Department of Medicine, Indiana University School of Medicine, Indianapolis, 46202-5120, USA.
Am J Physiol Lung Cell Mol Physiol. 2011 Dec;301(6):L836-46. doi: 10.1152/ajplung.00385.2010. Epub 2011 Aug 26.
The epithelial and endothelial cells lining the alveolus form a barrier essential for the preservation of the lung respiratory function, which is, however, vulnerable to excessive oxidative, inflammatory, and apoptotic insults. Whereas profound breaches in this barrier function cause pulmonary edema, more subtle changes may contribute to inflammation. The mechanisms by which cigarette smoke (CS) exposure induce lung inflammation are not fully understood, but an early alteration in the epithelial barrier function has been documented. We sought to investigate the occurrence and mechanisms by which soluble components of mainstream CS disrupt the lung endothelial cell barrier function. Using cultured primary rat microvascular cell monolayers, we report that CS induces endothelial cell barrier disruption in a dose- and time-dependent manner of similar magnitude to that of the epithelial cell barrier. CS exposure triggered a mechanism of neutral sphingomyelinase-mediated ceramide upregulation and p38 MAPK and JNK activation that were oxidative stress dependent and that, along with Rho kinase activation, mediated the endothelial barrier dysfunction. The morphological changes in endothelial cell monolayers induced by CS included actin cytoskeletal rearrangement, junctional protein zonula occludens-1 loss, and intercellular gap formation, which were abolished by the glutathione modulator N-acetylcysteine and ameliorated by neutral sphingomyelinase inhibition. The direct application of ceramide recapitulated the effects of CS, by disrupting both endothelial and epithelial cells barrier, by a mechanism that was redox and apoptosis independent and required Rho kinase activation. Furthermore, ceramide induced dose-dependent alterations of alveolar microcirculatory barrier in vivo, measured by two-photon excitation microscopy in the intact rat. In conclusion, soluble components of CS have direct endothelial barrier-disruptive effects that could be ameliorated by glutathione modulators or by inhibitors of neutral sphingomyelinase, p38 MAPK, JNK, and Rho kinase. Amelioration of endothelial permeability may alleviate lung and systemic vascular dysfunction associated with smoking-related chronic obstructive lung diseases.
肺泡衬里的上皮细胞和内皮细胞形成了维持肺呼吸功能的屏障，但它们易受到过度氧化、炎症和凋亡的损伤。虽然这种屏障功能的严重破坏会导致肺水肿，但更细微的变化也可能导致炎症。虽然香烟烟雾（CS）暴露导致肺炎症的确切机制尚未完全阐明，但已记录到上皮屏障功能的早期改变。我们试图研究 CS 暴露引发肺炎症的机制，以及可溶性 CS 成分如何破坏肺内皮细胞屏障功能。我们使用培养的原代大鼠微血管细胞单层，报告 CS 以与上皮细胞屏障相似的幅度和时间依赖性方式诱导内皮细胞屏障破坏。CS 暴露触发了中性鞘磷脂酶介导的神经酰胺上调以及 p38 MAPK 和 JNK 激活的机制，该机制依赖于氧化应激，并且与 Rho 激酶激活一起介导内皮屏障功能障碍。CS 引起的内皮细胞单层形态变化包括肌动蛋白细胞骨架重排、连接蛋白闭合蛋白-1 丢失和细胞间间隙形成，这些变化被谷胱甘肽调节剂 N-乙酰半胱氨酸消除，并通过中性鞘磷脂酶抑制得到改善。CS 的直接应用通过一种不依赖于氧化还原和凋亡且需要 Rho 激酶激活的机制，破坏内皮细胞和上皮细胞的屏障，重现了 CS 的作用。此外，神经酰胺通过二光子激发显微镜在完整大鼠中体内诱导了剂量依赖性的肺泡微循环屏障改变。总之，CS 的可溶性成分具有直接的内皮屏障破坏作用，可通过谷胱甘肽调节剂或中性鞘磷脂酶、p38 MAPK、JNK 和 Rho 激酶抑制剂得到改善。内皮通透性的改善可能减轻与吸烟相关的慢性阻塞性肺病相关的肺和全身血管功能障碍。
