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Advisor(s)
Abstract(s)
Tuberculosis (TB) is a disease caused by Mycobacterium tuberculosis (Mtb) that results in 1.6 million deaths yearly. The TB granuloma is the hallmark cellular structure of latent TB that contains the spread of infection1 . More comprehensive in vitro models of TB that better resemble the cellular and immunoregulatory complexity of the granuloma would facilitate the study of the interplay between the bacteria and the different immune system cells2 . We aim to generate an in vitro, 3D cell culture model of the TB granuloma that can be easily implemented using readily available commercial reagents and materials. A commercial encapsulation system based on sodium cellulose sulphate (NaCS) and Poly (diallyldimethylammonium chloride) (PDADMAC)3 was used to generate small capsules containing human peripheral blood mononuclear cells (PBMC) in the presence of GFPexpressing Mtb H37Rv and maintained in culture for several weeks. The 3D structure formed by the cells inside and outside the capsules was evaluated by fluorescence microscopy and flow cytometry to distinguish the different cell types, and how they are organised inside the sphere and to measure cell survival and bacteria replication. The results show that human PBMCs readily form 3D cellular aggregates around infected cells and that cells cultivated outside the capsules are attracted and surround the capsules in response to infection. The model could be maintained for several weeks before bacteria-induced cell necrosis. PBMC’s viability remained stable, with more than 80 % live cells following two weeks of culture. Moreover, adding an exterior layer of cells helped control bacterial replication, suggesting relevant communication between cells inside and outside the capsules to control the infection.