HBe Ag对髓样树突状细胞Toll样受体4及NF-κB信号通路的影响
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摘要
目的探讨HBe Ag对髓样树突状细胞(mDC)Toll样受体4(TLR4)及NF-κB信号通路的影响。方法将健康者外周血单核细胞诱导分化为未成熟mDC;加入HBe Ag体外刺激,经脂多糖(LPS)刺激获得成熟mDC。采用Western blot法检测TLR4、NF-κB信号通路蛋白相对表达量,MTS法检测mDC刺激同种异体淋巴细胞增殖能力。结果培养至第9天,HBe Ag刺激组TLR4、NF-κB相对表达量为0.12±0.01、0.75±0.12,分别低于对照组的0.27±0.03、1.20±0.13(均P<0.05)。在DC/淋巴细胞比例为1∶5、1∶10、1∶20的反应中,HBe Ag刺激组刺激指数为2.93±0.05、2.56±0.19、1.44±0.09,分别低于对照组的4.83±0.05、3.57±0.35、2.13±0.11(均P<0.05)。结论 HBe Ag抑制mDC TLR4信号通路蛋白的表达,减弱mDC刺激同种异体淋巴细胞增殖能力。
Objective To investigate the effect of HBeAg on the toll-like receptor 4(TLR4) and nuclear factor κB(NF-κB)signaling pathway in human monocyte-derived dendritic cells(m DCs). Methods The peripheral blood mononuclear cells were isolated from healthy volunteers, and then induced and proliferated to immature m DC in vitro. Mature m DCs were obtained after HBeAg and lipopolysaccharide stimulation. The expression of TLR4 and NF-κB proteins was detected wit Western blot, and the response of allogeneic lymphocytes was examined with MTS method. Results The expression of TLR4 and NF-κB proteins decreased in the HBeAg-stimulated groups( 0. 12 ± 0. 01, 0. 75 ± 0. 12) compared with that of control group(0.27 ±0.03, 1.20±0.13, P<0.05). In response to the DC/lymphocyte with ratio of 1:5, 1:10 and 1:20, the stimulation indexes of the HBeAg stimulation group was 2.93 ±0.05,2.56 ±0.19, 1.44 ±0.09 respectively, which were lower than that of the control group(4.83 ±0.05, 3.57±0.35, 2.13 ±0.11, P<0.05).Conclusion HBeAg can inhibit TLR4 and NF-κB signaling pathway of m DC, weaken the ability of stimulating T cell proliferation.
Objective To investigate the effect of HBeAg on the toll-like receptor 4(TLR4) and nuclear factor κB(NF-κB)signaling pathway in human monocyte-derived dendritic cells(m DCs). Methods The peripheral blood mononuclear cells were isolated from healthy volunteers, and then induced and proliferated to immature m DC in vitro. Mature m DCs were obtained after HBeAg and lipopolysaccharide stimulation. The expression of TLR4 and NF-κB proteins was detected wit Western blot, and the response of allogeneic lymphocytes was examined with MTS method. Results The expression of TLR4 and NF-κB proteins decreased in the HBeAg-stimulated groups( 0. 12 ± 0. 01, 0. 75 ± 0. 12) compared with that of control group(0.27 ±0.03, 1.20±0.13, P<0.05). In response to the DC/lymphocyte with ratio of 1:5, 1:10 and 1:20, the stimulation indexes of the HBeAg stimulation group was 2.93 ±0.05,2.56 ±0.19, 1.44 ±0.09 respectively, which were lower than that of the control group(4.83 ±0.05, 3.57±0.35, 2.13 ±0.11, P<0.05).Conclusion HBeAg can inhibit TLR4 and NF-κB signaling pathway of m DC, weaken the ability of stimulating T cell proliferation.
引文
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[18]Das D,Sarkar N,Sengupta I,et al.Anti-viral role of toll like receptor 4 in hepatitis B virus infection:An in vitro study[J].World J Gastroenterol,2016,22(47):10341-10352.DOI:10.3748/wjg.v22.i47.10341.
[19]Lang T,Lo C,Skinner N,et al.The hepatitis B e antigen(HBe Ag)targets and suppresses activation of the toll-like receptor signaling pathway[J].J Hepatol,2011,55(4):762-769.DOI:10.1016/j.jhep.2010.12.042.
[2]Cheng X,Xia Y,Serti E,et al.Hepatitis B virus evades innate immunity of hepatocytes but activates cytokine production by macrophages[J].Hepatology,2017,66(6):1779-1793.DOI:10.1002/hep.29348.
