Connexin (Cx) family members form hemichannels (HCs) and gap junctions (GJs). Biological functions of Cx HCs have not been adequately characterized due to the inability to selectively target HCs or GJs. Recently, we developed a 6-mer peptide mimetic (P5) of the first extracellular loop of Cx43 and showed that it can block the permeability of HCs but not GJs formed by Cx43. In this study, we further characterized the HC blocking property of P5 and investigated the role of Cx HCs in acute lung injury (ALI). We found that P5 administration decreased HC permeability, in pulmonary microvascular endothelial cells, HepG2 cells, and even Cx43-deficient astrocytes, which express different sets of Cxs, suggesting that P5 is a broad spectrum Cx HC blocker. In addition, P5 reduced HC permeability of alveolar cells in vivo. Moreover, P5 decreased endotoxin-induced release, by vascular endothelial cells in vitro, of high mobility group box protein 1 (HMGB1), a critical mediator of acute lung injury (ALI), and reduced HMGB1 accumulation in bronchoalveolar lavage fluid (BALF) of mice subjected to intratracheal endotoxin instillation. Furthermore, P5 administration resulted in a significant decrease in the concentrations of ALT, AST, and LDH in the BALF, the accumulation of leukocytes in alveoli, and the mortality rate of mice subjected to ALI. Wright-Giemsa staining showed that P5 caused similar reductions of both neutrophils and monocytes in BALF of ALI mice. Together, these results suggest that Cx HCs mediate HMGB1 release, augment leukocyte recruitment, and contribute to ALI pathology.

中文翻译：

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连接蛋白半通道在急性肺损伤发病机制中的作用

连接蛋白 (Cx) 家族成员形成半通道 (HC) 和间隙连接 (GJ)。由于无法选择性靶向 HC 或 GJ，Cx HC 的生物学功能尚未得到充分表征。最近，我们开发了 Cx43 的第一个细胞外环的 6 聚体肽模拟物 (P5)，并表明它可以阻断 HCs 的渗透性，但不能阻断 Cx43 形成的 GJs。在本研究中，我们进一步表征了 P5 的 HC 阻断特性，并研究了 Cx HCs 在急性肺损伤 (ALI) 中的作用。我们发现，在肺微血管内皮细胞、HepG2 细胞，甚至 Cx43 缺陷的星形胶质细胞中，P5 给药降低了 HC 通透性，这些星形胶质细胞表达不同的 Cxs，表明 P5 是一种广谱的 Cx HC 阻滞剂。此外，P5 降低了体内肺泡细胞的 HC 通透性. 此外，P5 减少了内毒素诱导的体外血管内皮细胞释放高迁移率族框蛋白 1 (HMGB1)，这是一种急性肺损伤 (ALI) 的关键介质，并减少了支气管肺泡灌洗液 (BALF) 中 HMGB1 的积累。小鼠接受气管内内毒素滴注。此外，P5 给药导致 BALF 中 ALT、AST 和 LDH 的浓度、肺泡中白细胞的积累以及遭受 ALI 的小鼠的死亡率显着降低。Wright-Giemsa 染色显示 P5 导致 ALI 小鼠 BALF 中中性粒细胞和单核细胞的类似减少。总之，这些结果表明 Cx HCs 介导 HMGB1 释放，增加白细胞募集，并有助于 ALI 病理。