mTOR通路在交通相关PM2.5短期暴露致心肌细胞损伤中的作用

Between 350,000 and 500,000 Chinese died prematurely each year as a result of outdoor air pollution in China. With the evaluation of the contribution of traffic-related PM2.5 to total PM2.5 has become the focus of interest, the cardiotoxic effects of the traffic-related PM2.5 short-term exposure is also censored. The toxicologic studies showed that PM2.5 short-term exposure could induce oxidative stress, which was link to cardiomyocyte injury via direct and indirect pathways. So far, the mechanism of the cardiomyocyte injury caused by traffic-related PM2.5 short-term exposure has been still unclear. Previous study showed that the genomic changes of cardiomyocyte induced by PM2.5 exposure might be associated with Akt/mTOR pathway. Moreover, it is clear that cardiomyocyte apoptosis and autophagy caused by oxidative stress were associated with mTOR signaling pathway. Therefore, in the present study, Fourier transform infrared spectroscopy, X-ray power diffraction, and Inductively coupled plasma mass spectrometry were applied to analyze the contribution of traffic-related PM2.5 to total PM2.5 at different areas and different seasons, then the traffic-related PM2.5 short-term exposure models with different doses and exposure time will be established, and then pathology, molecular biology, untrosonic cardiogram, FTIR, and Raman spectroscopy were applied to assess the cardiotoxic effects via mTOR signaling pathway, such as cardiomyocyte morphology, survival, molecular components, cardiac function, and so on. It will be useful to illuminate the molecular mechanism of cardiomyocyte injury caused by traffic-related PM2.5 short-term exposure, and it will provide a novel idea in prevention and treatment.

空气污染每年导致35-50万中国人过早死亡。随着交通相关PM2.5的贡献成为热点，其短期暴露致心血管损伤也备受关注。毒理学研究显示，PM2.5暴露引起氧化应激，通过直接、间接途径致心肌细胞损伤，但交通相关PM2.5短期暴露的心肌细胞损伤机制仍处于探索阶段。研究提示PM2.5暴露致心肌基因变化可能与Akt/mTOR信号通路相关，且氧化应激损伤中mTOR信号通路与心肌细胞凋亡、自噬等密切相关。因此，本研究拟应用傅里叶变换红外光谱（FTIR）及电感耦合等离子体质谱技术解析不同地区、季节交通相关PM2.5的贡献，并建立不同剂量、暴露时间的交通相关PM2.5短期暴露模型，应用病理学、分子生物学、超声、FTIR及拉曼光谱技术研究心脏mTOR信号通路介导的心肌细胞形态、活性、分子构成及心功能改变等，阐明交通相关PM2.5短期暴露致心肌细胞氧化应激性损伤的分子机制，为预防及治疗提供新思路。

交通相关PM2.5在城市PM2.5的贡献约占17%，而且，交通相关PM2.5中含有大量的致癌物质，因此，交通相关PM2.5对机体的毒性作用近年来备受关注。流行病学调查研究显示PM2.5与心血管疾病的发生及死亡相关，而毒理学研究提示交通相关PM2.5短期暴露可引起机体的氧化应激作用，因此推测氧化应激可能是交通相关PM2.5短期暴露的心肌细胞毒性损伤的机制之一。. PM2.5可以定植在肺泡中，其中PM0.1可以进入血液循环对心肌细胞起毒性作用，因此，本实验主要应用新生乳鼠原代心肌细胞及H9C2心肌细胞株建立交通相关PM0.1短期暴露的模型，暴露时间为24h内，剂量为10μg/ml-200μg/ml，应用分子生物学技术及傅里叶变换红外光谱技术技术分析心肌细胞的蛋白质、脂质及DNA等大分子的含量及构型变化，细胞坏死、凋亡、自噬的变化，以及氧化应激水平的变化。应用昆明小鼠尾静脉注射PM0.1混悬液建立动物模型，应用红外光谱技术检测血清样本的分子特征及氧化应激水平。. 结果显示，暴露2h即可以引起心肌细胞坏死，并呈现出剂量-反应关系； 200μg/ml水平暴露，细胞坏死呈时间-反应关系。暴露24h，心肌细胞蛋白质、脂质、DNA氧化应激水平呈剂量-反应关系；流式细胞术提示随着暴露时间增加，心肌细胞从早期凋亡向晚期凋亡及坏死转变；细胞凋亡、自噬相关蛋白表达均增加，同时mTOR通路蛋白表达也呈剂量-反应关系；红外光谱提示氧化应激水平呈剂量-反应关系，与分子生物学结果一致，蛋白质、脂质及DNA的分子含量和构型也存在差异。小鼠血清红外光谱提示暴露24h机体的氧化应激作用及分子特征也基本呈现剂量-反应关系。. 交通相关PM2.5短期暴露引起氧化应激，通过调控mTOR信号通路引起细胞坏死、凋亡及自噬等造成心肌细胞损伤，提示健康成年人，尤其是高暴露人群有防护的必要性，也为进一步预防、治疗的靶点的选择奠定理论基础。