[3]Boonstra A,Woltman AM,Janssen HL.Immunology of hepatitis B and hepatitis C virus infections[J].Best Pract Res Clin Gastroenterol,2008,22(6):1049-1061.DOI:10.1016/j.bpg.2008.11.015.
[4]Kim SJ,Diamond B.Modulation of tolerogenic dendritic cells and autoimmunity[J].Semin Cell Dev Biol,2015,41:49-58.DOI:10.1016/j.semcdb.2014.04.020.
[5]Kis-Toth K,Szanto A,Thai TH,et al.Cytosolic DNA-activated human dendritic cells are potent activators of the adaptive immune response[J].JImmunol,2011,187(3):1222-1234.DOI:10.4049/jimmunol.1100469.
[6]Ma YJ,He M,Han JA,et al.A clinical study of HBs Ag-activated dendritic cells and cytokine-induced killer cells during the treatment for chronic hepatitis B[J].Scand J Immunol,2013,78(4):387-393.DOI:10.1111/sji.12097.
[7]Zhang Z,Zhang JY,Wang LF,et al.Immunopathogenesis and prognostic immune markers of chronic hepatitis B virus infection[J].J Gastroenterol Hepatol,2012,27(2):223-230.DOI:10.1111/j.1440-1746.2011.06940.x.
[8]Nieto JC,Sanchez E,Roman E,et al.Cytokine production in patients with cirrhosis and TLR4 polymorphi sms[J].World J Gastroenterol,2014,20(46):17516-17524.DOI:10.3748/wjg.v20.i46.17516.
[9]Balmasova IP,Yushchuk ND,Mynbaev OA,et al.Immunopathogenesis of chronic hepatitis B[J].World J Gastroenterol,2014,20(39):14156-14171.DOI:10.3748/wjg.v20.i39.14156.
[10]Chen LM,Fan XG,Ma J,et al.Molecular mechanisms of HBe Ag in persistent HBV infection[J].Hepatol Int,2017,11(1):79-86.DOI:10.1007/s12072-016-9734-5.
[11]Liaw YF.Natural history of chronic hepatitis B virus infection and long-term outcome under treatment[J].Liver Int,2009,29(Suppl 1):100-107.DOI:10.1111/j.1478-3231.2008.01941.x.
[12]Vander MRG,Sprengers D,Biesta PJ,et al.Favorable effect of adefovir on the number and functionality of myeloid dendritic cells of patients with chronic HBV[J].Hepatology,2006,44(4):907-914.DOI:10.1002/hep.21340.
[13]Lin C,Zou H,Wang S.Hepatitis B e Antigen Seroconversion Is Related with the Function of Dendritic Cells in Chronic Hepatitis B Virus Infection[J].Gastroenterol Res Pract,2014,2014:413952.DOI:10.1155/2014/413952.
[14]燕飞,汤永志,潘柯传,等.乙型肝炎表面抗原对单核细胞来源的树突状细胞Toll样受体4信号通路的影响[J].中华传染病杂志,2014,32(2):94-99.DOI:10.3760/cma.j.issn.1000-6680.2014.02.007.
[15]Han Y,Li J,Jiang L,et al.Regulation of B7-H1 expression on peripheral monocytes and IFN-gamma secretion in T lymphocytes by HBe Ag[J].Cell Immunol,2013,283(1-2):25-30.DOI:10.3760/cma.j.issn.0254-6450.2017.10.023.
[16]Purvina M,Hoste A,Rossignol JM,et al.Human hepatitis B viral e antigen and its precursor P20 inhibit T lymphocyte proliferation[J].Biochem Biophys Res Commun,2012,417(4):1310-1315.DOI:10.1016/j.bbrc.2011.12.138.
[17]Jegaskanda S,Ahn SH,Skinner N,et al.Downregulation of interleukin-18-mediated cell signaling and interferon gamma expression by the hepatitis B virus e antigen[J].J Virol,2014,88(18):10412-10420.DOI:10.1128/JVI.00111-14.
[18]Das D,Sarkar N,Sengupta I,et al.Anti-viral role of toll like receptor 4 in hepatitis B virus infection:An in vitro study[J].World J Gastroenterol,2016,22(47):10341-10352.DOI:10.3748/wjg.v22.i47.10341.
[19]Lang T,Lo C,Skinner N,et al.The hepatitis B e antigen(HBe Ag)targets and suppresses activation of the toll-like receptor signaling pathway[J].J Hepatol,2011,55(4):762-769.DOI:10.1016/j.jhep.2010.12.042.