Immunogenetics Center, Department of Pathology and Laboratory Medicine, David Geffen School of Medicine, University of California, Los Angeles, 90095, USA.
Am J Physiol Cell Physiol. 2012 Oct 15;303(8):C872-82. doi: 10.1152/ajpcell.00199.2012. Epub 2012 Aug 22.
Following transplantation, HLA class I antibodies targeting donor endothelium stimulate cell proliferation and migration, which contribute to the development of transplant vasculopathy and chronic allograft rejection. Dynamic remodeling of the actin cytoskeleton regulates cell proliferation and migration in endothelial cells (ECs), but the mechanism(s) involved remain incompletely understood. We explored anti-HLA class I antibody-mediated alterations of the cytoskeleton in human aortic ECs (HAECs) and contrasted these findings to thrombin-induced cytoskeleton remodeling. Our results identify two different signaling pathways leading to myosin light chain (MLC) phosphorylation in HAECs. Stimulation of HAECs with thrombin at 1 U/ml induced a robust elevation of intracellular Ca(2+) concentration, increased MLC phosphorylation, and promoted stress fiber formation via MLC kinase (MLCK) and Rho kinase (ROK) in an ERK-independent manner. In contrast, HAECs stimulated with HLA class I antibodies did not promote any detectable change in intracellular Ca(2+) concentration but instead induced MLC phosphorylation and stress fiber assembly via MLCK and ROK in an ERK1/2-dependent manner. Stimulation of HAECs with low-dose thrombin (1 mU/ml) induced signaling cascades that were similar to stimulation with HLA class I antibodies. HLA class I antibodies also stimulated the translocation of mammalian target of rapamycin complex 2 (mTORC2) and ERK1/2 from the cytoplasm to the plasma membrane independently of stress fiber assembly. These findings identify novel roles for HLA class I signaling in ECs and provide new insights into the role of ERK1/2 and mTORC2 in cytoskeleton regulation, which may be important in promoting transplant vasculopathy, tumor angiogenesis, and atherosclerosis.
移植后，针对供体内皮细胞的 HLA Ⅰ类抗体刺激细胞增殖和迁移，这有助于移植血管病和慢性移植物排斥的发展。肌动蛋白细胞骨架的动态重塑调节内皮细胞（ECs）中的细胞增殖和迁移，但涉及的机制仍不完全清楚。我们探讨了抗 HLA Ⅰ类抗体介导的人主动脉 ECs（HAECs）细胞骨架的改变，并将这些发现与凝血酶诱导的细胞骨架重塑进行了对比。我们的结果确定了两种不同的信号通路，导致 HAECs 中的肌球蛋白轻链（MLC）磷酸化。在 1 U/ml 的凝血酶刺激下，HAECs 诱导细胞内 Ca(2+)浓度的强烈升高，通过 MLCK 和 ROK 增加 MLC 磷酸化，并以 ERK 非依赖性的方式促进应力纤维形成。相比之下，用 HLA Ⅰ类抗体刺激的 HAECs 不会促进任何可检测到的细胞内 Ca(2+)浓度变化，但通过 ERK1/2 依赖性方式诱导 MLC 磷酸化和应力纤维组装。用低剂量凝血酶（1 mU/ml）刺激 HAECs 诱导的信号级联与用 HLA Ⅰ类抗体刺激相似。HLA Ⅰ类抗体还刺激哺乳动物雷帕霉素靶蛋白复合物 2（mTORC2）和 ERK1/2 从细胞质向质膜的易位，而不依赖于应力纤维组装。这些发现确定了 HLA Ⅰ类信号在 ECs 中的新作用，并提供了关于 ERK1/2 和 mTORC2 在细胞骨架调节中的作用的新见解，这可能对促进移植血管病、肿瘤血管生成和动脉粥样硬化很重要。
