克拉霉素对烟雾暴露哮喘小鼠肺组织中组蛋白去乙酰化酶2及糖皮质激素受体α表达的影响
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摘要
目的探讨克拉霉素对烟雾暴露哮喘小鼠肺组织中组蛋白去乙酰化酶2及糖皮质激素受体α表达的影响。方法将40只SPF级BALB/c雌性小鼠按随机数字表法随机分为对照组(Control group),哮喘组(OVA group),哮喘+烟雾暴露组(SEA group),哮喘+烟雾暴露+克拉霉素干预组(CAM group),每组10只。通过卵蛋白致敏激发建立哮喘模型,哮喘小鼠烟雾暴露建立烟雾暴露哮喘小鼠,给予烟雾暴露小鼠克拉霉素干预。小鼠肺组织切片HE染色,组织病理学观察不同组小鼠肺组织病理变化及炎症评分(UNNDERWOOD标准);Elisa法检测小鼠支气管肺泡灌洗液中白介素(IL-4)及CXCL8炎症介质的表达水平;采用组蛋白去乙酰化酶2(HDAC2)活性检测试剂盒检测不同组小鼠肺组织中组蛋白去乙酰化酶2活性;Western blot法检测小鼠肺组织组蛋白去乙酰化酶2及糖皮质激素受体α蛋白表达。结果肺组织病理切片示哮喘组及烟雾暴露哮喘组小鼠肺组织炎症显著,克拉霉素干预组小鼠肺组织炎症减轻,病理炎症评分克拉霉素干预组炎症评分(1.833±.0373)低于烟雾暴露哮喘组(4.917±0.187)(P=0.031);克拉霉素干预组小鼠肺组织灌洗液中IL-4(60.78411±11.8858)及CXCL8(313.421±96.5448)低于烟雾暴露哮喘组(P=0.042,P=0.027);与烟雾暴露哮喘组(124.444±16.960)相比,克拉霉素干预组小鼠肺组织中HDAC2(266.889±28.205)活性提高(P=0.035),同时HDAC2及GRα表达升高(P=0.041,P=0.035)。结论克拉霉素可降低烟雾暴露哮喘小鼠肺组织炎症反应,改善小鼠肺组织中组蛋白去乙酰化酶2及糖皮质激素受体α的表达。
Objective To investigate the effects of clarithromycin on histone deacetylase 2(HDAC2) and glucocorticoid receptor of the smoke-exposed asthmatic mice.Methods Forty SPF female BALB/c mice were randomly divided into a control group(control group),an asthma group(OVA group),a smoke-exposed asthmatic group(SEA group),and a clarithromycin intervention group(CAM group) according to the random number table.There were 10 mice in each group respectively.The asthmatic mice models were established with ovalbumin.Asthmatic mice were exposed to smoke; clarithromycin were then administrated to the smoke-exposed asthmatic mice.The pathological changes of the lung tissues were presented with hematoxylin-eosin staining,and the inflammation of lung tissues was quantified using the UNDERWOOD standard.The inflammatory mediators including IL-4 and CXCL8 in bronchoalveolar lavage fluid(BALF) were tested using the ELISA method.The activity of HDAC2 was examined with HDAC2 activity test kit.Western blotting was adopted to investigate the expressions of histone deacetylase 2 and glucocorticoid receptor.Results Compared to the control group,the inflammation of lung tissues in the smoke-exposed asthmatic group(4.917±0.187)(SEA group) and clarithromycin intervention group(1.833±0.0373)(CAM group) were significant,while clarithromycin intervention attenuated the inflammatory degree(P=0.031); IL-4(60.78411±11.8858) and CXCL8(313.421±96.5448) in the CAM group were decreased in comparison to SEA group(P=0.042,P=0.027); compared with the SEA group(124.444±16.960),HDAC2 activity was increased in the CAM group(266.889±28.205)(P=0.035); the expressions of HDAC2 and glucocorticoid receptor in the CAM group were higher than those in the SEA group,and the differences were statistically significant(P=0.041,P=0.035).Conclusion Clarithromycin may decrease the airway inflammation of the smoke exposed asthmatic mice and enhance the expressions of HDAC2 and glucocorticoid receptor.
