Induction of T lymphocyte apoptosis by sulphasalazine in patients with Crohn's disease
Induction of T lymphocyte apoptosis by sulphasalazine in patients with Crohn's disease
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DOI：
10.1136/gut.2003.037911
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发表时间：
2004-11-01
期刊：
影响因子：
24.5
通讯作者：
Ruemmele, FM
中科院分区：
文献类型：
--
作者：
Doering, J;Begue, B;Ruemmele, FM
背景：克罗恩病（CD）患者肠粘膜固有层T淋巴细胞（LPL）呈慢性激活状态。激活LPL的凋亡缺陷被认为是一个重要的致病机制。事实上，在CD LPL中观察到抗凋亡分子的表达增加。在目前的工作中，我们的目的是分析5-氨基水杨酸（5-阿萨）和衍生物对LPL和外周血淋巴细胞（PBL）的CD患者的凋亡的影响和潜在的分子机制与溃疡性结肠炎（UC）和非炎症controls.Methods：PBL和LPL分离Ficoll-Hypopaque梯度离心和EGTA-胶原酶法，分别。用FasL、5-阿萨、磺胺吡啶和柳氮磺胺吡啶刺激PBL/LPL 24/48小时，并通过流式细胞术（膜联蛋白V-碘化丙啶方法）和免疫荧光定量细胞凋亡。药物诱导凋亡的分子机制进行了分析，在野生型和FADD 2/2 Jurkat T细胞使用western blot和caspase assays.Results：虽然PBL显示正常的凋亡模式Fas刺激后，在活动性CD患者，LPL从炎症领域是高度抵抗。在UC患者的LPL中观察到相当的抗凋亡性。与不诱导淋巴细胞凋亡的5-阿萨相比，柳氮磺胺嘧啶被证明是一种有效的促凋亡剂。柳氮磺胺吡啶诱导的T淋巴细胞凋亡不依赖于Fas通路，但与抗凋亡基因bcl-xl和bcl-2的显著下调、线粒体凋亡信号通路的激活以及随后caspase-9和caspase-3的激活有关。柳氮磺胺吡啶在治疗炎症性肠病中的有益作用至少部分归因于其对LPL的促凋亡作用，淋巴细胞活化的过程。
Background: Lamina propria T lymphocytes (LPL) of the intestinal mucosa are chronically activated in Crohn's disease ( CD). Defective apoptosis of activated LPL was proposed as a key pathogenic mechanism. In fact, increased expression of antiapoptotic molecules was observed in CD LPL. In the present work, we aimed to analyse the effects and underlying molecular mechanisms of 5-amino salicylic acid (5-ASA) and derivatives on apoptosis of LPL and peripheral blood lymphocytes (PBL) in patients with CD compared with ulcerative colitis (UC) and in non-inflammatory controls.Methods: PBL and LPL were isolated by Ficoll-Hypopaque gradient centrifugation and the EGTA-collagenase method, respectively. PBL/LPL were stimulated with FasL, 5-ASA, sulphapyridine, and sulphasalazine for 24/48 hours and apoptosis was quantified by flow cytometry (annexin V-propidium iodide method) and immunofluorescence. The molecular mechanisms of drug induced apoptosis were analysed in wild-type and FADD2/2 Jurkat T cells using western blots and caspase assays.Results: While PBL displayed a normal apoptosis pattern after Fas stimulation in patients with active CD, LPL from inflammatory areas were highly resistant. Comparable resistance to apoptosis was observed in LPL of UC patients. In contrast with 5-ASA, which did not induce apoptosis in lymphocytes, sulphasalazine proved to be a potent proapoptotic agent. Sulphasalazine induced T lymphocyte apoptosis was independent of the Fas pathway but associated with marked downregulation of antiapoptotic bcl-xl and bcl2, activation of the mitochondrial apoptosis signalling pathway, and subsequent activation of caspase-9 and caspase-3.Conclusion: The beneficial effect of sulphasalazine in treating inflammatory bowel disease is at least in part attributable to its proapoptotic effects on LPL which allows potent downregulation of lymphocyte activation.