Connexin43 hemichannels contributes to the disassembly of cell junctions through modulation of intracellular oxidative status.-猫眼课题宝

DOI：

10.1016/j.redox.2016.08.008

发表时间：

2016-10

期刊：

REDOX BIOLOGY

影响因子：

11.4

通讯作者：

中科院分区：

生物学1区

文献类型：

作者：

Chi, Yuan;Zhang, Xiling;Zhang, Zhen;Mitsui, Takahiko;Kamiyama, Manabu;Takeda, Masayuki;Yao, Jian

关键词：

Connexin43 Hemichannel GSH Oxidative stress Tight junction Adherens junction

来源链接：

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5007435/?report=classic

连接蛋白(CX)半通道调节许多细胞过程，但对其机制知之甚少。鉴于许多激活半通道的病理因素也破坏了细胞连接的完整性，我们推测半通道可能参与了细胞连接的调节。在这里，我们检验了这一假设。肾小管上皮细胞暴露于无钙介质中可导致紧密连接和粘连连接的解体，表现为ZO-1和钙粘附素水平降低，F-肌动蛋白解体，跨上皮电阻值严重下降。在这些变化之前，Cx43半通道被激活，细胞外的ATP外流和荧光黄的内流揭示了这一点。用化学抑制剂或Cx43 siRNA抑制半管连接可大大减弱细胞连接的解体。对来自Cx43野生型(Cx43+/+)、杂合子(Cx43+/-)和基因敲除(Cx43-/-)的胎儿成纤维细胞的进一步分析表明，与Cx43阴性细胞(Cx43-/-)相比，Cx43阳性细胞(Cx43+/+)在细胞形态、F-肌动蛋白和钙粘素方面表现出更显著的变化。这些细胞始终具有较高的蛋白质羰基修饰和磷酸化水平，以及更强的P38和JNK活性。半通道开放导致细胞外主要抗氧化剂谷胱甘肽(GSH)的损失。补充外源性GSH或抑制氧化敏感酶可在很大程度上阻止上述变化。综上所述，我们的研究表明，Cx43半通道通过调节细胞内的氧化状态促进细胞连接的分解。关于细胞连接的协调调控机制尚不清楚。Ca~(2+)的耗竭激活了半通道，破坏了细胞连接。半通道开放通过GSH的外流加重了氧化应激。阻断半通道可减轻氧化应激和细胞连接解体。半通道通过调节细胞内氧化还原状态来调节细胞连接。

Connexin (Cx) hemichannels regulate many cellular processes with little information available regarding their mechanisms. Given that many pathological factors that activate hemichannels also disrupts the integrity of cellular junctions, we speculated a potential participation of hemichannels in the regulation of cell junctions. Here we tested this hypothesis. Exposure of renal tubular epithelial cells to Ca2+-free medium led to disassembly of tight and adherens junctions, as indicated by the reduced level of ZO-1 and cadherin, disorganization of F-actin, and severe drop in transepithelial electric resistance. These changes were preceded by an activation of Cx43 hemichannels, as revealed by extracellular efflux of ATP and intracellular influx of Lucifer Yellow. Inhibition of hemichannels with chemical inhibitors or Cx43 siRNA greatly attenuated the disassembly of cell junctions. Further analysis using fetal fibroblasts derived from Cx43 wide-type (Cx43+/+), heterozygous (Cx43+/-) and knockout (Cx43-/-) littermates showed that Cx43-positive cells (Cx43+/+) exhibited more dramatic changes in cell shape, F-actin, and cadherin in response to Ca2+ depletion, as compared to Cx43-null cells (Cx43-/-). Consistently, these cells had higher level of protein carbonyl modification and phosphorylation, and much stronger activation of P38 and JNK. Hemichannel opening led to extracellular loss of the major antioxidant glutathione (GSH). Supplement of cells with exogenous GSH or inhibition of oxidative sensitive kinases largely prevented the above-mentioned changes. Taken together, our study indicates that Cx43 hemichannels promote the disassembly of cell junctions through regulation of intracellular oxidative status. The mechanisms about the coordinated regulation of cell junctions are obscure. Ca2+ depletion activates hemichannels and disrupts cell junctions. Hemichannel opening exaggerates oxidative stress via efflux of GSH. Blocking hemichannels attenuates oxidative stress and cell junction disassembly. Hemichannels regulate cell junctions via modulation of intracellular redox status.