A new study reveals that aging hematopoietic stem cells trigger a form of innate immune memory, known as trained immunity, that fuels the chronic low-grade inflammation that underlies age-related physiological decline. SIRT3, a mitochondrial deacetylase whose expression wanes with age, emerges as a key molecular brake on this process.

Clonal hematopoiesis (CH), an age-related expansion of somatically mutated hematopoietic clones, is associated with increased risk of severe infections including coronavirus disease (COVID)-19, yet the underlying mechanisms remain unclear. Here, we investigated the impact of Dnmt3a deficiency in a murine model of influenza A virus (IAV) pneumonia. Dnmt3a-deficient mice exhibited increased pulmonary viral burden and reduced neutrophil accumulation in IAV-infected lungs despite comparable circulating neutrophil numbers. Functional analyses of neutrophils showed impaired chemotactic migration in vitro, whereas maturation, antimicrobial enzyme content, and metabolic capacity were unchanged. Transcriptomic profiling revealed downregulation of pathways involved in chemotaxis, cytokine signaling, and cellular activation, including reduced expression of Cxcr1. Supporting the translational relevance of these findings, proteomic analysis of plasma from individuals with germline DNMT3A mutations (Tatton-Brown-Rahman syndrome) revealed alterations in proteins associated with cell migration and cytoskeletal dynamics. Collectively, our findings demonstrate that Dnmt3a loss compromises innate immune defense by impairing neutrophil migration in a cell-intrinsic manner, leading to ineffective pathogen clearance. This work provides mechanistic insight into how CH-associated mutations contribute to age-associated susceptibility to infection and highlights altered leukocyte trafficking as a potential therapeutic target in aging populations with CH.

Abstract Background: Clonal hematopoiesis (CH), defined using mutations in myeloid-lineage driver genes (N=74 [classic], Bick et al. Nature 2020), is a risk factor for hematologic malignancy (HM). Bernstein et al. (Nature Genetics 2024) identified 17 additional genes with mutations that are positively selected in the population, and documented a positive association with HM. Continuing to refine CH mutations that convey risk is critical for clinical use and research on modifiable drivers of CH to HM. Thus, we developed a CH-calling pipeline that detects CH from population-based sequencing data using an expanded gene list as well as a variant pathogenicity score based on Google AlphaFold 3. We investigated associations between CH called by the new pipeline and HM risk in a community-based cohort with long follow up. Methods: We conducted a prospective analysis of 9,365 participants in the Atherosclerosis Risk in Communities study aged 44-80 years, without a cancer history, and with whole exome sequencing data. 410 HMs (147 myeloid) were ascertained mainly by cancer registry linkage in a median follow up of 21 years. Follow up is longer than in the original well-known CH studies. With a mean depth of 100x, mutations with a variant allele frequency (VAF)>3.8% were considered for CH. We classified definitive CH as mutations in classic CH genes and truncating mutations in genes additionally identified by Bernstein et al. For missense mutations in the additional genes, we applied AlphaFold 3 to predict pathogenicity and called probable if score≥0.9. We estimated hazard ratios (HR) and 95% confidence intervals (CI) for the association of small (3.8 Citation Format: Elizabeth A. Platz, Hidetaka Uryu, Vernon A. Burk, Meng Ru, Sergiu Pasca, Lukasz P. Gondek, Katherine Y. King, Anna E. Prizment, Corinne E. Joshu, Pradeep Natarajan, Margaret A. Goodell, Christie M. Ballantyne, Koichi Takahashi. Clonal hematopoiesis and incident hematologic and myeloid malignancy in ARIC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 6291.

