Distinct temporal-spatial roles for rho kinase and myosin light chain kinase in epithelial purse-string wound closure
Distinct temporal-spatial roles for rho kinase and myosin light chain kinase in epithelial purse-string wound closure
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DOI：
10.1053/j.gastro.2005.01.004
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发表时间：
2005-04-01
期刊：
影响因子：
29.4
通讯作者：
Turner, JR
中科院分区：
文献类型：
--
作者：
Russo, JM;Florian, P;Turner, JR
背景和目标：小的上皮伤口通过由肌球蛋白轻链（MLC）激酶（MLCK）和rho激酶（ROCK）调节的肌动球蛋白环的荷包收缩而愈合。这些研究旨在确定这些激酶在荷包缝合伤口闭合中的作用。方法：在肠上皮单层上造成寡细胞和单细胞伤口。在伤口闭合过程中收集荧光成像和电生理数据。人类活组织检查进行了化学研究。结果：增强型绿色荧光蛋白-β-肌动蛋白的活细胞成像确定了创伤后2分钟内快速的肌动蛋白环组装。这在8分钟内发展为周向环，随后收缩并闭合伤口。因此，我们将此过程分为2个阶段：环组装和伤口收缩。在成形环组装过程中，活化rho和ROCK定位于伤口边缘。与组装阶段的主要作用一致，ROCK抑制阻止肌动蛋白环组装和伤口闭合。环组装完成后的ROCK抑制没有效果。MLCK的募集和激活发生在环组装完成后，与环收缩一致。MLCK抑制减慢，然后停止收缩，但不能阻止环组装。MLCK抑制也延迟屏障功能恢复。人类结肠活检标本的研究表明，荷包缝合伤口也发生在体内，因为MLC磷酸化增强周围寡细胞伤口。结论：这些结果表明这些激酶在实验和体内寡细胞上皮伤口的荷包闭合中的互补作用; rho和ROCK对肌动蛋白环组装至关重要，而MLCK的活性驱动收缩。
Background & Aims: Small epithelial wounds heal by purse-string contraction of an actomyosin ring that is regulated by myosin light chain (MLC) kinase (MLCK) and rho kinase (ROCK). These studies aimed to define the roles of these kinases in purse-string wound closure. Methods: Oligocellular and single-cell wounds were created in intestinal epithelial monolayers. Fluorescence imaging and electrophysiologic data were collected during wound closure. Human biopsies were studied immunohistochemically. Results: Live-cell imaging of enhanced green fluorescent protein-p-actin defined rapid actin ring assembly within 2 minutes after wounding. This progressed to a circumferential ring within 8 minutes that subsequently contracted and closed the wound. We therefore divided this process into 2 phases: ring assembly and wound contraction. Activated rho and ROCK localized to the wound edge during ring assembly. Consistent with a primary role in the assembly phase, ROCK inhibition prevented actin ring assembly and wound closure. ROCK inhibition after ring assembly was complete had no effect. Recruitment and activation of MLCK occurred after ring assembly was complete and coincided with ring contraction. MLCK inhibition slowed and then stopped contraction but did not prevent ring assembly. MLCK inhibition also delayed barrier function recovery. Studies of human colonic biopsy specimens suggest that purse-string wound closure also occurs in vivo, because MLC phosphorylation was enhanced surrounding oligocellular wounds. Conclusions: These results suggest complementary roles for these kinases in purse-string closure of experimental and in vivo oligocellular epithelial wounds; rho and ROCK are critical for actin ring assembly, while the activity of MLCK drives contraction.