Abstract

The study aims to explore the fluctuating expression of C/EBP Homologous Protein (CHOP) following rat carotid artery injury and its central role in vascular stenosis. Using in vivo rat carotid artery injury models and in vitro ischemia and hypoxia cell models employing human aortic endothelial cells (HAECs) and vascular smooth muscle cells (T/G HA-VSMCs), a comprehensive investigative framework was established. Histological analysis confirmed intimal hyperplasia in rat models. CHOP expression in vascular tissues was assessed using Western blot and immunohistochemical staining, and its presence in HAECs and T/G HA-VSMCs was determined through RT-PCR and Western blot. The study evaluated HAEC apoptosis, inflammatory cytokine secretion, cell proliferation, and T/G HA-VSMCs migration through Western blot, ELISA, CCK8, and Transwell migration assays. The rat carotid artery injury model revealed substantial fibrous plaque formation and vascular stenosis, resulting in an increased intimal area and plaque-to-lumen area ratio. Notably, CHOP is markedly elevated in vessels of the carotid artery injury model compared to normal vessels. Atorvastatin effectively mitigated vascular stenosis and suppresses CHOP protein expression. In HAECs, ischemia and hypoxia-induced CHOP upregulation, along with heightened TNFα, IL-6, caspase3, and caspase8 levels, while reducing cell proliferation. Atorvastatin demonstrated a dose-dependent suppression of CHOP expression in HAECs. Downregulation of CHOP or atorvastatin treatment led to reduced IL-6 and TNFα secretion, coupled with augmented cell proliferation. Similarly, ischemia and hypoxia conditions increased CHOP expression in T/G HA-VSMCs, which was concentration-dependently inhibited by atorvastatin. Furthermore, significantly increased MMP-9 and MMP-2 concentrations in the cell culture supernatant correlated with enhanced T/G HA-VSMCs migration. However, interventions targeting CHOP downregulation and atorvastatin usage curtailed MMP-9 and MMP-2 secretion and suppressed cell migration. In conclusion, CHOP plays a crucial role in endothelial injury, proliferation, and VSMCs migration during carotid artery injury, serving as a pivotal regulator in post-injury fibrous plaque formation and vascular remodeling. Statins emerge as protectors of endothelial cells, restraining VSMCs migration by modulating CHOP expression.

中文翻译：

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C/EBP同源蛋白在颈动脉损伤后血管狭窄中的作用

摘要

本研究旨在探讨大鼠颈动脉损伤后C/EBP同源蛋白（CHOP）的表达波动及其在血管狭窄中的核心作用。利用体内大鼠颈动脉损伤模型以及采用人主动脉内皮细胞（HAEC）和血管平滑肌细胞（T/G HA-VSMC）的体外缺血和缺氧细胞模型，建立了一个全面的研究框架。组织学分析证实了大鼠模型中的内膜增生。使用蛋白质印迹和免疫组织化学染色评估血管组织中的 CHOP 表达，并通过 RT-PCR 和蛋白质印迹确定其在 HAEC 和 T/G HA-VSMC 中的存在。该研究通过蛋白质印迹、ELISA、CCK8 和 Transwell 迁移测定评估了 HAEC 凋亡、炎性细胞因子分泌、细胞增殖和 T/G HA-VSMC 迁移。大鼠颈动脉损伤模型显示大量纤维斑块形成和血管狭窄，导致内膜面积和斑块与管腔面积比增加。值得注意的是，与正常血管相比，颈动脉损伤模型的血管中 CHOP 显着升高。阿托伐他汀有效减轻血管狭窄并抑制 CHOP 蛋白表达。在 HAEC 中，缺血和缺氧诱导 CHOP 上调，同时 TNFα、IL-6、caspase3 和 caspase8 水平升高，同时减少细胞增殖。阿托伐他汀对 HAEC 中 CHOP 表达具有剂量依赖性抑制作用。 CHOP 或阿托伐他汀治疗的下调导致 IL-6 和 TNFα 分泌减少，同时细胞增殖增强。类似地，缺血和缺氧条件增加了 T/G HA-VSMC 中的 CHOP 表达，而阿托伐他汀浓度依赖性地抑制了该表达。此外，细胞培养上清液中 MMP-9 和 MMP-2 浓度的显着增加与 T/G HA-VSMC 迁移的增强相关。然而，针对 CHOP 下调和阿托伐他汀使用的干预措施减少了 MMP-9 和 MMP-2 的分泌并抑制了细胞迁移。总之，CHOP 在颈动脉损伤过程中的内皮损伤、增殖和 VSMC 迁移中发挥着至关重要的作用，是损伤后纤维斑块形成和血管重塑的关键调节因子。他汀类药物作为内皮细胞的保护剂，通过调节 CHOP 表达来抑制 VSMC 迁移。