µCT Analysis of the Human Tuberculous Lung Reveals Remarkable Heterogeneity in 3D Granuloma Morphology.

Abstract

Rationale: Our current understanding of tuberculosis pathophysiology is limited by a reliance on animal models, the paucity of human tuberculosis lung tissue, and traditional histopathological analysis, a destructive two-dimensional approach that provides limited spatial insight. Determining the three-dimensional (3D) structure of the necrotic granuloma, a characteristic feature of tuberculosis, will more accurately inform preventative TB strategies. Objectives: To ascertain the 3D shape of the human tuberculous granuloma and its spatial relationship with airways and vasculature within large lung tissues. Methods: We characterized the 3D microanatomic environment of human tuberculous lungs using micro-computed tomography (µCT), histopathology and immunohistochemistry. Using 3D segmentation software, we accurately reconstructed TB granulomas, vasculature, and airways in 3D and confirmed our findings using histopathology and immunohistochemistry. Measurements and main results: We observed marked heterogeneity in the morphology, volume, and number of TB granulomas in human lung sections. Unlike depictions of granulomas as simple spherical structures, human necrotic granulomas exhibit complex, cylindrical, branched morphologies which are connected to the airways and shaped by the bronchi. 3D imaging of human tuberculosis lung sections provides unanticipated insight into the spatial organization of tuberculosis granulomas in relation to airways and the vasculature. Conclusions: Our findings highlight the likelihood that a single structurally complex lesion could be mistakenly viewed as multiple independent lesions when evaluated in 2D. Also, lack of vascularization within obstructed bronchi establishes a paradigm for antimycobacterial drug tolerance. Lastly, our results suggest that bronchogenic spread of Mycobacterium tuberculosis re-seeds the lung.