Baldwin Cindy, Chen Zhongchuan Will, Bedirian Arda, Yokota Naoko, Nasr Samih H, Rabb Hamid, Lemay Serge
Dept. of Medicine, Division of Nephrology, McGill Univ., Lyman-Duff Bldg. Rm. 228, 3775 Univ. St., Montreal, Quebec, Canada.
Am J Physiol Renal Physiol. 2006 Nov;291(5):F960-71. doi: 10.1152/ajprenal.00020.2006. Epub 2006 May 30.
Ischemia-reperfusion injury (IRI) is a major cause of acute kidney injury in both native kidneys and renal allografts. Disruption of the actin cytoskeleton is a key event with multiple repercussions on cell adhesion and function during IRI. However, receptors involved in regulating cytoskeletal repair following injury have not been identified. In an in vivo model of renal IRI, we used multiprobe RNase protection assay to examine the expression of Eph receptor tyrosine kinases, key regulators of actin dynamics in embryonic development. We found that one receptor in particular, EphA2, was strongly upregulated in the kidney following IRI. Ephrins, the cell-bound ligands of Eph receptors, were also strongly expressed. In cultured renal tubular cells, diverse injurious stimuli mimicking IRI also resulted in upregulation of EphA2 protein expression. Upregulation of EphA2 was inhibited by the Src kinase inhibitor PP2. Conversely, overexpression of Src kinases strongly enhanced the expression of endogenous EphA2 as well as the activity of a human EphA2 promoter construct. Activation of the Erk pathway was necessary, but not sufficient for full induction of EphA2 upreglation by Src kinases. Stimulation of renal tubular epithelial cells with the EphA2 ligand ephrin-A1 caused tyrosine phosphorylation of endogenous EphA2, paxillin, and an unidentified approximately 65-kDa protein and resulted in increased cortical F-actin staining. In summary, under in vitro conditions mimicking IRI, EphA2 expression is strongly upregulated through a Src kinase-dependent pathway. Interactions between upregulated EphA2 and its ephrin ligands may provide critical cell contact-dependent, bidirectional cues for cytoskeletal repair in renal IRI.
缺血再灌注损伤（IRI）是导致天然肾和肾移植急性肾损伤的主要原因。肌动蛋白细胞骨架的破坏是IRI期间对细胞黏附和功能产生多种影响的关键事件。然而，损伤后参与调节细胞骨架修复的受体尚未被确定。在肾IRI的体内模型中，我们使用多探针核糖核酸酶保护试验来检测Eph受体酪氨酸激酶的表达，Eph受体酪氨酸激酶是胚胎发育中肌动蛋白动力学的关键调节因子。我们发现，在IRI后，肾脏中一种特别的受体EphA2强烈上调。Eph受体的细胞结合配体Ephrin也有强烈表达。在培养的肾小管细胞中，模拟IRI的各种损伤性刺激也导致EphA2蛋白表达上调。EphA2的上调被Src激酶抑制剂PP2抑制。相反，Src激酶的过表达强烈增强了内源性EphA2的表达以及人EphA2启动子构建体的活性。Erk途径的激活对于Src激酶完全诱导EphA2上调是必要的，但并不充分。用EphA2配体ephrin-A1刺激肾小管上皮细胞导致内源性EphA2、桩蛋白和一种未鉴定的约65 kDa蛋白的酪氨酸磷酸化，并导致皮质F-肌动蛋白染色增加。总之，在模拟IRI的体外条件下，EphA2的表达通过Src激酶依赖性途径强烈上调。上调的EphA2与其Ephrin配体之间的相互作用可能为肾IRI中的细胞骨架修复提供关键的细胞接触依赖性双向信号。
