【作者】 刘颖；

【导师】 许安；

【作者基本信息】 中国科学技术大学 ， 生物物理学， 2021， 博士

【摘要】 微生物组是维持机体内环境稳定的重要因素之一,与宿主代谢和免疫功能息息相关,紊乱的微生物组与环境污染暴露损伤及多种疾病的发生发展紧密关联。在当前,大气细颗粒物(PM2.5)污染已成为制约社会和谐发展的公共卫生问题。然而,PM2.5暴露对机体微生物组的影响尚不明确,微生物组在PM2.5暴露导致的损伤及疾病中的作用有待确证。本研究聚焦机体微生物组,分析研究PM2.5暴露对肺部菌群和肠道菌群的影响;通过菌群移植,探索肺部菌群在PM2.5导致的肺损伤和影响肺癌发生进展中的作用及机制。1、PM2.5对小鼠肺部菌群和肠道菌群及肺组织代谢组的影响。通过气管滴注PM2.5构建亚急性小鼠暴露模型,分析暴露组小鼠微生物组改变和机体损伤。结果表明:1)亚急性PM2.5暴露导致小鼠肺部菌群和肠道菌群菌群结构和组成显著改变。单位质量小鼠肺组织和肠道排泄物菌群总量显著增加,但菌群多样性降低,主要菌群种类比例变化明显,益生菌乳酸杆菌和双歧杆菌相对丰度显著下降,条件致病菌志贺菌属和具核梭杆菌属比例显著上升。2)亚急性PM2.5暴露导致小鼠肺组织代谢组变化明显,并与显著变化的肺部菌群存在关联作用。3)亚急性PM2.5暴露导致遗传毒性显著增强。亚急性PM2.5暴露后小鼠股骨骨髓嗜多染红细胞(PCE)的微核率显著增加;肺组织Tunel实验和γ-H2AX分析表明,亚急性PM2.5暴露导致小鼠显著的肺细胞基因组不稳定性。2、肺部菌群和肠道菌群在PM2.5导致机体损伤中的作用和机制。通过菌群移植改变PM2.5暴露组小鼠肺部菌群和肠道菌群,结果显示:移植正常小鼠肠道/肺部菌群后,PM2.5暴露组小鼠骨髓嗜多染红细胞微核率未明显降低,系统性炎症未明显改善;然而,肺部菌群移植显著改善PM2.5暴露导致的肺组织炎症、细胞凋亡和DNA损伤,但肠道菌群移植对改善PM2.5暴露所致肺损伤作用不明显。乳酸杆菌、具核梭杆菌和志贺菌属移植亦可显著改变PM2.5暴露引起的肺组织炎症、纤维化和DNA损伤。肺部菌群影响PM2.5暴露导致的肺损伤可能与cGAS/STING/TLR9信号通路激活有关。3、肺部菌群在PM2.5促进肺癌发生进展中的作用和机制。1)通过对比分析肺癌患者和对照人群肺部菌群差异,发现肺癌患者与对照病人肺部菌群显著不同,具核梭杆菌和志贺菌属相对丰度显著增加。2)通过菌群移植发现,肺癌患者肺部菌群会加速肺癌模型小鼠肺癌进展,肺癌细胞增多,肺癌进展指标Ki67和PCNA表达显著上调。3)为明确肺部菌群在PM2.5暴露加速肺癌发生进展中的作用,利用抗生素清除PM2.5暴露组小鼠肺部菌群,和/或移植肺癌患者肺部菌群和健康小鼠肺部菌群,观察肺癌模型小鼠肺癌进展变化。结果显示:使用抗生素清除PM2.5暴露组肺部菌群,可显著减轻肺癌模型小鼠肺癌进展以及肺组织炎症和DNA损伤;移植肺癌患者肺部菌群,显著加重肺癌模型小鼠肺癌进展以及肺组织炎症和DNA损伤;单一菌种(如乳酸杆菌、具核梭杆菌或志贺菌属)移植,亦可显著改变PM2.5暴露引起的肺癌进展与肺组织炎症和DNA损伤变化。综上所述,本研究观察分析PM2.5暴露所导致的小鼠肺部菌群/肠道菌群改变,并通过菌群移植确证肺部菌群和肠道菌群在PM2.5暴露所致小鼠肺损伤、纤维化和基因组不稳定性中的作用,为解析PM2.5暴露的毒性效应和机制提供了新的视角。另一方面,本研究分析肺部菌群与肺癌相关性,通过菌群移植确证肺部菌群与肺癌发生进展的因果关系,鉴定数种肺癌指示菌;分析确证肺部菌群在PM2.5促癌效应中的作用,并从肺部菌群引起的炎症和DNA损伤角度解析其中机制。本课题从微生物组视角解析PM2.5毒性作用和机制,并为肺癌预防、诊断和治疗提供潜在新的干预靶点。更多还原

