从胰岛素信号转导通路探讨BPA促进动脉粥样硬化的作用机制

Bisphenol A (BPA) is one of environmental endocrine disrupting chemicals that is present ubiquitously in daily life. Although many epidemiologic studies have established the association between BPA exposure and the development of atherosclerotic cardiovascular disease, the underlying mechanisms clarified by in vitro studies or in vivo experimental work in animals are limited. As we known, normal insulin signaling in classical insulin target organs such as liver, muscle, and adipose are important for maintaining the whole body glucose homeostasis. And there are increasing studies suggest that defective insulin signaling in cells of the arterial wall including macrophages may promote the development of atherosclerosis. In previous study, exposure to BPA has been shown to induce insulin resistance by disrupting insulin signaling in peripheral tissues in rat or mice model. Because that the rabbit is considered more suitable animal model for studying cardiovascular disease, the present study selected a Watanabe heritable hyperlipidemic (WHHL) rabbit model, which is deficient for LDL receptor function, to test whether exposure to BPA was able to induce or/and promote the development of atherosclerosis, and if so, whether these adverse effects were mediated by impaired regulation of insulin signaling. We firstly tested the effects of exposure to BPA on glucose homeostasis and atherosclerotic lesions in the WHHL rabbits. Then, effects of BPA on components/modulators of the insulin signaling pathway were screened and identified in both classic insulin-sensitive organs and arterial vascular cell (including endohtelial, smooth muscle and macrophage foam cells) by performing modern molecular biology experiments. After that, cell culture and intervention experiments were performed in vitro to ensure the possibility that exposure to BPA worsened atherosclerosis by selectively inhibiting insulin signaling. Taken together, this study is expected to address the mechanism of BPA-induced atherosclerosis at molecular levels, furthermore, create new insights into the role of environmental factors on the onset of atherosclerosis.

双酚A（BPA）是典型的环境内分泌干扰物。大量流行病学调查证实BPA暴露与动脉粥样硬化（AS）的发生存在显著正相关，但对于危害现象的分子毒理机制报道较少。胰岛素抵抗可导致动脉血管胰岛素信号受损，进而破坏血管壁舒缩功能、促进血管平滑肌细胞增殖、诱导单核-巨噬细胞凋亡，在AS的发生发展过程中发挥重要作用。现已证实BPA可通过抑制胰岛素信号诱发机体胰岛素抵抗，所以本项目拟以渡边可遗传高脂血症（WHHL）家兔为研究对象，确定BPA暴露引发WHHL家兔胰岛素抵抗和AS的风险和量效关系；阐明BPA暴露致AS过程中受胰岛素信号通路调控的关键靶基因及其信号网络；利用离体培养的血管内皮细胞、平滑肌细胞及巨噬细胞模型，比较过表达或抑制筛选靶基因前后BPA对血管细胞脂肪病变、炎症、增殖、凋亡等的影响，确证BPA通过干扰胰岛素信号转导诱发AS的分子机制。项目有望为环境因素致AS的发病机制研究提供新的思路与线索。

双酚A（BPA）是一类在环境中广泛分布的环境内分泌干扰物质，它在人体血液和尿液中含量的升高和多种心血管疾病存在关联，但目前仍缺乏实验室研究来验证上述相关性并阐明潜在的分子毒理学机制。本项目以WHHL家兔为模型，研究BPA暴露对动脉粥样硬化形成和发展的影响。发现BPA可被WHHL家兔快速吸收和代谢。低浓度暴露12周促进了WHHL家兔动脉粥样硬化的形成。高浓度BPA暴露8周没有显著促进主动脉各部位总体病变面积增加，但促使主动脉病变区域的巨噬细胞数量和后期病变面积显著增加，说明BPA对动脉粥样硬化的影响和暴露剂量及时间相关。结合细胞实验揭示BPA主要作用于血管内皮细胞，通过诱导炎症反应和内质网应激等作用导致内皮细胞的功能受损，从而间接诱导平滑肌细胞迁移和增殖。同时，BPA还可直接作用于心肌细胞和动脉壁，通过促进炎症反应、心肌细胞缺血坏死及钙化斑块增多等现象加剧动脉粥样硬化病变程度。. 进一步研究BPA暴露对WHHL家兔脂类和糖类代谢的影响，发现高浓度BPA暴露造成WHHL家兔出现胰岛素抵抗，血糖代谢减缓。BPA暴露导致皮下和腹部脂肪细胞肥大，肝细胞和心肌细胞中出现脂质蓄积、炎症细胞入侵及细胞凋亡等现象，若干和内质网应激、脂类代谢及炎症反应密切相关基因在肝脏中显著上调表达。体外细胞实验发现BPA同样显著诱导了人体肝癌细胞（HepG2）中与内质网应激、炎症反应、脂类代谢及污染物代谢相关基因显著上调表达，脂滴蓄积和甘油三酯含量升高，表明BPA可通过调节上述信号通路对肝脏的脂类代谢及正常结构功能造成损伤。通过小鼠及体外细胞实验揭示BPA通过抑制miR-192导致其直接作用的靶基因——肝细胞脂质稳态关键转录因子SREBF1上调，从而引起肝细胞脂质稳态的调节异常。. 综上所述，BPA一方面可直接作用于动脉壁和心肌细胞，另一方面可通过诱发多重代谢紊乱，从而直接或间接促进动脉粥样硬化的形成和发展。