Friedman G B, Taylor C T, Parkos C A, Colgan S P
Center for Experimental Therapeutics and Reperfusion Injury, Department of Anesthesia, Boston, Massachusetts, USA.
J Cell Physiol. 1998 Jul;176(1):76-84. doi: 10.1002/(SICI)1097-4652(199807)176:1<76::AID-JCP9>3.0.CO;2-5.
Mucosal tissues, such as the lung and intestine, are primary targets for ischemic damage. Under these conditions, neutrophil (polymorphonuclear leukocyte; PMN) infiltration into the protective epithelium has been implicated as a pathophysiologic mediator. Because PMN transepithelial migration results in increased paracellular permeability, and because our previous data revealed that epithelial hypoxia enhances PMN transmigration, we hypothesized that macromolecular permeability may be altered in epithelium exposed to hypoxia and reoxygenation (H/R) in the presence of PMNs. Human intestinal epithelia (T84) were grown on permeable supports, exposed to cellular hypoxia (pO2 20 torr) for 0-72 hr, and examined for increases in PMN-evoked permeability by using standard flux assays. Increasing epithelial hypoxia potentiated PMN-induced permeability of labeled paracellular tracers (size range 3-500 kD). Such increases were blocked by monoclonal antibody (mAb) to the PMN integrin CD11b (82 +/- 1% decreased compared with control mAb) and were partially blocked by anti-CD47 mAb (51 +/- 1%). Assessment of barrier recovery revealed that monolayers exposed to H/R were significantly diminished in their ability to reseal following PMN transmigration (recovery of 36 +/- 6% in H/R vs. 94 +/- 2% in normoxic controls). Because intracellular cyclic AMP (cAMP) has been demonstrated to regulate epithelial permeability, and because PMN-derived compound(s), (i.e., 5'-adenosine monophosphate; AMP) elevate epithelial cAMP, we examined the impact of hypoxia on epithelial cAMP responses. These experiments revealed that hypoxic epithelia were diminished in their ability to generate cAMP, and pharmacologic elevation (8-bromo-cAMP) of intracellular cAMP in hypoxic cells normalized both PMN-induced permeability changes and restoration of barrier function. These results support a role for PMN in increased intestinal permeability associated with reperfusion injury and imply a substantial role for cAMP signaling in maintenance of permeability during PMN transmigration.
肺和肠道等黏膜组织是缺血性损伤的主要靶点。在这些情况下，中性粒细胞（多形核白细胞；PMN）浸润到保护性上皮细胞中被认为是一种病理生理介质。由于PMN跨上皮迁移会导致细胞旁通透性增加，并且我们之前的数据显示上皮细胞缺氧会增强PMN的迁移，因此我们推测在存在PMN的情况下，暴露于缺氧和复氧（H/R）的上皮细胞中大分子通透性可能会发生改变。人肠上皮细胞（T84）在可渗透支持物上生长，暴露于细胞缺氧（pO2 20托）0至72小时，并通过标准通量测定法检测PMN诱导的通透性增加情况。上皮细胞缺氧程度的增加增强了PMN诱导的标记细胞旁示踪剂（大小范围为3 - 500 kD）的通透性。这种增加被针对PMN整合素CD11b的单克隆抗体（mAb）阻断（与对照mAb相比降低了82 +/- 1%），并被抗CD47 mAb部分阻断（51 +/- 1%）。屏障恢复评估显示，暴露于H/R的单层细胞在PMN迁移后重新封闭的能力显著降低（H/R组恢复率为36 +/- 6%，而常氧对照组为94 +/- 2%）。由于细胞内环状AMP（cAMP）已被证明可调节上皮细胞通透性，并且由于PMN衍生的化合物（即5'-腺苷单磷酸；AMP）会升高上皮细胞cAMP，我们研究了缺氧对上皮细胞cAMP反应的影响。这些实验表明，缺氧上皮细胞产生cAMP的能力降低，并且缺氧细胞中细胞内cAMP的药理学升高（8-溴-cAMP）使PMN诱导的通透性变化和屏障功能恢复正常。这些结果支持PMN在与再灌注损伤相关的肠道通透性增加中起作用，并暗示cAMP信号在PMN迁移过程中维持通透性方面起重要作用。
