Toll样受体mRNA和相关细胞因子在肝多房棘球蚴病患者体内的变化
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摘要
目的探讨Toll样受体2(Toll-like receptor 2,TLR2)和TLR4 m RNA在肝多房棘球蚴病(HAE)患者外周血单核细胞(PBMC)和肝组织中的表达情况及其与血浆相关细胞因子水平之间的相关性。方法收集2012年1月至2015年6月在新疆医科大学第一附属医院住院治疗的肝多房棘球蚴病患者(HAE组)28例,以体检健康的志愿者(HC组,n=28例)作为对照,用ELISA检测血浆中γ干扰素(IFN-γ)、白细胞介素5(IL-5)、IL-23、IL-10水平,用q RT-PCR法检测PBMCs以及HAE患者肝组织中TLR2和TLR4 m RNA的表达水平,用血细胞分析仪检测外周血嗜酸粒细胞百分比(Eo%)。Spearman秩相关分析PBMCs中TLR2和TLR4 m RNA水平与血浆中细胞因子水平和外周血Eo%的相关性。结果ELISA检测结果显示,HAE组血浆中IFN-γ、IL-5、IL-23、IL-10水平分别为(301.100±47.290)、(43.420±11.380)、(86.580±31.990)、(8.766±7.568)pg/ml,均高于HC组的(301.100±67.790)、(40.970±6.310)、(46.770±15.490)、(6.272±10.360)pg/ml,其中两组的IL-23水平差异有统计学意义(P<0.01)。q RT-PCR检测结果显示,HAE组PBMCs中TLR2 m RNA相对表达量为0.100±0.084,高于HC组的0.055±0.040(P<0.05);HAE组和HC组的TLR4 m RNA相对表达量分别为0.004±0.003和0.003±0.002,两者间差异无统计学意义(P>0.05)。HAE患者肝病灶组织中TLR2和TLR4 m RNA相对表达量分别为29.680±25.650和21.340±16.640,均高于病灶旁的2.308±4.140和5.541±9.233以及正常组织的1.112±1.431和1.100±1.734(P<0.01)。HAE组和HC组外周血Eo%分别为0.448±0.240和0.110±0.100,两者差异有统计学意义(P<0.01)。Spearman秩相关分析结果显示,PBMCs中TLR2 m RNA水平与血浆中IL-23水平和外周血Eo%之间均存在正相关关系(r=0.368,r=0.382)。结论肝多房棘球蚴病患者PBMCs、肝病灶中的TLR2和TLR4 m RNA表达水平升高,PBMCs中TLR2 m RNA水平与血浆中IL-23水平和外周血Eo%之间均存在正相关关系。
Objective To investigate the expression of Toll-like receptor 2(TLR2) and TLR4 m RNA in peripheral blood mononuclear cells(PBMC) and in the liver of patients with hepatic alveolar echinococcosis(HAE),and their correlations with related cytokines in plasma. Methods Twenty-eight HAE patients hospitalized in the First Affiliated Hospital of Xinjiang Medical University during January 2012 and June 2015 and 28 healthy volun-teers as a control were enrolled in this study. Plasma levels of interferon-γ(IFN-γ), interleukin-5(IL-5), IL-23, and IL-10 were measured by ELISA. q RT-PCR was performed to detect TLR2 and TLR4 m RNA levels in PBMCs andhepatic tissues. The percentage of peripheral blood eosinophil(Eo%) was determined by a hematology analyzer. The correlations of TLR2 and TLR4 m RNA levels in PBMCs with levels of related cytokines and Eo% were analyzed with the Spearman Correlation method. Results ELISA results showed that the plasma levels of IFN-γ, IL-5, IL-23,and IL-10 in the HAE group were(301.100 ±47.290),(43.420 ±11.380),(86.580 ±31.990) and(8.766 ±7.568) pg/ml respectively, which were higher than those in the control [(301.100±67.790),(40.970±6.310),(46.770±15.490) and(6.272±10.360) pg/ml] with a statistical significance for IL-23(P <0.01). Results of q RT-PCR showed that the expression level of TLR2 in the HAE group(0.100 ±0.084) was significantly higher than that in the control(0.055 ±0.040)(P<0.05), while the expression level of TLR4 in the HAE group(0.004 ±0.003) was comparable to that in the control(0.003 ±0.002)(P >0.05). The expression of TLR2 and TLR4 m RNA in HAE lesions in the HAE group(29.680 ±25.650 and 21.340 ±16.640, respectively) were both significantly higher than that in para-lesion regions(2.308 ±4.140 and 5.541 ±9.233) and that in tissues of the control(1.112 ±1.431 and 1.100 ±1.734)(P <0.01).There was also a significant difference in Eo% between the HAE(0.448 ±0.240) and the control(0.110 ±0.100)groups. Spearman correlation coefficients revealed a positive correlation of TLR2 m RNA in PBMCs with plasma IL-23 level and peripheral blood Eo% in HAE subjects(r=0.368, r=0.382, respectively). Conclusion There are increases in TLR2 and TLR4 m NRA expression in PBMCs and in HAE lesions in HAE patients. The TLR2 m NRA expression in PBMCs positively correlates with plasma IL-23 level and peripheral Eo%.
