Investigating tuberculosis pathology in the human lung with 3D X-ray Histology; a correlative imaging approach

Abstract

Tuberculosis kills more than 1.5 million people every year. (1) Most studies on tuberculosis use either animal or in vitro cell culture models that do not recapitulate human disease. The most common method of disease investigation (used in both research and clinical workflows) is the route of traditional histology, which involves thinly sectioning the sample and chemically staining said sections to extract specific molecular/cellular information about the tissue. However, such two-dimensional systems present with limitations regarding tissue connectivity and three-dimensionality of tissue structures. Consequently, fundamental mechanisms in human tuberculosis disease are poorly understood. (2-6) In this project, we have developed a correlative imaging approach by integrating non-invasive 3D micro-CT imaging (7-9) with traditional 2D histology of formalin-fixed paraffin-embedded (FFPE) human lung biopsies from tuberculosis patients. We leverage the third dimension added to our histology imaging data by micro-CT to explore disease process in tuberculosis, interrogating the structure of tuberculous granulomas and the interconnectivity of tissue microenvironments which is lost when imaging representative 2D sections by traditional optical microscopy.Human tuberculous lung biopsies were first imaged with micro-CT. The FFPE biopsies were then thinly sectioned and each section was stained with either histological or tinctorial stains to reveal areas of tuberculous pathology, fibrosis and calcification. Sections were scanned using a slide-scanner and optical microscopy. Manual segmentation in 3D was based on annotations performed by a consultant histopathologist on the digital histology slides. Preliminary data suggests that these analyses will provide unique insight into human disease and the interrelationships of microenvironments in 3D. 
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Identifiers

ORCID for Elena Konstantinopoulou: orcid.org/0000-0003-4077-9648

ORCID for Orestis L. Katsamenis: orcid.org/0000-0003-4367-4147

ORCID for Philip J Basford: orcid.org/0000-0001-6058-8270

ORCID for Philipp Schneider: orcid.org/0000-0001-7499-3576

ORCID for Paul Elkington: orcid.org/0000-0003-0390-0613

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