Clonal hematopoiesis of indeterminant potential (CHIP) is an age-related phenomenon associated with increased risk of hematologic malignancy. Preclinical studies have shown that infection is a driver of CHIP; clinical studies in people living with HIV suggest a relationship between chronic infection and CHIP, but the association between infection frequency and incident CHIP in the general population remains unknown. We leveraged the atherosclerosis risk in communities study to design a closed prospective cohort study. CHIP was determined based on whole-exome sequencing at 2 time points 20 years apart. Included were 3,367 individuals without cancer or CHIP at time 1 and without hematologic malignancy by time 2. The 3,367 study participants had an average age of 55.3 years at time 1; 59.1% were women, 40.9% were men; 24% were Black, and 76% were White. Documented infection was assessed from routinely collected hospital discharge summaries. Frequency was categorized as no documented infection, 1 infection, 2 infections, or ≥3 infections. Of the participants, 19.7% had incident CH, 6.9% had large CHIP, and 5.2% had large non-DNMT3A CHIP. Participants with ≥3 documented infections had an increased odds of incident CHIP (odds ratios [OR] 1.41, p = 0.03), especially large CHIP (OR 1.83, p = 0.008) and large non-DNMT3A CHIP (OR 1.81; p = 0.02). To our knowledge, this study is the first to demonstrate an association between infection and incident CHIP in a general population, highlighting a modifiable risk factor for CHIP. Further work is required to describe the mutation-specific impact underlying this observed relationship.

Clonal hematopoiesis is emerging as a surprising modifier of Alzheimer's disease. Recent findings suggest that mutant myeloid cells may enter or expand within the brain, adopting either inflammatory or reparative states. We propose that their effects depend on the mutation, timing, clone size, brain niche, and disease stage.

Clonal hematopoiesis (CH) arises when hematopoietic stem cells (HSCs) gain a fitness advantage from somatic mutations and expand, resulting in an increase in variant allele frequency (VAF) over time. To analyze CH trajectories, we develop a state-dependent stochastic model of wild-type and mutant HSCs, in which an environmental parameter α ∈ [ 0 , 1 ] regulates death rates and interpolates between homeostatic (Moran-like, α = 1 ) and growth-facilitating ( α < 1 ) regimes. Using functional law of large numbers and central limit theorems, we derive explicit mean-field dynamics and a Gaussian-Markov approximation for VAF fluctuations. We show that the mean VAF trajectory has an explicit logistic form determined by selective advantage, while environmental effects affect only the variance and autocovariance structure. Building on these results, we introduce BESTish (Bayesian estimate for selection incorporating scaling-limit to detect mutant heterogeneity), a novel, efficient and accurate Bayesian inference method that can be applied to analyze both cohort-level and longitudinal VAF datasets. BESTish implements the closed-form finite-dimensional distributions that we derive to estimate mutation fitness, mutation rate, and environmental strength for individual CH drivers. When applied to existing CH datasets, BESTish produces consistent mutation fitness inferences across different studies, and estimates CH driver mutation rates in agreement with independent experimental studies. Furthermore, BESTish reveals patient-specific heterogeneity in the selective behavior of recurrent mutations, and identifies variants whose dynamics are compatible with non-homeostatic, growth-facilitating environments. BESTish provides a unified and mechanistic framework for quantifying CH evolution, with potential applications for other biological systems where clonal expansions can be measured.

Analysis of lung alveolar type 2 (AT2) progenitor stem cells has highlighted fundamental mechanisms that direct their differentiation into alveolar type 1 cells (AT1s) in lung repair and disease. However, microRNA (miRNA) mediated post-transcriptional mechanisms which govern this nexus remain understudied. We show here that the let-7 miRNA family serves a homeostatic role in governance of AT2 quiescence, specifically by preventing the uncontrolled accumulation of AT2 transitional cells and by promoting AT1 differentiation. Using mice and organoid models, we demonstrate genetic ablation of let-7a1/let-7f1/let-7d cluster (let-7afd) in AT2 cells prevents AT1 differentiation and results in accumulation of AT2 transitional cells in progressive pulmonary fibrosis. Integration of AGO2-eCLIP with RNA-sequencing from AT2 cells uncovered the induction of direct targets of let-7 in an oncogene feed-forward regulatory network including BACH1/EZH2/MYC which drives an aberrant fibrotic cascade. Additional analyses using CUT&RUN-sequencing revealed an epigenetic role of let-7 in induction of chromatin histone acetylation and methylation and maladaptive AT2 cell reprogramming. This study identifies let-7 as a key gatekeeper of post-transcriptional and epigenetic chromatin signals to prevent AT2-driven pulmonary fibrosis.

