Inhibition of endothelial wound repair by dominant negative connexin inhibitors
Inhibition of endothelial wound repair by dominant negative connexin inhibitors
复制标题
DOI：
10.1091/mbc.12.4.831
复制
发表时间：
2001-04-01
影响因子：
3.3
通讯作者：
Meda, P
中科院分区：
文献类型：
--
作者：
Kwak, BR;Pepper, MS;Meda, P
内皮细胞的损伤与细胞间直接通讯的改变有关。为了确定间隙连接通讯是否参与伤口修复过程，我们比较了 PymT 转化的小鼠内皮细胞（克隆 bEnd.3）单层培养物和表达不同显性失活 Cx 抑制剂的 bEnd.3 细胞中的连接蛋白 (Cx) 表达、细胞间偶联和伤口修复动力学。在亲代 bEnd.3 细胞中，机械损伤导致损伤部位的 Cx43 表达增加，Cx37 表达减少，而 Cx40 表达不受影响。根据不同荧光示踪剂的评估，这些伤口诱导的 Cx 表达变化与细胞间偶联的功能变化相关。用编码嵌合连接蛋白 3243H7 或融合蛋白 Cx43-beta Gal 的 cDNA 进行稳定转染，会导致在基础条件和损伤条件下 bEnd.3 细胞之间的间隙连接通讯受到干扰。在表达显性阴性 Cx 抑制剂的细胞中，完全修复汇合单层内确定的伤口所需的时间增加了大约 50%，而其他细胞特性，如增殖率、单细胞迁移、囊肿形成和细胞外蛋白水解活性，则没有改变。这些发现表明，在内皮伤口修复过程中，协调迁移需要适当的 Cx 表达。
Wounding of endothelial cells is associated with altered direct intercellular communication. To determine whether gap junctional communication participates to the wound repair process, we have compared connexin (Cx) expression, cell-to-cell coupling and kinetics of wound repair in monolayer cultures of PymT-transformed mouse endothelial cells (clone bEnd.3) and in bEnd.3 cells expressing different dominant negative Cx inhibitors. In parental bEnd.3 cells, mechanical wounding increased expression of Cx43 and decreased expression of Cx37 at the site of injury, whereas Cx40 expression was unaffected. These wound-induced changes in Cx expression were associated with functional changes in cell-to-cell coupling, as assessed with different fluorescent tracers. Stable transfection with cDNAs encoding for the chimeric connexin 3243H7 or the fusion protein Cx43-beta Gal resulted in perturbed gap junctional communication between bEnd.3 cells under both basal and wounded conditions. The time required for complete repair of a defined wound within a confluent monolayer was increased by similar to 50% in cells expressing the dominant negative Cx inhibitors, whereas other cell properties, such as proliferation rate, migration of single cells, cyst formation and extracellular proteolytic activity, were unaltered. These findings demonstrate that proper Cx expression is required for coordinated migration during repair of an endothelial wound.