Wednesday, April 24, 2024
4/24/2024
PROJECT SUMMARY Tuberculosis was the leading infectious cause of mortality worldwide from 2007 to 2019, only dethroned in 2020 by the Covid-19 pandemic. Unlike Covid-19, there is no effective vaccine for tuberculosis, so it will remain a leading infectious cause of death worldwide. With multidrug-resistant strains emerging, there is a critical need to develop new therapies that enhance the immune response to infection. The mechanisms used by Mtb to evade elimination are incompletely understood. The Ernst lab has observed that in mice infected with M. tuberculosis, certain phagocytes (mononuclear cell subset 1; MNC1) descended from bone marrow monocytes are especially “permissive”: they are more frequently infected than are other phagocytes and poorly eliminate engulfed mycobacteria. However, accumulation of monocyte-derived cells in inflamed tissues is associated with pathogen clearance in other infections. Recent work has also shown that the bone marrow monocyte population is not homogeneous; rather, subpopulations of monocytes have differing transcriptional profiles and have varying responses to distinct inflammatory stimuli. These data prompt the hypothesis that M. tuberculosis reprograms the bone marrow to produce monocytes that differentiate into lung cells that are highly permissive of mycobacterial growth and survival. To test this hypothesis, spectral flow cytometry will be used to determine whether acquisition of the phenotype observed in MNC1 cells begins in bone marrow monocytes from Mtb- infected mice. Bone marrow monocytes from uninfected and infected mice will also be co-transferred into infected recipients to determine whether monocyte origin influences differentiation into MNC1 cells. The heterogeneity of bone marrow monocytes will also be assessed by single-cell RNA sequencing to test multiple hypotheses about mechanisms underlying the differentiation of permissive lung cells. Monocytes from Mtb- infected mice will be compared to those from uninfected mice and from mice administered lipopolysaccharide. In a separate experiment, gene expression in bone marrow monocytes from Mtb-infected mice will be compared to that in monocytes from mice infected with the attenuated mycobacterial strain M. bovis bacillus Calmette- Guérin Pasteur, to link Mtb-specific genes to altered monocyte development. The proposed studies will improve understanding of how M. tuberculosis affects monocytes recruited to the lung from the bone marrow and could lead to host-directed therapies to augment their mycobactericidal activity, an innovative target that would increase the effectiveness of current anti-mycobacterial drugs. All work will be performed at UCSF, which offers a supportive, well-resourced training environment for physician-scientists like the applicant. The proposed studies will provide him with expertise in monocyte development, experimentation in a high-containment facility, and bioinformatic analysis, facilitating his transition to an independent investigator studying immune responses to lung infections.
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