Human and murine studies reveal that innate immune cells are able to mount enhanced responses to pathogens after primary inflammatory exposure. Innate immune memory has been shown to last for months to years, longer than the lifespan of most innate immune cells. Indeed, long-lived hematopoietic stem and progenitor cells (HSPCs) serve as a cellular reservoir for innate immune memory. In this review, we summarize the evidence that innate immune memory is epigenetically encoded in HSPCs, and we consider whether HSPC subpopulations with differentiation bias, cell autonomous epigenetic reprogramming, or both features underlie the phenomenon of central trained immunity. We further profile the significant implications of central trained immunity in stem cell transplant, aging, inflammatory diseases, and vaccination strategies for the future.

Recent studies suggest that infection reprograms hematopoietic stem and progenitor cells (HSPCs) to enhance innate immune responses upon secondary infectious challenge, a process called "trained immunity."However, the specificity and cell types responsible for this response remain poorly defined. We established a model of trained immunity in mice in response to Mycobacterium avium infection. scRNA-seq analysis revealed that HSPCs activate interferon gamma-response genes heterogeneously upon primary challenge, while rare cell populations expand. Macrophages derived from trained HSPCs demonstrated enhanced bacterial killing and metabolism, and a single dose of recombinant interferon gamma exposure was sufficient to induce similar training. Mice transplanted with influenza -trained HSPCs displayed enhanced immunity against M. avium challenge and vice versa, demonstrating cross protection against antigenically distinct pathogens. Together, these results indicate that heterogeneous responses to infection by HSPCs can lead to long-term production of bone marrow derived macrophages with enhanced function and confer cross-protection against alternative pathogens.

Infection leads to durable cell-autonomous changes in hematopoietic stem and progenitor cells (HSPCs), resulting in production of innate immune cells with heightened immunity. The mechanisms underlying this phenomenon, termed central trained immunity, remain poorly understood. We hypothesized that infection induces histone modifications leading to changes in chromatin accessibility that are conserved during differentiation from HSPCs to myeloid progenitors and monocytes. We conducted genome-wide surveillance of histone marks H3K27ac and H3K4me3 and chromatin accessibility in hematopoietic stem cells, multipotent progenitor 3, granulocyte-monocyte progenitors, and monocytes and macrophages of naive and Mycobacterium avium-infected mice. IFN signaling pathways and related transcription factor binding motifs including IRFs, NF-κB, and CEBP showed increased activating histone marks and chromatin accessibility across cell types. However, histone marks and increased chromatin accessibility were conserved at only a few loci, notably Irf1 and Gbp6. Knock out of IRF1 disrupted enhanced mitochondrial respiration and bacterial killing in human monocyte cell lines, while GBP6-KO monocyte cell lines showed dysregulated mitochondrial respiration. In summary, this study identifies IRF1 and GBP6 as 2 key loci at which infection-induced systemic inflammation leads to epigenetic changes that are conserved from HSPCs to downstream monocytes, providing a mechanistic avenue for central trained immunity.

