Macrophage global metabolomics identifies cholestenone as host/pathogen cometabolite present in human Mycobacterium tuberculosis infection

Abstract

Mycobacterium tuberculosis ( M. tuberculosis ) causes an enormous burden of disease worldwide. As a central aspect of its pathogenesis, M. tuberculosis grows in macrophages, and host and microbe influence each other’s metabolism. To define the metabolic impact of M. tuberculosis infection, we performed global metabolic profiling of M. tuberculosis –infected macrophages. M. tuberculosis induced metabolic hallmarks of inflammatory macrophages and a prominent signature of cholesterol metabolism. We found that infected macrophages accumulate cholestenone, a mycobacterial-derived, oxidized derivative of cholesterol. We demonstrated that the accumulation of cholestenone in infected macrophages depended on the M. tuberculosis enzyme 3 β -hydroxysteroid dehydrogenase (3 β -Hsd) and correlated with pathogen burden. Because cholestenone is not a substantial human metabolite, we hypothesized it might be diagnostic of M. tuberculosis infection in clinical samples. Indeed, in 2 geographically distinct cohorts, sputum cholestenone levels distinguished subjects with tuberculosis (TB) from TB-negative controls who presented with TB-like symptoms. We also found country-specific detection of cholestenone in plasma samples from M. tuberculosis –infected subjects. While cholestenone was previously thought to be an intermediate required for cholesterol degradation by M. tuberculosis , we found that M. tuberculosis can utilize cholesterol for growth without making cholestenone. Thus, the accumulation of cholestenone in clinical samples suggests it has an alternative role in pathogenesis and could be a clinically useful biomarker of TB infection.

Immune evasion and provocation by Mycobacterium tuberculosis

“…A variety of parameters have been shown to correlate with bacterial load and/or treatment response. For example, sputum abundance of cholestenone, a mycobacterium-derived cholesterol metabolite, correlates with Mycobacterium tuberculosis lung burden 198 . M. tuberculosis -specific CD4 + T cells that express PD1 define a population of T cells that have seen antigen and correlate with mycobacterial load 199 .…”

Defining the Genes Required for Survival of Mycobacterium bovis in the Bovine Host Offers Novel Insights into the Genetic Basis of Survival of Pathogenic Mycobacteria

“…In a recent extended Tn screen utilizing diverse mouse genotypes, Tn insertions in hsd caused reduced fitness in a small panel of selected genotypes indicates there may be some host genetic component to the requirement for cholesterol oxidation by hsd ( 40 ). Given the potential for the use of host cholesterol metabolites, specifically cholestenone, as diagnostic biomarkers, this observation might have applications in the development of diagnostics ( 41 ).…”

Understanding the contribution of metabolism to Mycobacterium tuberculosis drug tolerance

“…In humans, direct RNA-sequencing of Mtb from patient sputum revealed up-regulation of transcripts encoding enzymes required for cholesterol degradation ( Lai et al., 2021 ). Furthermore, the Mtb-specific cholesterol byproduct 4-cholesten-3-one is increased in patients with active tuberculosis, suggesting that Mtb actively metabolizes cholesterol during infection ( Chandra et al., 2022 ). These data indicate that Mtb carbon metabolism is shifted in the host to preferentially rely on lipids over carbohydrate carbon sources.…”