阻断Notch1信号通路对慢性乙肝患者外周血T细胞免疫功能的影响

中国临床药理学与治疗学 ›› 2011, Vol. 16 ›› Issue (2): 174-179.

阻断Notch1信号通路对慢性乙肝患者外周血T细胞免疫功能的影响

刘雪妮, 裴金仙, 李小亮, 臧国庆, 余永胜

上海交通大学附属第六人民医院感染病科,上海 200233

收稿日期:2011-02-01 修回日期:2011-02-12 发布日期:2011-04-20
通讯作者: 余永胜,男,主任医师,硕导,研究方向:乙型病毒性肝炎的免疫机制。E-mail: yuyongsheng@medmail.com
作者简介:刘雪妮,女,硕士研究生,研究方向:乙型病毒性肝炎的免疫机制。Tel: 15800853235 E-mail: zsk370@163.com

Effect of blockage of Notch1 signaling on the activation of T cellular immunity in peripheral blood mononuclear cells of patients with chronic hepatitis B

LIU Xue-ni, PEI Jin-xian, LI Xiao-liang, ZANG Guo-qing, YU Yong-sheng

Department of Infectious Diseases, Shanghai Sixth People’s Hospital, Shanghai Jiao Tong University, Shanghai 200233, China

Received:2011-02-01 Revised:2011-02-12 Published:2011-04-20

摘要/Abstract

摘要： 目的:观察阻断Notch1信号对慢性乙型病毒性肝炎患者外周血单个核细胞(PBMC)T-bet表达,Th1和Th2型细胞因子表达以及细胞毒性T淋巴细胞(CTL)细胞毒活性的影响,探讨Notch1信号通路在调节慢性乙型病毒性肝炎患者外周血T细胞免疫功能的作用。方法: 将慢性乙型病毒性肝炎患者PBMC随机分为实验组,阴性对照组和空白对照组,分别给予抗-Notch1,IgG2b和RPMI-1640培养液,采用实时荧光定量PCR(real-time Q-PCR)检测各组T-bet mRNA的表达情况,采用ELISA法检测Th1型细胞因子IFN-γ,IL-2和Th2型细胞因子IL-4,IL-10的分泌水平,采用乳酸脱氢酶释放法检测CTL对靶细胞HepG2.2.15细胞毒活性的变化。结果: 用抗体阻断Notch1信号通路后,慢性乙型病毒性肝炎患者PBMC中T-bet mRNA表达增强,Th1型细胞因子IFN-γ和IL-2浓度增高,而Th2型细胞因子IL-4和IL-10浓度减少,且CTL细胞毒活性明显增强,与阴性对照组和空白对照组相比差异有统计学意义(P均<0.01)。结论:阻断Notch1信号通路可增强慢性乙肝患者PBMC中T-bet mRNA表达,使Th1型细胞因子分泌增加,Th2型细胞因子分泌减少,且使CTL细胞毒活性增强,说明阻断Notch1信号通路可增强慢性乙肝患者T细胞免疫反应,Notch1信号通路可能参与慢性乙型病毒性肝炎患者机体的免疫耐受,且可能与T-bet有关。

关键词: 慢性乙肝, Notch1, T细胞免疫, T-bet, CTL

Abstract: AIM: To investigate the influence of Notch1 signaling on expression of T-bet, Th1- and Th2-type cytokines and cytotoxic T-lymphocyte (CTL) cytotoxicity of peripheral blood mononuclear cells (PBMC) of patients with chronic hepatitis B, and analyze the possible function of Notch1 signaling on regulating the activation of T cellular immunity.METHODS: PBMC of patients with chronic hepatitis B were divided into test group, negative control group and blank control group, respectively was given anti-Notch1, IgG2b and RPMI-1640. The expression of T-bet mRNA was examined by real-time PCR, Th1-type cytokines IFN-γ,IL-2, Th2-type cytokines IL-4,IL-10 were assayed by enzyme-linked immunosorbent assay (ELISA) and CTL cytotoxicity was measured by Cytotoxicity Detection Assay (LDH).RESULTS:After Notch1 signaling of PBMC was blocked in chronic hepatitis B patients, expression of T-bet mRNA and the production of Th1-type cytokines IFN-γ,IL-2 were enhanced while Th2-type cytokines IL-4,IL-10 were decreased, and CTL cytotoxicity was enhanced, compared with the blank control groups (P<0.01).CONCLUSION: After blocking of Notch1 signaling of PBMC of chronic hepatitis B patients, the expression of T-bet mRNA and the production of Th1-type cytokines IFN-γ,IL-2 were increased, while Th2-type cytokines IL-4,IL-10 were decreased and CTL cytotoxicity was enhanced. It suggested that blockage of Notch1 signaling could activate T cellular immunity of patients with chronic hepatitis B, Notch1 signaling might contribute to immune tolerance of patients with chronic hepatitis B, perhaps related with T-bet.