While the clonal expansion of mutant hematopoietic stem cells (HSCs) known as clonal hematopoiesis (CH) is associated with many age-associated diseases, the connection to neurodegenerative diseases including Alzheimer’s Disease (AD) is less clear. Prior evidence suggests systemic inflammatory signals can be transmitted to the central nervous system (CNS), where they trigger microglia activation and amyloid-beta clearance. Here we investigated the role of the two most commonly mutated clonal hematopoiesis-associated genes, DNMT3A and TET2, in the pathogenesis of AD using a mouse model. We utilized 5xFAD (Familial Alzheimer’s Disease) mice which express human disease variants of amyloid precursor protein and presenilin and develop b-amyloid plaques and cognitive decline by 6-9 months of age. At 6-8 weeks of age, 5xFAD mice were non-competitively transplanted with Dnmt3a-/-, Tet2-/- or wildtype (WT) bone marrow (BM). Mice were then challenged weekly with LPS to mimic age-associated chronic inflammation. At 6 months of age, transplanted mice were assessed for signs of AD pathogenesis as well as the presence of infiltrating peripheral immune cells within the brain. 5xFAD mice transplanted with Dnmt3a-/- BM displayed exacerbated AD, including worsened cognitive impairment and decreased microglia activation compared to those transplanted with WT BM. They also had fewer peripheral immune cells infiltrating the brain compared to WT-transplanted recipients. In contrast, 5xFAD mice transplanted with Tet2-/- BM showed improved cognitive status, decreased amyloid plaques, and increased microglia activation. Tet2-/- transplanted mice had a higher percentage of activated infiltrating myeloid cells in the brain compared to WT controls. To assess whether peripherally derived immune cells directly replace microglia in the CNS, we quantified donor-derived peripheral immune cells positive for the microglia marker Tmem119 within the brain using flow cytometry. Interestingly, mice transplanted with Tet2-/- BM, had a higher proportion of donor-derived mutant microglia-like cells when compared to mice transplanted with WT BM. To evaluate the impact of DNMT3A or TET2 mutation on human microglial function, we utilized microglia derived from isogenic human iPSC lines harboring DNMT3A (DNMT3AR882H/WT) or TET2 mutations (TET2DelE3-E11/WT). Cytokine bead array of culture supernatant showed that TET2-mutant induced microglia (iMGs) released significantly more inflammatory cytokines and demonstrated a greater capacity to phagocytose both myelin and b-amyloid upon LPS activation compared to WT or DNMT3A-mutant iMGs. In summary, Tet2 provided protection against AD through increased myeloid cell infiltration and microglia activation, leading to more efficient amyloid plaque clearance and improved cognitive performance; while the opposite was true for loss of Dnmt3a. Our study of CH and AD marks the first report in which DNMT3A and TET2, which have opposite roles in DNA methylation, induced opposing effects on disease progression. Citation Format: Katherine Y King. Dnmt3a and Tet2-driven clonal hematopoiesis have opposing effects on the pathogenesis of Alzheimer’s disease [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: DNA Methylation, Clonal Hematopoiesis, and Cancer; 2025 Feb 1-4; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2025;85(3 Suppl):Abstract nr IA017

Clonal hematopoiesis (CH) is associated with many age-related diseases, but its interaction with Alzheimer’s disease (AD) remains unclear. Here, we show that TET2-mutant CH is associated with a 47% reduced risk of late-onset AD (LOAD) in the UK Biobank, whereas other drivers of CH do not confer protection. In a mouse model of AD, transplantation of Tet2-mutant bone marrow reduced cognitive decline and β-amyloid plaque formation, effects not observed with Dnmt3a-mutant marrow. Bone-marrow-derived microglia-like cells were detected at an increased rate in Tet2-mutant marrow recipients, and TET2-mutant human induced pluripotent stem cell (iPSC)-derived microglia were more phagocytic and hyperinflammatory than DNMT3A-mutant or wild-type microglia. Strikingly, single-cell RNA sequencing (scRNA-seq) revealed that macrophages and patrolling monocytes were increased in brains of mice transplanted with Tet2-mutant marrow in response to chemokine signaling. These studies reveal a TET2-specific protective effect of CH on AD pathogenesis mediated by peripheral myeloid cell infiltration.

MicroRNA-mediated post-transcriptional regulation of lung alveolar type 2 (AT2) and AT1 cell differentiation remains understudied. Here, we demonstrate that the let-7 miRNA family plays a homeostatic role in AT2 quiescence by preventing the uncontrolled accumulation of AT2 transitional cells and promoting AT1 differentiation. Using mouse and organoid models, we show that genetic ablation of let-7a1/let-7f1/let-7d cluster (let-7afd) in AT2 cells prevents AT1 differentiation and leads to KRT8 transitional cell accumulation in progressive pulmonary fibrosis. Integration of AGO2-eCLIP with RNA-sequencing identified direct let-7 targets within an oncogene feed-forward regulatory network, including BACH1/EZH2/MYC, which drives an aberrant fibrotic cascade. Additional CUT&RUN-sequencing analyses revealed that let-7afd loss disrupts histone acetylation and methylation, driving epigenetic reprogramming and altered gene transcription in profibrotic AT2 cells. This study identifies let-7 as a central hub linking unchecked oncogenic signaling to impaired AT2 cell plasticity and fibrogenesis.

