2):169-177.
[14]MIURA T,OUCHIDA R,YOSHIKAWA N,et al.Functionalmodulatio of the glucocorticoid receptor and suppression of NF-kappa B-dependent transcription by ursodeoxycholic acid[J].J Biol Chem,2001,276(50):47371-47378.
[15]MATSUYAMA T,KITANI A.The role of VCAM-1molecule in the pathogenesis ofrheumatoid synovitis[J].Hum Cell,1996,9(3):187-192.
[16]INOUE H,UMESONO K,NISHIMORI T,et al.Glucocorticoid-mediated suppression of the promoter activity of the cyclooxygenase-2gene is modulated by expression of its receptor in vascular endothelial cells[J].Biochem Bioph Res Commun,1999,254(2):292-298.
[17]任公平,那辉,佟雷,等.辣椒素通过下调COX-2和mPGES-1抑制IL-1β诱导的NCI-H460细胞PGE2含量的实验研究[J].西安交通大学学报(医学版),2016,37(2):283-287.
[18]毛伟.TCDCA对AA模型大鼠GR介导的基因组学转录激活效应的影响[D].内蒙古呼和浩特:内蒙古农业大学,2011.
[19]FERLAZZP V,MILANO S.Anti-inflammatory effects of annexin-1:stimulation of IL-10release and inhibition of nitric oxide synthesis[J].Int Immunopharmacol,2003,3(10/11):1363-1369.
[20]何国雄.痹肿消汤对胶原诱导性关节炎大鼠滑膜组织annexin 1、aldolase A表达的影响[D].湖南长沙:中南大学,2006:24-27.
[21]刘明强.TCDCA对佐剂性关节炎模型大鼠的治疗作用及其作用机制研究[D].内蒙古呼和浩特:内蒙古农业大学,2012.
[22]张婷婷.TCDCA对AA模型大鼠成纤维样滑膜细胞EP2/EP4-G蛋白-cAMP信号通路的影响[D].内蒙古呼和浩特:内蒙古农业大学,2015.
[23]BONDESON J,FOXWELL B,BRENNAN F,et al.Defining therapeutic targets by using adenovirus:blocking NF-kappa B inhibits both inflammatory and destructive mechanisms in rheumatoid synovium but spares anti-inflammatory mediators[J].Proc Natl Acad Sci USA,1999,96(10):5668-5673.
[24]MAKAROV S S,JOHNSTON W N,OLSEN J C,et al.NF-kappa B as a target for anti-inflammatory gene therapy:suppression of inflammatory responses in monocytic and stromal cells by stable gene transfer of I kappa B alpha cDNA[J].Gene Ther,1997,4(8):846-852.
[25]LI P,SANZ I.NF-kappa B regulates VCAM-1expression on f ibroblast-1ike synoviocytes[J].J Immunol,2000,164(11);5990-5997.
[26]BARCHOWSKY A,FRLETA D,VINCENTI M P.Integration of the NF-kappa B and mitogen-activated protein kinase/AP-1 pathways at the collagenase-1promoter:divergence of IL-1and TNF-dependent signal transduction in rabbit primary synovial fibroblast[J].Cytokine,2000,12(10):1469-1479.
[27]GUSTIN J A,OZES O N,AKCA H,et al.Cell typespecific expression of the I kappa B kinases determines the significance of phosphatidylinositol 3-kinase/Akt signaling to NF-kappa B activation[J].J Biol Chem,2004,279(3):1615-1620.
[2]胡霞敏,石朝周.牛磺结合及其游离胆汁酸在小鼠镇咳祛痰抗炎模型上的作用比较[J].中国临床药学杂志,2001,10(2):85-88.
[3]何秀玲,李培锋,关红,等.牛磺酸鹅去氧胆酸对小鼠免疫功能的影响[J].中药材,2005,28(12):1089-1092.
[4]李培锋,刘明强,关红.牛磺鹅去氧胆酸对小鼠脾脏中白介素-2基因表达的影响[J].中国畜牧兽医,2009,36(3):66-71.
[5]李欣,牛春宇,毛伟,等.TCDCA对大鼠脂肪代谢及其相关受体FXR表达量的影响[J].中国兽医学报,2018,38(7):1430-1434.
[6]王旭,张子英,李培锋.牛磺鹅去氧胆酸对TGR5介导的NR8383细胞TNF-ɑ的影响[J].中国兽医学报,2016,36(1):108-111.
[7]李磊.TCDCA与糖皮质激素受体的相互作用及其对活化蛋白-1的影响[D].内蒙古呼和浩特:内蒙古农业大学,2014.
[8]TANAKA H,MAKIN O I.Ursodeoxycholic acid-dependent activation of the glucocorticoid receptor[J].Biochen Bioph Res Commun,1992,188(2):942-948.
