Emerson, L. | bioRxiv Science

bioRxiv2023

A microfluidic model of colonocyte-microbiota interaction mimicking the colorectal cancer microenvironment

Changes in the abundance of certain bacterial species within the colorectal microbiota correlate with colorectal cancer development. While carcinogenic mechanisms of single pathogenic bacteria have been characterized in vitro, limited tools are available to investigate interactions between pathogenic bacteria and both commensal microbiota and colonocytes in a physiologically relevant tumor microenvironment. To address this, we developed a microfluidic device that can be used to co-culture colonocytes and colorectal microbiota. The device was used to explore the effect of Fusobacterium nucleatum, an opportunistic pathogen associated with colorectal cancer development in humans, on colonocyte gene expression and microbiota composition. F. nucleatum altered the transcription of genes involved in cytokine production, epithelial-to-mesenchymal transition, and proliferation in colonocytes in a contact-independent manner; however, most of these effects were diminished by the presence of fecal microbiota. Interestingly, F. nucleatum significantly altered the abundance of multiple bacterial clades associated with mucosal immune responses and cancer development in the colon. Our results highlight the importance of evaluating the potential carcinogenic activity of pathogens in the context of a commensal microbiota, and the potential to discover novel inter-species microbial interactions in the colorectal cancer microenvironment.

bioengineering↗

bioRxiv2022

Leishmania infection-derived extracellular vesicles drive transcription of genes involved in M2 polarization.

Although it is known that the composition of EVs is determined by the characteristics of the cell and its environment, the effects of intracellular infection on EV composition and functions are not well understood. We had previously shown that cultured macrophages infected with Leishmania parasites release EVs that contain parasite derived molecules (LiEVs). In this study we show that LdVash, a molecule previously identified in LiEVs from L. donovani infected RAW264.7 macrophages is widely distributed in the liver of L. donovani infected mice. This result shows for the first time that parasite molecules are released in EVs and distributed in infected tissues where they can be endocytosed by cells in the liver, including macrophages that undergo a significant increase in numbers as the infection progresses. To commence evaluating the potential impact of LiEVs on macrophage functions, we show that primary peritoneal exudate macrophages (PECs) express transcripts of signature molecules of M2 macrophages such as arginase 1, IL-10 and IL-4R when incubated with LiEVs. In comparative studies that illustrate how intracellular pathogens control the composition and functions of EVs released from macrophages, we show that EVs from RAW264.7 macrophages infected with Salmonella enterica serovar Typhimurium activate PECs to express transcripts of signature molecules of M1 macrophages such as iNOS, TNF alpha and IFN gamma and not M2 signature molecules. Finally, we show that in contrast to the polarized responses observed in in vitro studies of macrophages, both M1 and M2 signature molecules are detected in L. donovani infected livers although they exhibit differences in their spatial distribution in infected tissues.

microbiology↗