Objective To investigate the effects of clarithromycin on histone deacetylase 2(HDAC2) and glucocorticoid receptor of the smoke-exposed asthmatic mice.Methods Forty SPF female BALB/c mice were randomly divided into a control group(control group),an asthma group(OVA group),a smoke-exposed asthmatic group(SEA group),and a clarithromycin intervention group(CAM group) according to the random number table.There were 10 mice in each group respectively.The asthmatic mice models were established with ovalbumin.Asthmatic mice were exposed to smoke; clarithromycin were then administrated to the smoke-exposed asthmatic mice.The pathological changes of the lung tissues were presented with hematoxylin-eosin staining,and the inflammation of lung tissues was quantified using the UNDERWOOD standard.The inflammatory mediators including IL-4 and CXCL8 in bronchoalveolar lavage fluid(BALF) were tested using the ELISA method.The activity of HDAC2 was examined with HDAC2 activity test kit.Western blotting was adopted to investigate the expressions of histone deacetylase 2 and glucocorticoid receptor.Results Compared to the control group,the inflammation of lung tissues in the smoke-exposed asthmatic group(4.917±0.187)(SEA group) and clarithromycin intervention group(1.833±0.0373)(CAM group) were significant,while clarithromycin intervention attenuated the inflammatory degree(P=0.031); IL-4(60.78411±11.8858) and CXCL8(313.421±96.5448) in the CAM group were decreased in comparison to SEA group(P=0.042,P=0.027); compared with the SEA group(124.444±16.960),HDAC2 activity was increased in the CAM group(266.889±28.205)(P=0.035); the expressions of HDAC2 and glucocorticoid receptor in the CAM group were higher than those in the SEA group,and the differences were statistically significant(P=0.041,P=0.035).Conclusion Clarithromycin may decrease the airway inflammation of the smoke exposed asthmatic mice and enhance the expressions of HDAC2 and glucocorticoid receptor.
引文
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[11]Kobayashi Y,Wada H,Rossios C,et al.A novel macrolide/fl uoroketolide,solithromycin(CEM-101),reverses corticosteroid insensitivity via phosphoinositide3-kinase pathway inhibition[J].Br J Pharmacol,2013,169(5):1024-1034.
[12]Barnes P J.Chronic obstructive pulmonary disease[J].Clin Chest Med,2014,35(1):xiii.
[13]Kobayashi Y,Bossley C,Gupta A,et al.Passive smoking impairs histone deacetylase 2 in children with severe asthma[J].Chest,2014,145(2):305-312.
[14]Reiter J,Demirel N,Mendy A,et al.Macrolides for the longterm management of asthma-a meta-analysis of randomized clinical trials[J].Allergy,2013,68(8):1040-1049.
[15]罗琳,尤列.皮尔曼,周向东,等.克拉霉素抑制气道黏液高分泌细胞内机制的研究[J].中国抗生素杂志,2013,36(8):630-634.
[16]Simpson J L,Powell H,Baines K J,et al.The effect of azithromycin in adults with stable neutrophilic COPD:a double blind randomised,placebo controlled trial[J].PLo S One,2014,9(8):e105609.
[17]Mercado N,Hakim A,Kobayashi Y,et al.Restoration of corticosteroid sensitivity by p38 mitogen activated protein kinase inhibition in peripheral blood mononuclear cells from severe asthma[J].PLo S One,2012,7(7):e41582.
[18]Kobayashi Y,Mercado N,Miller-Larsson A,et al.Increased corticosteroid sensitivity by a long acting beta2 agonist formoterol via beta2 adrenoceptor independent protein phosphatase 2A activation[J].Pulm Pharmacol Ther,2012,25(3):201-207.
[19]Sriskantharajah S,Hamblin N,Worsley S,et al.Targeting phosphoinositide 3-kinaseδfor the treatment of respiratory diseases[J].Ann N Y Acad Sci,2013,1280:35-39.
