Molecular mapping of microbial communities at the host-pathogen interface by multi-modal 3-dimensional imaging mass spectrometry

项目概要 细胞与环境的相互作用构成了所有生物体健康和疾病的基础。曝光 对营养物质、毒素和邻近细胞的影响会触发协调的分子反应，从而影响细胞功能和 以有益的、适应性的或有害的方式进行新陈代谢。尽管多细胞性的好处 复杂组织结构的形成或整个器官系统的功能长期以来一直受到人们的重视， 直到最近人们才了解到，脊椎动物的微生物居民也对宿主产生巨大的影响 细胞功能和功能障碍。尽管如此，对这些相互作用的理解并没有超出简单的范围 关联，并且几乎没有可用的分子技术能够充分定义复杂的 微生物生态系统影响宿主细胞功能，或者宿主对微生物定植的反应如何影响 细菌群落。当人们考虑到以下问题所产生的广泛而重大的影响时，这种知识差距是惊人的： 微生物对人体健康有影响。在此应用中，我们建议通过以下方式明确填补这一知识空白： 开发一种新型多模态成像管道，该管道将提供分子的 3 维信息 微生物群落的异质性和宿主-病原体界面的免疫反应。 该提案结合了我们在免疫学、感染生物学、质谱、小动物方面的专业知识 成像、机器学习和计算机视觉，开发集成的多模态可视化方法 研究传染病。我们独特的方法将在计算上结合超高速（~50px/s） MALDI-TOF 图像，超高质量分辨率（>200,000 分辨率） MALDI FTICR IMS，金属成像 通过 LA-ICP-IMS、高空间分辨率光学显微镜和使用数据驱动图像融合的 MR 成像。这个 该策略将使 3D 分子图像能够生成数千种元素、代谢物、脂质和 蛋白质前所未有地结合了化学特异性和空间保真度，速度快于 50 倍以上 目前是可能的。我们将使用这种下一代成像功能来（i）定义异质微生物 金黄色葡萄球菌群落整个 3D 体积的亚群，(ii) 揭示形成的宿主分子 脓肿和积聚以限制小鼠模型中的微生物生长，以及（iii）阐明分子标记 区分不同阶段脓肿之间宿主-病原体界面的体内生物膜 进展，并在不同程度的营养压力下。这些研究将揭示新的治疗靶点 由于该提案而开发的干预措施和技术将广泛适用于所有 生理相关过程，深刻影响生物医学研究。

PROJECT SUMMARY Cellular interactions with the environment form the basis of health and disease for all organisms. Exposure to nutrients, toxins, and neighboring cells trigger coordinated molecular responses that impact cell function and metabolism in a beneficial, adaptive, or detrimental manner. Although the benefits of multicellularity for the formation of complex tissue structures or the function of entire organ systems has been long appreciated, it has only recently been understood that microbial inhabitants of vertebrates also have a tremendous impact on host cell function and dysfunction. Despite this, an understanding of these interactions has not moved beyond simple associations, and there are virtually no molecular technologies available that adequately define how a complex microbial ecosystem impacts host cell function, or how the host response to microbial colonization affects the bacterial community. This gap in knowledge is striking when one considers the broad and significant impact that microbes have on human health. In this application, we propose to expressly fill this knowledge gap through development of a novel multimodal imaging pipeline that will provide 3-dimensional information on the molecular heterogeneity of microbial communities and the immune response at the host-pathogen interface. This proposal combines our expertise in immunology, infection biology, mass spectrometry, small animal imaging, machine learning, and computer vision to develop an integrated multimodal visualization method for studying infectious disease. Our unique approach will computationally combine ultra-high speed (~50px/s) MALDI-TOF images, ultra-high mass resolution (>200,000 resolving power) MALDI FTICR IMS, metal imaging by LA-ICP-IMS, high-spatial resolution optical microscopy, and MR imaging using data-driven image fusion. This strategy will enable 3-D molecular images to be generated for thousands of elements, metabolites, lipids, and proteins with an unprecedented combination of chemical specificity and spatial fidelity more than 50x faster than is currently possible. We will use this next-generation imaging capability to (i) define the heterogeneous microbial subpopulations throughout the 3-D volume of a S. aureus community, (ii) uncover the host molecules that form the abscess and accumulate to restrict microbial growth in murine models, and (iii) elucidate molecular markers that differentiate in vivo biofilms at the host-pathogen interface, between abscesses at various stages of progression, and under distinct degrees of nutrient stress. These studies will uncover new targets for therapeutic intervention and the techniques developed as a result of this proposal will be broadly applicable to all physiologically relevant processes, profoundly impacting biomedical research.