Key words: Chronic hepatitis B, Notch1, T cellular immunity, T-bet, CTL

中图分类号:

R512.6

刘雪妮, 裴金仙, 李小亮, 臧国庆, 余永胜. 阻断Notch1信号通路对慢性乙肝患者外周血T细胞免疫功能的影响[J]. 中国临床药理学与治疗学, 2011, 16(2): 174-179.

LIU Xue-ni, PEI Jin-xian, LI Xiao-liang, ZANG Guo-qing, YU Yong-sheng. Effect of blockage of Notch1 signaling on the activation of T cellular immunity in peripheral blood mononuclear cells of patients with chronic hepatitis B[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2011, 16(2): 174-179.

参考文献

[1] Radtke F, Wilson A, Mancini SJ, et al. Notch regulation of lymphocyte development and function [J]. Nat Immunol, 2004, 5(3): 247-53.
[2] Osborne BA, Minter LM. Notch signaling during peripheral T-cell activation and differentiation [J]. Nat Rev Immunol, 2007, 7(1): 64-75.
[3] Szabo SJ,Kim ST,Costa GL,et a1. A novel transcription factor, T-bet, directs Thl lineage commitment [J].Cell, 2000, 100(6):655-669.
[4] Szabo SJ, Sullivan BM, Stemmann C, et al .Distinct effects of T-bet in Th1 lineage commitment and IFN2 gamma production in CD₄ and CD₈ T cell [J]. Science, 2002, 295(5553): 338-342.
[5] Rehermann B, Nascimbeni M. Immunology of hepatitis B virus and hepatitis C virus infection [J]. Nat Rev Immunol, 2005, 5(3):215-229.
[6] Bcher WO, Galun E, Marcus H, et al. Reduced hepatitis B virus surface antigen-specific Th1 helper cell frequency of chronic HBV carriers is associated with a failure to produce antigen-specific antibodies in the Trimera mouse [J]. Hepatology, 2000, 31(2):480-7.
[7] Iso T, Kedes L, Hamamori Y. HES and HERP families: multiple effectors of the notch signaling pathway [J]. J Cell Physiol, 2003, 194(3):237-255.
[8] Amsen D, Vlander JM, Lee GR, et al. Instruction of distinct CD₄ T helper cell fates by different Notch ligands on antigen-presenting cells [J]. Cell, 2004, 117(4):515-526.
[9] Brooks DG, Teyton L, Oldstone MB, et al. Intrinsic functional dysregulation of CD₄ T cells occurs rapidly following persistent viral infection [J]. J Virol, 2005, 79(16):10514-27.
[10]裴金仙, 臧国庆, 余永胜, 等. 乙型肝炎患者外周血CD₄⁺T细胞Notch1蛋白的表达 [J].胃肠病学和肝病学杂志, 2009, 18 (12): 1136-38.
[11]Usui T, Preiss JC, Kanno Y, et al. T-bet regulates Th1 responses through essential effects on GATA-3 function rather than on IFNG gene acetylation and transcription [J]. J Exp Med, 2006, 203(3):755-766.
[12]Wang SY, Yang M, Xu XP, et al. Intranasal delivery of T-bet modulates the profile of helper T cell immune responses in experimental asthma [J]. J Investig Allergol Clin Immunol, 2008, 18(5): 357-65.
[13]Lametschwandtner G, Biedermann T, Schwarzler C, et al. Sustained T-bet expression confers polarized human Th2 cell with Th1-like cytokine production and migratory capacities [J]. J Allergy Clin Immunol, 2004, 113(5):987-994.