Modeling immune responses of cattle to Mycobacterium bovis using magnetic bioprinted granulomas

Tuberculosis (TB) remains a threat for human and livestock health. Mycobacteria causing TB are host-adapted pathogens that occasionally spill over into other species. Mycobacterium bovis causes bovine TB, a well-known zoonosis. Mycobacterium tuberculosis is adapted to humans and can occasionally trigger symptomatic infection in cattle. However, immunocompetent cattle are resistant to experimental infection with M. tuberculosis. Hallmarks of TB in susceptible hosts are organized multicellular tissue lesions termed granulomas. In the absence of suitable in vitro systems that enable investigations of bovine tuberculous granuloma, we developed a three-dimensional granuloma model using bovine leukocytes and magnetic cell labeling. This model was termed the in vitro granuloma-like structure (IVGLS). We generated stable IVGLS resembling TB granulomas at the innate stage, composed of macrophages, and at adaptive stages, containing lymphocytes in addition. M. bovis Bacillus Calmette-Guérin (BCG) replicated within IVGLS and triggered the progression of macrophages to foamy phenotypes. Within the IVGLS, the lymphocytes accelerated BCG-induced apoptotic cell death over time. IVGLS released abundant chemoattractants and Th1-associated cytokines and adopted a glycolytically polarized metabolism. Magnetic bioprinted bovine granulomas recapitulate features of TB granulomas and thus facilitate the study of immune responses to mycobacteria, including spatial and temporal mapping, as well as establishing precise cell death patterns within multicellular microenvironments. Deciphering protective immune responses within IVGLS can contribute to vaccine development for bovine TB, and elucidation of resistance mechanisms can facilitate the design of novel interventions for human TB.

Mycobacterial infections, including bovine tuberculosis (TB), have a profound impact on global health. This is exemplified by zoonotic TB in humans and animal TB, which is a life-threatening disease in livestock and wildlife. Mycobacteria cause the formation of granulomas, which significantly impact disease progression. Therefore, decoding granulomas is essential for an in-depth understanding of immune responses to mycobacteria. Conventional mouse models frequently fail to develop organized granulomas, and the procurement of samples from granulomatous lesions in cattle and humans is challenging, offering limited insights into the course of infection. Most in vitro TB research is confined to two-dimensional cell cultures, which neglect the spatial characteristics and cellular architecture of granulomas in vivo. To address this gap in knowledge, we have developed a novel multicellular in vitro model for TB. Our spheroid granuloma model, derived from bovine leukocytes using nanotechnologies, offers an adaptable platform for deciphering immune events within granulomas.