Objective To investigate the expression of Toll-like receptor 2(TLR2) and TLR4 m RNA in peripheral blood mononuclear cells(PBMC) and in the liver of patients with hepatic alveolar echinococcosis(HAE),and their correlations with related cytokines in plasma. Methods Twenty-eight HAE patients hospitalized in the First Affiliated Hospital of Xinjiang Medical University during January 2012 and June 2015 and 28 healthy volun-teers as a control were enrolled in this study. Plasma levels of interferon-γ(IFN-γ), interleukin-5(IL-5), IL-23, and IL-10 were measured by ELISA. q RT-PCR was performed to detect TLR2 and TLR4 m RNA levels in PBMCs andhepatic tissues. The percentage of peripheral blood eosinophil(Eo%) was determined by a hematology analyzer. The correlations of TLR2 and TLR4 m RNA levels in PBMCs with levels of related cytokines and Eo% were analyzed with the Spearman Correlation method. Results ELISA results showed that the plasma levels of IFN-γ, IL-5, IL-23,and IL-10 in the HAE group were(301.100 ±47.290),(43.420 ±11.380),(86.580 ±31.990) and(8.766 ±7.568) pg/ml respectively, which were higher than those in the control [(301.100±67.790),(40.970±6.310),(46.770±15.490) and(6.272±10.360) pg/ml] with a statistical significance for IL-23(P <0.01). Results of q RT-PCR showed that the expression level of TLR2 in the HAE group(0.100 ±0.084) was significantly higher than that in the control(0.055 ±0.040)(P<0.05), while the expression level of TLR4 in the HAE group(0.004 ±0.003) was comparable to that in the control(0.003 ±0.002)(P >0.05). The expression of TLR2 and TLR4 m RNA in HAE lesions in the HAE group(29.680 ±25.650 and 21.340 ±16.640, respectively) were both significantly higher than that in para-lesion regions(2.308 ±4.140 and 5.541 ±9.233) and that in tissues of the control(1.112 ±1.431 and 1.100 ±1.734)(P <0.01).There was also a significant difference in Eo% between the HAE(0.448 ±0.240) and the control(0.110 ±0.100)groups. Spearman correlation coefficients revealed a positive correlation of TLR2 m RNA in PBMCs with plasma IL-23 level and peripheral blood Eo% in HAE subjects(r=0.368, r=0.382, respectively). Conclusion There are increases in TLR2 and TLR4 m NRA expression in PBMCs and in HAE lesions in HAE patients. The TLR2 m NRA expression in PBMCs positively correlates with plasma IL-23 level and peripheral Eo%.
引文
[1]温浩,吐尔干艾力·阿吉,邵英梅,等.棘球蚴病防治成就及面临的挑战[J].中国寄生虫学与寄生虫病杂志,2015,33(6):466-471.
[2]Bresson-Hadni SMJ,Mantion GA,Vuitton DA.Echinococcosis of the liver[M]//Rodes JBJ,Blei AT,Reichen M.Textbook of Hepatology.3rd edition.Oxford,UK:Blackwell Publishing,2007:1047-1057.
[3]单骄宇,吐尔洪江·吐逊,王俊华,等.慢性囊型包虫病患者外周血单个核细胞中TLR2、4、7 m RNA表达水平的研究[J].中国微生物学与免疫学杂志,2011,31(3):273-278.
[4]Trinchieri G,Sher A.Cooperation of Toll-like receptor signals in innate immune defenc e[J].Nat Rev Immunol,2007,7(3):179-90.
[5]黄蓉,吴青,李茂军,等.Toll样受体4在免疫相关细胞中的作用[J].中华临床医师杂志,2016,10(8):1178-1182.
[6]Ghosh D,Stumhofer JS.Do you see what I see:recognition of protozoan parasites by Toll-like receptors[J].Curr Immunol Rev,2013,9(3):129-140.