【Abstract】 Microbiome is one of the important factors for the stability of the host’s internal environment,which is closely associated with metabolism and immunity of the host.However,microbiome dysbiosis is closely related to injury induced by environmental exposure and occurrence/development of several diseases.Nowadays,fine particulate matter(PM2.5)pollution has been a public health issue,which restrained social development.The influence of PM2.5 exposure on microbiome and the role of microbiome in injury and diseases caused by PM2.5 are still unclear.Current study aimed to explore the effects of PM2.5 exposure on the microbiome(lung microbiota and gut microbiota),the role of microbiome in injury and diseases caused by PM2.5,the association between lung microbiota and lung cancer and the role of lung microbiota in lung cancer induced by PM2.5.1.The studies of PM2.5 exposure on lung/gut microbiota and lung metabonomics showed that the subacute PM2.5 exposure caused dysbiosis of lung microbiota and gut microbiota,namely increase of the total amount of microbiota in unit mass of lung/feces,reduction of alpha diversity,significant changes of microbiotal composition and structure,reduction of probiotics and augment of pathogens.Subacute PM2.5 exposure altered the metabonomics of lung tissue,which were associated with significantly changed lung microbiota.The micronucleus rate of polychromatic erythrocytes(PCE)in bone marrow significantly augmented after exposure to PM2.5,suggesting the genomic instability of mice.Tunel experiment of lung tissue and γ-H2AX analysis showed that PM2.5 exposure resulted in significant DNA damage and genomic instability of the lung tissues in mice.2.To investigate the role of lung and gut microbiota in lung damage and genomic instability caused by PM2.5 exposure,we altered the lung and gut microbiota of mice in the exposure group by microbiota transplantation.The results showed that after transplantation of lung/gut microbiota of normal mice,the micronucleus rate of PCE in bone marrow of mice exposed to PM2.5 was not significantly reduced,and systemic inflammation was not significantly improved.Lung microbiota transplantation significantly alleviaviated lung inflammation,cell apoptosis and DNA damage caused by PM2.5 exposure,while gut microbiota transplantation had no significant effect on lung injury caused by PM2.5 exposure.Transplantation of Lactobacillus,Fusobacterium and Shigella also significantly altered lung inflammation,fibrosis and DNA damage caused by PM2.5 exposure.The influence of lung microbiota transplantation on lung damage induced by PM2.5 exposure may be related to the activation of cGAS/STING/TLR9 signaling pathway.3.In order to investigate the correlation between lung microbiota and lung cancer,we conducted a case-control study to analyze the differences of lung microbiota between health controls and lung cancer patients.The data demonstrated an obviously difference in the lung microbiota among lung cancer patients and controls,and the relative abundances of Fusobacterium and Shigella were significantly increased in lung cancer patients.Furthermore,the causal relationship between lung microbiota and lung cancer was confirmed by microbiota transplantation.The results showed that lung microbiota of lung cancer patients accelerated the progression of lung cancer in lung cancer model mice,increased lung cancer cells,and significantly upregulated the expression of lung cancer progression indicators Ki67 and PCNA.To further confirm the role of lung microbiota in accelerating the progression of lung cancer caused by PM2.5 exposure,we used antibiotics to clear the lung microbiota of mice in PM2.5 exposure group,and/or transplanted the lung microbiota of lung cancer patients and controls.The results showed that elimination of lung microbiota in PM2.5 exposed mice with antibiotics could significantly reduce lung cancer progression and inflammation in lung cancer model mice and transplantation of lung microbiota from lung cancer patients significantly increased lung cancer progression,lung tissue inflammation and DNA damage in lung cancer model mice.Transplantation of Lactobacillus,Fusobacterium and Shigella could also significantly alter the progression of lung cancer,lung inflammation and DNA damage caused by PM2.5 exposure.In this study,we observed and analyzed changes in lung/gut microbiota of mice caused by exposure to subacute PM2.5,and confirmed the role of lung/gut microbiota in lung injury,fibrosis and DNA damage caused by exposure to PM2.5 through microbia transplantation.On the other hand,this study analyzed the correlation between lung microbiota and lung cancer,confirmed the causal relationship between lung microbiota and the development of lung cancer through microbiota transplantation,and identified several bacterial indicators of lung cancer.We also analyzed and confirmed the role of lung microbiota in the cancer-promoting effect of PM2.5 via the inflammation and DNA damage caused by lung microbiota.This study analyzed the toxicity of PM2.5 from the perspective of microbiome,and provided a potential target for the diagnosis,prevention and treatment of lung cancer.更多还原