Luminal Cysteine-Proteases Degrade Colonic Tight Junction Structure and Are Responsible for Abdominal Pain in Constipation-Predominant IBS
Luminal Cysteine-Proteases Degrade Colonic Tight Junction Structure and Are Responsible for Abdominal Pain in Constipation-Predominant IBS
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DOI：
10.1038/ajg.2013.152
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发表时间：
2013-08-01
影响因子：
9.8
通讯作者：
Bueno, Lionel
中科院分区：
文献类型：
--
作者：
Annahazi, Anita;Ferrier, Laurent;Bueno, Lionel
目的：肠腔丝氨酸蛋白酶导致肠易激综合征（IBS-D）患者结肠细胞旁通透性增加和内脏高敏感性。其他蛋白酶，即半胱氨酸蛋白酶（CP），通过消化上皮紧密连接蛋白来增加气道通透性。在这项研究中，我们专注于便秘型IBS（IBS-C），我们的目的是（i）评估两组IBS患者的CP水平，（ii）测试IBS-C粪便上清液（FSN）是否影响小鼠重复给药后的通透性和内脏敏感性，以及（iii）评估IBS-C结肠活检组织中的occludin表达。使用选择性底物和抑制剂（E64）测定粪便CP活性。评价木瓜蛋白酶（作为阳性对照）和IBS-C FSN施用对小鼠和T84单层中结肠细胞旁通透性和粘膜闭合蛋白水平的影响。在IBS-C结肠活组织检查中评价了闭塞蛋白水平。重复给药IBS-C FSN.RESULTS：我们发现在IBS-C患者的一个子集中，粪便CP活性增强，与健康对照组和IBS-D患者相比，结肠直肠扩张（CRD）的敏感性进行了测定。CP活性水平与疾病严重程度和腹痛评分呈正相关。受试者工作特征曲线分析证实了这种关联。在小鼠中，反复将IBS-C FSN应用于结肠引发了渗透性增加，与occludin的酶降解有关，并与内脏对CRD的敏感性增强有关。最后，在IBS-C患者的结肠活检中发现闭合蛋白水平降低，IBS-C FSN能够在体外降解重组人闭合蛋白。所有这些作用都被预先孵育IBS-CFSN和CP抑制剂E64所消除。结论：这些数据表明，管腔CP可能代表了一个新的因素，有助于IBS症状的发生。
OBJECTIVES: Luminal serine-proteases lead to increased colonic paracellular permeability and visceral hypersensitivity in patients with diarrhea-predominant irritable bowel syndrome (IBS-D). Other proteases, namely cysteine-proteases (CPs), increase airway permeability by digesting epithelial tight junction proteins. In this study, we focused on constipation-predominant IBS (IBS-C) and we aimed to (i) evaluate CP levels in two cohorts of IBS patients, (ii) test if IBS-C fecal supernatant (FSN) affects permeability, and visceral sensitivity after repeated administrations in mice, and (iii) evaluate occludin expression in IBS-C colonic biopsies.METHODS: Fecal CP activity was determined using selective substrate and inhibitor (E64). The effect of papain, as positive control, and IBS-C FSN administrations were evaluated on colonic paracellular permeability and mucosal occludin levels in mice and T84 monolayers. Occludin protein levels were evaluated in IBS-C colonic biopsies. Sensitivity to colorectal distension (CRD) was measured after repeated administrations of IBS-C FSN.RESULTS: We found in a subset of IBS-C patients an enhanced fecal CP activity, in comparison with healthy controls and IBS-D patients. CP activity levels positively correlated with disease severity and abdominal pain scoring. This association was confirmed by receiver operating characteristic curve analysis. In mice, repeated application of IBS-C FSN into colon triggered increased permeability, linked to the enzymatic degradation of occludin, and was associated with enhanced visceral sensitivity to CRD. Finally, occludin levels were found decreased in colonic biopsies from IBS-C patients, and IBS-C FSNs were able to degrade recombinant human occludin in vitro. All these effects were abolished by preincubation of IBS-C FSN with a CP inhibitor, E64.CONCLUSIONS: These data suggest that luminal CPs may represent a new factor contributing to the genesis of symptoms in IBS.