[9]SOLA S.Nuclear translocation of UDCA by the glucocorticoid receptor is required to reduce TGF-β1-induced apoptosis in rat hepatocytes[J].Hepatology,2005,42(4):925-934.
[10]SHARMA R,LONG A,GILMER J F.Advances in bile acid medicinal chemistry[J].Curr Med Chem,2011,18(26):4029-4052.
[11]TANAK A,MAKIN O.Ligand-independent activation of the glucocorticoid receptor by ursodeoxycholic acid:repression of IFN-γ-induced MHC class II gene expression viaa glucocorticoid receptor-dependent pathway[J].J Immunol,1996,156(4):1601-1608.
[12]SUSANA S,CASTRO R E,KREN B T,et al.Modulation of nuclear steroid receptors byursodeoxycholic acid inhibits TGF-beta1-induced E2F-1/p53-mediated apoptosis of rat hepatocytes[J].Biochemistry,2004,43(26):8429-38.
[13]WEITZELCHRISTOF A,STARKDAGMAR B,KULLMANNFRANK C,et al.Ursodeoxycholic acid induced activation of the glucocorticoid receptor in primary rat hepatocytes[J].Eur J Gastroen Hepat,2005,17(2):169-177.
[14]MIURA T,OUCHIDA R,YOSHIKAWA N,et al.Functionalmodulatio of the glucocorticoid receptor and suppression of NF-kappa B-dependent transcription by ursodeoxycholic acid[J].J Biol Chem,2001,276(50):47371-47378.
[15]MATSUYAMA T,KITANI A.The role of VCAM-1molecule in the pathogenesis ofrheumatoid synovitis[J].Hum Cell,1996,9(3):187-192.
[16]INOUE H,UMESONO K,NISHIMORI T,et al.Glucocorticoid-mediated suppression of the promoter activity of the cyclooxygenase-2gene is modulated by expression of its receptor in vascular endothelial cells[J].Biochem Bioph Res Commun,1999,254(2):292-298.
[17]任公平,那辉,佟雷,等.辣椒素通过下调COX-2和mPGES-1抑制IL-1β诱导的NCI-H460细胞PGE2含量的实验研究[J].西安交通大学学报(医学版),2016,37(2):283-287.
[18]毛伟.TCDCA对AA模型大鼠GR介导的基因组学转录激活效应的影响[D].内蒙古呼和浩特:内蒙古农业大学,2011.
[19]FERLAZZP V,MILANO S.Anti-inflammatory effects of annexin-1:stimulation of IL-10release and inhibition of nitric oxide synthesis[J].Int Immunopharmacol,2003,3(10/11):1363-1369.
[20]何国雄.痹肿消汤对胶原诱导性关节炎大鼠滑膜组织annexin 1、aldolase A表达的影响[D].湖南长沙:中南大学,2006:24-27.
[21]刘明强.TCDCA对佐剂性关节炎模型大鼠的治疗作用及其作用机制研究[D].内蒙古呼和浩特:内蒙古农业大学,2012.
[22]张婷婷.TCDCA对AA模型大鼠成纤维样滑膜细胞EP2/EP4-G蛋白-cAMP信号通路的影响[D].内蒙古呼和浩特:内蒙古农业大学,2015.
[23]BONDESON J,FOXWELL B,BRENNAN F,et al.Defining therapeutic targets by using adenovirus:blocking NF-kappa B inhibits both inflammatory and destructive mechanisms in rheumatoid synovium but spares anti-inflammatory mediators[J].Proc Natl Acad Sci USA,1999,96(10):5668-5673.
[24]MAKAROV S S,JOHNSTON W N,OLSEN J C,et al.NF-kappa B as a target for anti-inflammatory gene therapy:suppression of inflammatory responses in monocytic and stromal cells by stable gene transfer of I kappa B alpha cDNA[J].Gene Ther,1997,4(8):846-852.
[25]LI P,SANZ I.NF-kappa B regulates VCAM-1expression on f ibroblast-1ike synoviocytes[J].J Immunol,2000,164(11);5990-5997.
[26]BARCHOWSKY A,FRLETA D,VINCENTI M P.Integration of the NF-kappa B and mitogen-activated protein kinase/AP-1 pathways at the collagenase-1promoter:divergence of IL-1and TNF-dependent signal transduction in rabbit primary synovial fibroblast[J].Cytokine,2000,12(10):1469-1479.
[27]GUSTIN J A,OZES O N,AKCA H,et al.Cell typespecific expression of the I kappa B kinases determines the significance of phosphatidylinositol 3-kinase/Akt signaling to NF-kappa B activation[J].J Biol Chem,2004,279(3):1615-1620.