[7]WHO Informal Working Group.International classification of ultrasound images in cystic echinococcosis for application in clinical and field epidemiological settings[J].Acta Trop,2003,85(2):253-261.
[8]中国医师协会外科医师分会包虫病外科专业委员会.两型包虫病诊断和治疗的专家共识(2015版)[J].中华消化外科杂志,2015,14(4):253-264.
[9]黄玉娴,孔庆瑜,蔡高荣,等.三种检测嗜酸性粒细胞方法的比较[J].分子诊断与治疗杂志,2004,16(3):9-11.
[10]Tuxun T,Apaer S,Ma HZ,et al.The potential role of Th9cell related cytokine and transcription factors in patients with hepatic alveolar echinococcosis[J].J Immunol Res,2015,2015:895416.
[11]Vuitton DA.The ambiguous role of immunity in echinococcosis:protection of the host or of the parasite[J].Acta Trop,2003,85(2):119-132.
[12]Hübner MP,Manfras BJ,Margos MC,et al.Echinococcus multilocularis metacestodes modulate cellular cytokine and chemokine release by peripheral blood mononuclear cells in alveolar echinococcosis patients[J].Clin Exp Immunol,2006,145(2):243-51.
[13]Vuitton DA.Echinococcosis and allergy[J].Clin Rev Allergy Immunol,2004,26(2):93-104.
[14]Janeway CA Jr.Approaching the asymptote?Evolution and revolution in immunology[J].Cold Spring Harb Symp Quant Biol,1989,54 Pt 1:1-13.
[15]Kane CM,Jung E,Pearce EJ.Schistosoma mansoni egg antigenmediated modulation of Toll-like receptor(TLR)-induced activation occurs independently of TLR2,TLR4,and My D88[J].Infect Immun,2008,76(12):5754-5759.
[16]Smith P,Mangan NE,Fallon PG.Generation of parasite antigens for use in toll-like receptor research[J].Methods Mol Biol,2009,517:401-413.
[17]Kawai T,Akira S.The role of pattern-recognition receptors in innate immunity:update on toll-like receptors[J].Nat Immunol,2010,11(5):373-384.
[18]van Riet E,Everts B,Retra K,et al.Combined TLR2 and TLR4 ligation in the context of bacterial or helminth extracts in human monocyte derived dendritic cells:molecular correlates for Th1/Th2 polarization[J].BMC Immunol,2009,10:9.
[19]Kvarnhammar AM,Cardell LO.Pattern-recognition receptors in human eosinophils[J].Immunology,2012,136(1):11-20.
[20]Tuxun T,Wang JH,Lin RY,et al.Th17/Treg imbalance in patients with liver cystic echinococcosis[J].Parasite Immunol,2012,34(11):520-527.
[21]Pang N,Zhang F,Ma X,et al.TGF-β/Smad signaling pathway regulates Th17/Treg balance during Echinococcus multilocularis infection[J].Int Immunopharmacol,2014,20(1):248-257.
[22]Truchetet ME,Beven L,Renaudin H,et al.Potential role of Mycoplasma hominis in interleukin(IL)-17-producing CD4+Tcell generation via induction of IL-23 secretion by human dendritic cells[J].J Infect Dis,2011,204(11):1796-1805.
[23]Shainheit MG,Lasocki KW,Finger E,et al.The pathogenic Th17 cell response to major schistosome egg antigen is sequentially dependent on IL-23 and IL-1β[J].J Immunol,2011,187(10):5328-5335.
[24]Lechner CJ,Grüner B,Huang X,et al.Parasite-specific IL-17-type cytokine responses and soluble IL-17 receptor levels in alveolar echinococcosis patients[J].Clin Dev Immunol,2012,2012:735342.
[25]吐尔洪江·吐逊,单骄宇,李涛,等.Th17细胞和调节性T细胞在肝包虫病免疫逃避中的作用[J].中华消化外科杂志,2010,9(4):283-286.
[26]Floss DM,Schr der J,Franke M,et al.Insights into IL-23biology:from structure to function[J].Cytokine Growth Factor Rev,2015,26(5):569-578.
[2]Bresson-Hadni SMJ,Mantion GA,Vuitton DA.Echinococcosis of the liver[M]//Rodes JBJ,Blei AT,Reichen M.Textbook of Hepatology.3rd edition.Oxford,UK:Blackwell Publishing,2007:1047-1057.
[3]单骄宇,吐尔洪江·吐逊,王俊华,等.慢性囊型包虫病患者外周血单个核细胞中TLR2、4、7 m RNA表达水平的研究[J].中国微生物学与免疫学杂志,2011,31(3):273-278.
