邻苯二甲酸二丁酯和苯并(a)芘致肝损伤表观调控机制及其干预研究

Dibutyl phthalate and Benzo(a)pyrene , as the most common organic pollutants, have been listed as the first "environmental problems" by many countries. However, their pathogenic and carcinogenic mechanisms are still unknown, which is resulting in that the long-term health hazards aren't completely controlled. Governments and scholars are paying high attention to such problems. Inflammatory reactions and immune regulation are thought as starting factors for many diseases including liver injury. Recent studies suggest that epigenetic regulation plays an important role in immune inflammation. Therefore, this study is based on the previous research to investigate the current situation of organic pollution and liver toxicity in Guiyang city. Dibutyl phthalate and Benzo(a)pyrene are selected as our research object and fluorescence quantitative PCR, immunoprecipitation, immunoblotting and quantitative mass spectrometry technique are used. From the perspective of epigenetics (DNA methylation, histone modification and miRNAs), the molecular mechanism of dibutyl phthalate and benzo (a) pyrene-induced inflammatory response and immunosuppression in hepatocyte injury and even malignant transformation is explored systematically through cell and animal experiments. The current study aims to further the molecular target of drug toxicity, analyze their relationship with hepatocyte injury and even malignant transformation, and obtain a clear and effective comprehensive control strategy.

邻苯二甲酸二丁酯和苯并(a)芘作为最多见的有机污染物，已被很多国家列为“环境问题”之首，其致病致癌机制不明是导致其远期健康危害未得以彻底控制的瓶颈问题，备受各国政府和学者高度关注。炎症反应、免疫调控是包括肝损伤在内的多种疾病的始动因素已被众多学者认可，但机制尚不明确。新近研究推测，表观遗传学调控在免疫炎症中起重要作用。本项目针对本地区有机物污染及其肝毒性现状，在课题组前期研究基础上，以邻苯二甲酸二丁酯和苯并(a)芘为研究对象，采用荧光定量PCR、免疫共沉淀、免疫印迹、定量质谱等方法，从表观遗传角度（DNA甲基化、组蛋白修饰和miRNAs），通过细胞、动物二个层次较深入系统地探讨炎症反应、免疫抑制在邻苯二甲酸二丁酯及苯并(a)芘致肝细胞损伤及恶性转化的分子机制；寻找毒作用分子靶点，分析其与肝细胞损伤乃至恶性转化的关系；并进行针对性干预，以获得靶向明确、实效明显的综合防治策略。

邻苯二甲酸二丁酯(DBP)和苯并(a)芘(BaP)作为最多见的有机污染物，已被很多国家列为“环境问题”之首，其致病机制不明是导致其远期健康危害未得以彻底控制的瓶颈问题，备受各国政府和学者高度关注。本研究从表观遗传角度（DNA甲基化、组蛋白修饰和miRNAs），通过细胞、动物二个层次较深入系统地探讨炎症反应在DBP及BaP致肝细胞损伤的分子机制，寻找毒作用分子靶点，分析其与肝细胞损伤的关系；并进行针对性干预，以获得靶向明确、实效明显的综合防治策略。研究结果显示，（1）DBP、BaP亚慢性染毒可使尿液中主要代谢产物物MBP与1-OHP水平增加，大鼠肝功能异常，肝脏组织发生病理改变，炎症因子分泌紊乱，肝脏出现炎性损伤；（2）DBP、BaP亚慢性染毒可下调TLR4基因启动子区甲基化水平，诱导 TLR4/CD14/MD-2受体复合物表达水平升高，进而介导下游TRAF6/STAT1信号通路表达异常，启动炎症反应；（3）DBP、BaP亚慢性染毒通过诱导大鼠肝组织内TLR4的高乙酰化，进而激活TLR4/NF-κB信号通路，促进炎症因子的释放，介导肝脏炎性损伤的发生；（4）DBP、BaP亚慢性染毒通过下调miR-34a-5p的表达激活Notch信号，促使巨噬细胞M1/M2极化的不平衡，进而导致炎症因子分泌紊乱和肝损伤的发生；（5）破壁灵芝孢子粉（SSGL）分别通过回复TLR4基因启动子区低甲基化、降低TLR4乙酰化水平、增加miR-34a-5p的表达水平，抑制相应信号通路TLR4/CD14/MD-2受体复合物及其介导的下游TRAF6/STAT1信号转导通路、TLR4/NF-κB信号通路、Notch信号通路的激活，进而减少炎症因子的分泌，减缓DBP、BaP所致的大鼠肝损伤。（6）细胞实验表明BaP、 DBP暴露可降低L02细胞存活率，引起细胞形态改变，将细胞周期阻滞在S期， 诱导肝细胞凋亡；同时促进炎症因子的释放，导致肝细胞炎性损伤。（7）BaP、DBP暴露可下调L02细胞中miR-122-5p的表达，进而促进炎症因子的释放，介导肝细胞损伤；而上调 miR-122-5p可缓解肝细胞损伤。（8）miR-122-5p通过靶向SOCS1调控IL6/STAT3-Mcl-1轴，进而促进炎症反应是BaP、DBP诱导肝细胞损伤的主要机制之一。可为防治本地区POPs污染带来的健康危害提供科学依据。