ABSTRACT:Prolonged or broad-spectrum antibiotic courses are associated with intestinal dysbiosis and cytopenias, and depletion of hematopoietic progenitor populations after antibiotics is associated with loss of peripheral immune cells, leading to increased susceptibility to systemic infections. We evaluated the bone marrow hematopoietic compartment in a murine model of antibiotic exposure. Single-cell RNA sequencing revealed a substantial and previously unrecognized depletion of bone marrow B cells at all stages of development in antibiotic-treated mice, further confirmed by flow cytometric analysis. Depletion of the microbiota was associated with rapid changes in the peripheral B-cell compartment, yet fecal microbiota transfer did not rescue either peripheral or bone marrow B cells to a greater degree than natural recovery from antibiotic treatment. Antibiotic-mediated loss of B-cell progenitors was secondary to enhanced apoptosis and occurred independent of disrupted systemic type I and II interferon signaling, previously implicated in the maintenance of other hematopoietic compartments. Instead, the depletion of prosurvival MYC signaling was implicated in the depletion of circulating lymphocytes and bone marrow B-cell progenitor populations during antibiotic treatment. Furthermore, in vitro exposure of bone marrow cells to antibiotics demonstrated significantly decreased viability of B cells. We conclude that both microbiota depletion and cytotoxic effects of prolonged broad-spectrum antibiotic treatment disrupt cytokine and cell survival signaling critical for B-cell progenitor maintenance. These results contribute to our understanding of the compartment-specific mechanisms by which the microbiota maintains the hematopoietic system and suggest critical pathways for maintenance of bone marrow progenitors during prolonged antibiotic treatment.

Clonal hematopoiesis (CH) is associated with a higher hematologic malignancy (HM) risk, and risk increases with variant allele frequency (VAF). We determined whether inflammatory factors modify the association between CH and HM. We conducted a prospective cohort analysis of 9, 978 participants (55.9% female, 29.5% Black) in the Atherosclerosis Risk in Communities (ARIC) study who were 44-80 years old and did not have a cancer history. Median follow up was 20.9 years. CH (VAF≥2%: 9.8%; VAF≥10% [large CH]: 4.7%) was identified from whole exome sequencing data constrained to 74 genes (Bick et al. Nature 2020). Incident HM (N=434) was ascertained mainly by cancer registry linkage. Aspirin was assessed at study visits by review of containers of prescribed and over the counter drugs used in the last 2 weeks. Plasma proteins were measured by SomaScan® 5K assay. We prespecified interleukin (IL)-6, interferon-gamma (IFN-gamma), IL-8, IL-1beta, and tumor necrosis factor (TNF)-alpha. We estimated the association between CH and HM stratified by aspirin use and plasma protein levels (median; 8, 693 participants) using Cox regression adjusting for age, sex, race, and risk factors for HM. The association between large CH (vs no CH) and HM was weaker in aspirin users (HR=1.58, 95% CI 0.91-2.72, p-trend [across no, small, large CH]=0.26) than in non-users (HR=2.02, 95% CI 1.29-3.15, p-trend=0.008). The association between large CH and HM was stronger in participants with higher (HR=2.11, 95% CI 1.29-3.44, p-trend=0.10) than lower (HR=1.38, 95% CI 0.77-2.48, p-trend=0.56) plasma IL-6. Median VAF in those with large CH was comparable between higher (18%) and lower (16%) IL-6 strata, as was percentage of common genes mutated (DNMT3A: higher 48%, lower 48%; TET2: higher 21%, lower 23%). Patterns of association between CH and HM by strata of IFN-gamma (higher: p-trend=0.014; lower: p-trend=0.46) and IL-8 (higher: p-trend=0.01, lower: p-trend=0.46) were similar to IL-6. In contrast, for TNF-alpha strata, the association between large CH and HM was weaker in participants with higher (p-trend=0.20) than with lower (p-trend=0.06) levels, and the percentage with mutated DNMT3A appeared to differ by TNF-alpha level (higher 51%, lower 43%). Associations between large CH and HM were comparable in the higher (p-trend=0.05) and lower (p-trend=0.12) IL-1beta strata. Regular aspirin use may dampen CH-associated hematologic malignancy risk. Higher plasma levels of some pro-inflammatory cytokines, especially IL-6, may exacerbate CH-associated hematologic malignancy risk. While IL-6 is produced by immune cells with DMNT3A and/or TET2 mutations, the similar VAF and percentage of mutated gene by IL-6 level suggests that IL-6 is not simply marking CH extent. Laboratory experiments are needed to establish cause for these inflammation mediators. Support: NHLBI, NCI, NPCR Elizabeth A. Platz, Vernon A. Burk, Meng Ru, Sergiu Pasca, Lukasz P. Gondek, Katherine Y. King, Ajibike D. Lapite, Anna Prizment, Corinne E. Joshu, Pradeep Natarajan, Margaret A. Goodell, Christie M. Ballantyne, Koichi Takahashi. Mediators of inflammation modify the association between clonal hematopoiesis and incident hematologic malignancy in ARIC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 7357.