[4]Trinchieri G,Sher A.Cooperation of Toll-like receptor signals in innate immune defenc e[J].Nat Rev Immunol,2007,7(3):179-90.
[5]黄蓉,吴青,李茂军,等.Toll样受体4在免疫相关细胞中的作用[J].中华临床医师杂志,2016,10(8):1178-1182.
[6]Ghosh D,Stumhofer JS.Do you see what I see:recognition of protozoan parasites by Toll-like receptors[J].Curr Immunol Rev,2013,9(3):129-140.
[7]WHO Informal Working Group.International classification of ultrasound images in cystic echinococcosis for application in clinical and field epidemiological settings[J].Acta Trop,2003,85(2):253-261.
[8]中国医师协会外科医师分会包虫病外科专业委员会.两型包虫病诊断和治疗的专家共识(2015版)[J].中华消化外科杂志,2015,14(4):253-264.
[9]黄玉娴,孔庆瑜,蔡高荣,等.三种检测嗜酸性粒细胞方法的比较[J].分子诊断与治疗杂志,2004,16(3):9-11.
[10]Tuxun T,Apaer S,Ma HZ,et al.The potential role of Th9cell related cytokine and transcription factors in patients with hepatic alveolar echinococcosis[J].J Immunol Res,2015,2015:895416.
[11]Vuitton DA.The ambiguous role of immunity in echinococcosis:protection of the host or of the parasite[J].Acta Trop,2003,85(2):119-132.
[12]Hübner MP,Manfras BJ,Margos MC,et al.Echinococcus multilocularis metacestodes modulate cellular cytokine and chemokine release by peripheral blood mononuclear cells in alveolar echinococcosis patients[J].Clin Exp Immunol,2006,145(2):243-51.
[13]Vuitton DA.Echinococcosis and allergy[J].Clin Rev Allergy Immunol,2004,26(2):93-104.
[14]Janeway CA Jr.Approaching the asymptote?Evolution and revolution in immunology[J].Cold Spring Harb Symp Quant Biol,1989,54 Pt 1:1-13.
[15]Kane CM,Jung E,Pearce EJ.Schistosoma mansoni egg antigenmediated modulation of Toll-like receptor(TLR)-induced activation occurs independently of TLR2,TLR4,and My D88[J].Infect Immun,2008,76(12):5754-5759.
[16]Smith P,Mangan NE,Fallon PG.Generation of parasite antigens for use in toll-like receptor research[J].Methods Mol Biol,2009,517:401-413.
[17]Kawai T,Akira S.The role of pattern-recognition receptors in innate immunity:update on toll-like receptors[J].Nat Immunol,2010,11(5):373-384.
[18]van Riet E,Everts B,Retra K,et al.Combined TLR2 and TLR4 ligation in the context of bacterial or helminth extracts in human monocyte derived dendritic cells:molecular correlates for Th1/Th2 polarization[J].BMC Immunol,2009,10:9.
[19]Kvarnhammar AM,Cardell LO.Pattern-recognition receptors in human eosinophils[J].Immunology,2012,136(1):11-20.
[20]Tuxun T,Wang JH,Lin RY,et al.Th17/Treg imbalance in patients with liver cystic echinococcosis[J].Parasite Immunol,2012,34(11):520-527.
[21]Pang N,Zhang F,Ma X,et al.TGF-β/Smad signaling pathway regulates Th17/Treg balance during Echinococcus multilocularis infection[J].Int Immunopharmacol,2014,20(1):248-257.
[22]Truchetet ME,Beven L,Renaudin H,et al.Potential role of Mycoplasma hominis in interleukin(IL)-17-producing CD4+Tcell generation via induction of IL-23 secretion by human dendritic cells[J].J Infect Dis,2011,204(11):1796-1805.
[23]Shainheit MG,Lasocki KW,Finger E,et al.The pathogenic Th17 cell response to major schistosome egg antigen is sequentially dependent on IL-23 and IL-1β[J].J Immunol,2011,187(10):5328-5335.
[24]Lechner CJ,Grüner B,Huang X,et al.Parasite-specific IL-17-type cytokine responses and soluble IL-17 receptor levels in alveolar echinococcosis patients[J].Clin Dev Immunol,2012,2012:735342.
[25]吐尔洪江·吐逊,单骄宇,李涛,等.Th17细胞和调节性T细胞在肝包虫病免疫逃避中的作用[J].中华消化外科杂志,2010,9(4):283-286.
[26]Floss DM,Schr der J,Franke M,et al.Insights into IL-23biology:from structure to function[J].Cytokine Growth Factor Rev,2015,26(5):569-578.