Microbiome-Derived Metabolites Shape CD4⁺ T-Cell Differentiation and Immune Aging in Chronic HIV-1 Infection

Abstract The role of aromatic gut-derived bacterial metabolites (GDBMs) in shaping immune cell metabolism and function remains poorly explored. Using ex vivo metabolomic profiling of paired plasma and CD4⁺ T-cells from people living with HIV-1 (PLWH), we identified a network of aromatic GDBMs whose cell-associated abundance, rather than systemic levels, was linked to broad alterations in CD4⁺ T-cell metabolic and functional states. Among these metabolites, p-cresol sulfate (PCS) emerged as a mechanistic prototype investigated in depth. Ex vivo flow cytometry and single-cell RNA sequencing of CD4⁺ T-cells stratified by cell-associated PCS levels revealed dose-dependent enrichment of transcriptional programs associated with impaired differentiation capacity, regulatory-like identity, and cellular senescence. Consistently, in vitro transcriptomic and proteomic analyses of PCS-exposed CD4⁺ T cells demonstrated induction of cell-cycle arrest, mitochondrial dysfunction, and senescence-associated programs, including upregulation of p16 and p21. Integration of these immunometabolic features with measurements of HIV-1 reservoir size in PLWH revealed that CD4⁺ T-cell states defined by cell-associated GDBMs track with intact proviral DNA levels in vivo. Together, these findings define a microbiome-derived axis that reshapes CD4⁺ T-cell metabolism and fate and promotes immune aging–associated states in PLWH. Our data suggest that cell-associated GDBMs may foster immunometabolic CD4⁺ T-cell states previously linked to long-term HIV-1 reservoir persistence in vivo. Graphical Abstract. PCS-driven metabolic reprogramming and senescence promoting CD4+ T-cell immune cell aging. Dietary proteins are metabolized by proteolytic gut microbiota into p-cresol, which is absorbed and converted in the liver to PCS. Circulating PCS accumulates in CD4⁺ T-cells, where it activates the aryl hydrocarbon receptor (AhR). AhR signaling reduces glycolysis and mTOR activity, while enhancing TGF-β, Wnt/β-catenin, and TCF7 pathways, driving a regulatory-like and stem-like transcriptional program. These changes are associated with increased expression of p16 and p21, leading to cell cycle arrest and cellular senescence promoting CD4 + T-cell immune cell aging.