Investigating the host-pathogen interaction in tuberculosis by bioelectrospray 3D cell culture modelling and label-free imaging

Tuberculosis is a deadly infectious disease caused by the bacterium Mycobacterium tuberculosis (Mtb). It constitutes a major cause of death worldwide with approximately 1.5 million people dying from TB every year.1-6 Traditional animal models as well as conventional 'two-dimensional' cell cultures do not accurately mimic human tuberculosis infection in vivo, such as the formation of caseating granulomas and degradation of extracellular matrix. We study a bioelectrospray-generated 3D cell culture model of tuberculosis, using diverse imaging techniques. In the 3D culture, Mtb-infected PBMCs (strains: H37Rv, 0414B) are mixed with an alginate-collagen gel. We investigated the host-pathogen interaction at high resolution at various stages of infection using Transmission Electron Microscopy (TEM). We used Micro-Computed Tomography (μCT) to show distribution of PBMCs in 3D in comparison to a human tuberculous lung biopsy. This was then correlated to traditional H&E, and matrix staining of the sectioned tuberculous human lung block. Using label-free microscopy [Coherent Anti-Stokes Raman Scattering (CARS), Second Harmonic Generation (SHG), Two-Photon Excitation Fluorescence (TPEF)], PBMC aggregation and collagen ... (continues)