immune cell composition in the different sites was largely maintained across age, except for significantly decreased frequencies of cd8 t n cells in the blood and lns, concomitant increases in t em cells in the blood and t rm cells in the lns and lower frequencies of classical monocytes in the bm ( fig. 5c and supplementary table 14 ). these results indicate that tissue - driven immune cell composition and profile are largely maintained with age. to interrogate specific effects of aging on immune cells across lineages and sites, we conducted a separate de analysis for each tissue group and immune subset with sufficient representation, comparing younger ( < 40 years ) versus older ( > 40 years ) individuals, while controlling for sex, cmv status and other covariates ( supplementary tables 15 – 17 ). an embedding of similarities between age - related degs revealed that some immune cells exhibited changes specific to tissue site ( for example, t cells in the ln, lymphocytes in the jej ), while others showed subset - specific changes independent of site ( for example, monocytes in blood, bm and lung, and memory b cells in lns and lung ) ( fig. 5d ). we identified genes regulated with age in each subset and tissue by integrating de results with gsea ( supplementary fig. 8a ). for myeloid lineage cells, mucosal sites had the most age - related degs, with similar trends in lymphoid organs ( fig. 5e – g ). age - related changes in classical monocytes included increased expression of genes associated with proliferation and inflammation ( kras, calm1 ) and decreased expression of genes for macrophage differentiation ( for example, rab44 ) ( fig. 5e ); non - classical monocytes showed age - associated increase in expression of genes for cell – cell interactions ( lgals1, itga9 ) and decreased expression of metabolism and mitochondrial regulation genes ( lyrm7, sars2 ) ( fig. 5f ). macrophages had decreased expression of genes associated with metabolism and mitochondrial fitness ( marc1, mtor ) and increased expression of genes associated with m2 macrophages ( id1, adgre1 npr2 ) and interferon signaling ( mx2 ) ( fig. 5g ) along with increased cd95 ( fas ) at the protein level ( fig. 5h ). together, these age - related changes in monocytes and macrophages were subset - specific, enriched in mucosal sites and indicated decreased overall fitness. we used schpf to identify age - associated gene signatures for myeloid cells and gsea to assess their expression in different subsets and sites ( supplementary tables 18 and 19 ). we uncovered an apoe – trem2 signature, including apolipoprotein genes ( apoc1, apoc2 and apoe ) and trem2, a triggering receptor expressed on myeloid cells that binds apoe and facilitates macrophage functions, such as phagocytosis and chemotaxis, and induces metabolic changes ( fig. 5i ). this apoe – trem2 signature was significantly downregulated with age in monocytes and macrophages in the lungs, lymphoid organs and blood ( fig. 5j ), and in an independent published dataset from human lungs ( n = 29; see methods ) ( fig. 5k ). overall, this analysis showed subset and site - specific features of macrophage aging involving a major functional and metabolic pathway. we applied the above approaches to identify age - associated signatures in t cells and b cells. for cd4 t cells, cd8 t cells and b cells, there were relatively few genes ( for example, those associated with oxidation and inflammation: sod1, il18bp, il15 ) that changed over age in two or more subsets within each lineage ( extended data fig. 5a – d ). pathway analysis revealed increased inflammation, apoptosis and reduced tcr signaling across multiple t cell subsets with age ( supplementary fig. 8b ). despite the paucity of age - associated gene expression changes across subsets, cd8 t emra cells exhibited multiple age - associated changes conserved across sites; these included increased expression of nk cell genes ( ncam1, klrf1, gnly ) and the nk cell marker cd56, consistent with findings in blood, and reduced expression of genes associated with signaling ( cd6, jak3 ), proliferation and metabolism ( tcf7, rptor ) ( fig. 6a, b ). by schpf analysis, we identified two prominent age - associated transcriptional signatures shared across multiple cd8 t cell subsets. a cytokine signature, containing genes for effector cytokines and chemok ines ( for example, ccl3, ccl4, xcl1, ifng, tnf ), was increased with age in all cd8 t cell subsets and γδ t cells across all sites examined ( fig. 6c, d ). this aging signature was also detected in published datasets from human lungs ( fig. 6e ) and in peripheral blood mononuclear cells ( pbmcs ) from the sound life cohort ( n = 96, age 26 – 65 years ) within the human immune health atlas ( extended data fig. 5e ). a second signature contained gzmk encoding the cytolytic molecule granzyme k, the transcription factor eomes and activation or signaling markers ( pdcd1, hla - dr, fcrl3 ) ( fig. 6f ). this signature aligned with a granzyme k - containing, age - associated signature identified in t cells in aged mice and human blood and in pbmcs from the human immune health atlas ( extended data fig. 5f ). this gzmk signature was increased with age in cd8 t emra cells, γδ t cells and cd8 t em cells across tissues but not in cd8 t rm cells in lungs and jej ( fig. 6g ). the frequency and clonal expansion of cd8 t emra cells enriched in the gzmk signature was higher in older than in younger donors ( fig. 6h, i ). these results show distinc t age - associated signatures in mucosal resident t cells compared to circulation. for b cells, there were more age - related degs in memory compared to naive b cells in lns ( fig. 6j and extended data fig. 5d ). memory b cells from older donors had increased expression of genes associated with inflammatory cytokines ( il18, il18bp ), cell adhesion ( cd58, lgals1 ) and cell death or autophagy ( fas, itm2a ), along with reduced expression of proliferation ( cdca7, il2ra ), lipid metabolism ( acsm3, pnpla7 ) and differentiation markers ( fig. 6j ). select il - 18 pathway - associated genes in b cells and cd4 t cells in blood were validated in the human immune health atlas cohort ( supplementary fig. 9a, b ). transcript and / or surface expression of igm ( ighm ) and ighd were reduced in older compared to younger b cells, and the frequency of igm b cells decreased with age ( fig. 6k, l and supplementary table 20 ). we identified by schpf that a gene signature related to ras signaling ( rasa4b, rasgrf1, gab2 ) downstream of the bcr was downregulated with age in naive and memory b cells across all sites ( fig. 6m, n ). we validated this age - associated signature in bm - derived b cells from an independent dataset ( n = 39, age 2 – 84 years ) and in pbmcs from the human immune health atlas ( fig. 6o and extended data fig. 5g ). pathway analysis further revealed increased inflammation and reduced bcr signaling in ln b cells with age ( supplementary fig. 8c ). these results showed that b cells exhibited diminished signaling and functional dysregulation across tissues over age. cmv infection drives immune cell alterations, including increased accumulation of t emra cells with age in blood, spleen and lungs. we investigated the impact of cmv serostatus on cell composition and immune aging and found no significant associations with cd4 t cell, cd8 t cell or b cell frequencies ( supplementary fig. 10a, b ). two cd8 t cell signatures were associated with cmv serostatus after regression of other covariates: the gzmk signature and a gnly signature ( gnly, fgfbp2 and cx3cr1 ) ( supplementary fig. 10c – f ). the gnly signature was enriched across all cd8 t cell subsets in cmv donors, while the gzmk signature was variably enriched in different sites and subsets of cmv donors by gsea ( supplementary fig. 10d, f and supplementary tables 21 and 22 ).