[2]陈静,周杰,丁铁林,等.用生物化学方法从植物内生真菌提取液中高通量筛选细菌β-内酰胺酶抑制物[J].中国抗生素杂志,2013,38(5):321-326.
[3]Gross K L,Oakley R H,Scoltock A B,et al.Glucocorticoid receptor alpha isoform-selective regulation of antiapoptotic genes in osteosarcoma cells:a new mechanism for glucocorticoid resistance[J].Mol Endocrinol,2011,25(7):1087-1099.
[4]Mercado N,To Y,Kobayashi Y,et al.p38MAPK mitogenactivated protein kinase-gamma inhibition by long-acting beta2 adrenergic agonists reversed steroid insensitivity in severe asthma[J].Mol Pharmacol,2011,80(6):1128-1135.
[5]Kobayashi Y,Mercado N,Barnes P J,et al.Defects of protein phosphatase 2A causes corticosteroid insensitivity in severe asthma[J].PLo S One,2011,6(12):e27627.
[6]Chung K F.p38 mitogen-activated protein kinase pathway in asthma and COPD[J].Chest,2011,139(6):1470-1479.
[7]Milara J,Navarro A,Almudéver P,et al.Oxidative stressinduced glucocorticoid resistance is prevented by dual PDE3/PDE4 inhibition in human alveolar macrophages[J].Clin Exp All,2011,41(4):535-546.
[8]Zhang Y,Leung D Y,Richers B N,et al.Vitamin D inhibits monocyte/macrophage pro-inflammatory cytokine production by targeting MAPK phosphatase-1[J].J Immunol,2012,188(5):2127-2135.
[9]Nuji?K,Banjanac M,Muni?V,et al.Impairment of lysosomal functions by azithromycin and chloroquine contributes to anti-inflammatory phenotype[J].Cell Immunol,2012,279(1):78-86.
[10]Tanabe T,Kanoh S,Tsushima K,et al.Clarithromycin inhibits interleukin-13-induced goblet cell hyperplasia in human airway cells[J].Am J Respir Cell Mol Biol,2011,45(5):1075-1083.
[11]Kobayashi Y,Wada H,Rossios C,et al.A novel macrolide/fl uoroketolide,solithromycin(CEM-101),reverses corticosteroid insensitivity via phosphoinositide3-kinase pathway inhibition[J].Br J Pharmacol,2013,169(5):1024-1034.
[12]Barnes P J.Chronic obstructive pulmonary disease[J].Clin Chest Med,2014,35(1):xiii.
[13]Kobayashi Y,Bossley C,Gupta A,et al.Passive smoking impairs histone deacetylase 2 in children with severe asthma[J].Chest,2014,145(2):305-312.
[14]Reiter J,Demirel N,Mendy A,et al.Macrolides for the longterm management of asthma-a meta-analysis of randomized clinical trials[J].Allergy,2013,68(8):1040-1049.
[15]罗琳,尤列.皮尔曼,周向东,等.克拉霉素抑制气道黏液高分泌细胞内机制的研究[J].中国抗生素杂志,2013,36(8):630-634.
[16]Simpson J L,Powell H,Baines K J,et al.The effect of azithromycin in adults with stable neutrophilic COPD:a double blind randomised,placebo controlled trial[J].PLo S One,2014,9(8):e105609.
[17]Mercado N,Hakim A,Kobayashi Y,et al.Restoration of corticosteroid sensitivity by p38 mitogen activated protein kinase inhibition in peripheral blood mononuclear cells from severe asthma[J].PLo S One,2012,7(7):e41582.
[18]Kobayashi Y,Mercado N,Miller-Larsson A,et al.Increased corticosteroid sensitivity by a long acting beta2 agonist formoterol via beta2 adrenoceptor independent protein phosphatase 2A activation[J].Pulm Pharmacol Ther,2012,25(3):201-207.
[19]Sriskantharajah S,Hamblin N,Worsley S,et al.Targeting phosphoinositide 3-kinaseδfor the treatment of respiratory diseases[J].Ann N Y Acad Sci,2013,1280:35-39.