Haematopoietic stem cells maintain blood production throughout life1. Although extensively characterized using the laboratory mouse, little is known about clonal selection and population dynamics of the haematopoietic stem cell pool during murine ageing. We isolated stem cells and progenitors from young and old mice, identifying 221,890 somatic mutations genome-wide in 1,845 single-cell-derived colonies. Mouse stem cells and progenitors accrue approximately 45 somatic mutations per year, a rate only approximately threefold greater than human progenitors despite the vastly different organismal sizes and lifespans. Phylogenetic patterns show that stem and multipotent progenitor cell pools are established during embryogenesis, after which they independently self-renew in parallel over life, evenly contributing to differentiated progenitors and peripheral blood. The stem cell pool grows steadily over the mouse lifespan to about 70,000 cells, self-renewing about every 6 weeks. Aged mice did not display the profound loss of clonal diversity characteristic of human haematopoietic ageing. However, targeted sequencing showed small, expanded clones in the context of murine ageing, which were larger and more numerous following haematological perturbations, exhibiting a selection landscape similar to humans. Our data illustrate both conserved features of population dynamics of blood and distinct patterns of age-associated somatic evolution in the short-lived mouse.

Hematologic side effects are associated with prolonged antibiotic exposure in up to 34% of patients. Neutropenia, reported in 10-15% of patients, increases the risk of sepsis and death. Murine studies have established a link between the intestinal microbiota and normal hematopoiesis. We sought to identify predisposing factors, presence of microbiota-derived metabolites, and changes in intestinal microbiota composition in otherwise healthy pediatric patients who developed neutropenia after prolonged courses of antibiotics. In this multi-center study, patients with infections requiring anticipated antibiotic treatment of two or more weeks were enrolled. Stool samples were obtained at the start and completion of antibiotics and at the time of neutropenia. We identified 10 patients who developed neutropenia on antibiotics and 29 controls matched for age, sex, race, and ethnicity. Clinical data demonstrated no association between neutropenia and type of infection or type of antibiotic used; however intensive care unit admission and length of therapy were associated with neutropenia. Reduced intestinal microbiome richness and decreased abundance of Lachnospiraceae family members correlated with neutropenia. Untargeted stool metabolomic profiling revealed several metabolites that were depleted exclusively in patients with neutropenia, including members of the urea cycle pathway, pyrimidine metabolism and fatty acid metabolism that are known to be produced by Lachnospiraceae . Our study confirms a relationship between intestinal microbiota disruption and abnormal hematopoiesis and identifies taxa and metabolites likely to contribute to microbiota-sustained hematopoiesis. As the microbiome is a key determinant of stem cell transplant and immunotherapy outcomes, these findings are likely to be of broad significance. Key Points:Neutropenia occurred in 17% of patients receiving prolonged antibiotic therapy.We found no association between neutropenia and type of infection or class of antibiotic used. Development of neutropenia after prolonged antibiotic treatment was associated with decreased prevalence of Lachnospiraceae and Lachnospiraceae metabolites such as citrulline.