Janus激酶抑制剂 tyrphostin AG490在体外对原发性T细胞反应的抑制及同种异体抗原特异性低反应性的诱导

Suppression of primary T-cell responses and induction of alloantigen-specific hyporesponsiveness in vitro by the Janus kinase inhibitor tyrphostin AG490.

作者信息

Säemann M D, Böhmig G A, Osterreicher C H, Staffler G, Diakos C, Krieger P M, Hörl W H, Stockinger H, Zlabinger G J

机构信息

Institute of Immunology and Department of Internal Medicine III, University of Vienna, Austria.

出版信息

Transplantation. 2000 Oct 27;70(8):1215-25. doi: 10.1097/00007890-200010270-00016.

DOI:10.1097/00007890-200010270-00016

PMID:11063344

Abstract

BACKGROUND

Tyrphostin AG490 has recently been shown to block interleukin (IL)-2 receptor gamma-chain-associated Janus kinase 3. Here, we analyzed the effect of AG490 on T-cell alloresponses in vitro.

METHODS

For the evaluation of T-cell activation, DNA synthesis, surface marker expression, cytokine secretion, intracellular calcium mobilization, early protein tyrosine phosphorylation, and apoptosis were measured.

RESULTS

AG490 effectively inhibited T-cell proliferation in human mixed lymphocyte culture (MLC) even when added 4 days after culture initiation. Inhibition of IL-2-dependent proliferation in T-cell blasts and the incapability of IL-2 or IL-15 to restore proliferation in AG490-treated MLC suggests interference with cytokine receptor signaling. T-cell receptor-triggered early protein tyrosine phosphorylation, calcium mobilization, up-regulation of CD69, and initial CD25 expression were not affected. Interestingly, AG490 substantially inhibited production of IL-2 and interferon-gamma in T cells stimulated with alloantigen or via CD3 and CD28. In CD28-independent activation models (e.g., stimulation with phorbol myristate acetate plus ionomycin), however, cytokine secretion was not inhibited. Pretreatment of primary MLC with AG490 resulted in substantial down-regulation of secondary responses to cells from the original donor as opposed to third-party cells or phytohemagglutinin. Unresponsiveness was induced also in T cells stimulated with CD3 monoclonal antibody. Induction of apoptosis in polyclonally activated T cells and the incapability of IL-2 to reverse specific hyporesponsiveness, suggest programmed cell death as an important mechanism underlying antigen-specific down-regulation of alloresponses.

CONCLUSIONS

We demonstrate that AG490 blocks different manifestations of T-cell activation. This and its ability to induce alloantigen-specific hyporesponsiveness point to a potential use for interfering with alloreactivities in vivo.

摘要

背景

最近研究表明， tyrphostin AG490 可阻断白细胞介素（IL）-2 受体γ链相关的 Janus 激酶 3。在此，我们分析了 AG490 对体外 T 细胞同种异体反应的影响。

方法

为评估 T 细胞活化，检测了 DNA 合成、表面标志物表达、细胞因子分泌、细胞内钙动员、早期蛋白酪氨酸磷酸化及细胞凋亡情况。

结果

即使在培养开始 4 天后添加 AG490，其仍能有效抑制人混合淋巴细胞培养（MLC）中的 T 细胞增殖。AG490 处理的 MLC 中，IL-2 依赖性增殖受到抑制，且 IL-2 或 IL-15 无法恢复其增殖，这提示其对细胞因子受体信号传导有干扰作用。T 细胞受体触发的早期蛋白酪氨酸磷酸化、钙动员、CD69 的上调及初始 CD25 的表达均未受影响。有趣的是，AG490 能显著抑制同种异体抗原刺激或通过 CD3 和 CD28 刺激的 T 细胞中 IL-2 和干扰素-γ 的产生。然而，在不依赖 CD28 的活化模型中（如佛波酯肉豆蔻酸酯加离子霉素刺激），细胞因子分泌未受抑制。用 AG490 预处理原代 MLC 可导致对来自原始供体细胞的二次反应显著下调，而对第三方细胞或植物血凝素的反应则不然。用 CD3 单克隆抗体刺激的 T 细胞也会诱导无反应性。多克隆活化的 T 细胞中凋亡的诱导以及 IL-2 无法逆转特异性低反应性，提示程序性细胞死亡是同种异体反应抗原特异性下调的重要机制。