Thursday, May 9, 2024
5/9/2024
Project Summary Human tuberculosis (TB) granulomas are complex three-dimensional (3D) lung tissue lesions that form in response to infection by Mycobacterium tuberculosis (Mtb). TB granulomas are comprised of multiple cell types and are the critical site of host-pathogen interactions that determine disease outcome. The host-pathogen interaction in TB is complex and different granulomas can either progress or regress in the same individual concurrently, demonstrating that local granuloma factors determine progression versus protection. Granulomas expand and organize by the recruitment of additional cells including: monocytes recruited from the blood that differentiate into macrophages and dendritic cells, stromal fibroblasts which deposit extracellular matrix (ECM) and secreted factors that influence host cell survival and Mtb growth, and T and B lymphocytes that localize to the peripheral regions of the granuloma. Despite the importance of these events, the signals that mediate cell recruitment, aggregation, and function within human TB granulomas are poorly understood, largely due to the limitations of current experimental models of human TB pathogenesis. To address this need, we have developed tissue-engineered, self-assembling 3D biomimetic human TB granulomas (BHTGs) from human peripheral blood mononuclear cells (PBMCs). BHTGs recapitulate key architectural elements of human TB granulomas: a central core of mononuclear phagocytes containing live mycobacteria that grows via cell-to-cell spread, progressive tissue growth and reorganization through the recruitment of monocytes, and 3D cell organization including internal localization of monocytes and macrophages and the peripheral localization of lymphocytes. In this proposal, we seek to more fully recapitulate the cellular interactions in TB granulomas and will apply our model as a biological discovery platform to contribute unprecedented mechanistic and behavioral insight into the factors governing TB granuloma progression through two Specific Aims. In Aim 1, we will identify the contributions of specific cell types and ECM to BHTG structure, phenotype, and Mtb growth. We will specifically investigate the signals regulating monocyte differentiation and migratory behavior, and understand microenvironmental factors that optimize the containment and killing of Mtb. In Aim 2, we introduce a novel 3D model of vascularized human TB granuloma which will identify the mechanisms by which TB granulomas impact vascular morphogenesis, barrier function, and endothelial activation to facilitate monocyte homing and local vascular egress. To achieve these Aims, we have assembled a highly complementary research team with expertise in host immunity to Mtb (Ernst) and engineered biomimetic human culture models (Kutys). We will establish the long-term foundation for a new experimental platform to study TB granulomas and generate unprecedented knowledge and understanding of the dynamics of human TB granuloma generation, differentiation, and maintenance and the determinants of TB outcomes. Ultimately, these efforts will establish a predictive framework, which can be widely disseminated and adopted, for the identification and testing of new therapeutics for the control of human TB.
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