Structured Abstract
INTRODUCTION
Aging is accompanied by parallel functional decline across organs, but the cellular drivers remain unclear. Tissue-resident macrophages (TRMs), long-lived cells that comprise 60 to 90% of macrophages in major organs, maintain homeostasis through efferocytosis of apoptotic and senescent cells. Neutrophils, the most abundantly produced and shortest-lived leukocytes (more than 100 billion generated daily in humans), require continuous TRM clearance; uncleared aged neutrophils release proteases and extracellular traps that damage tissues and propagate aging. TRMs express the prostaglandin E2 (PGE2) receptor EP2, which suppresses macrophage metabolism and phagocytosis in aging. Whether impaired TRM efferocytosis drives the accumulation of senescent neutrophils that promote organ aging and whether inhibition of EP2 signaling can restore this process, remain unknown.
RATIONALE
We studied aged mice in which EP2 signaling on TRMs was selectively reduced, either genetically (TRM-specific EP2 deletion) or pharmacologically, to define how TRM dysfunction shapes organ-wide aging. TRMs are long-lived gatekeepers of tissue homeostasis, and EP2 offers a tractable target because its activity increases in aged macrophages and suppresses their metabolic and phagocytic function. Using complementary genetic and pharmacological approaches, we tested whether restoring TRM function reverses organ aging and identified which efferocytic substrate is most affected. The same design also defined the molecular step at which EP2 acts within TRMs and extended our findings to aged human tissues.
RESULTS
In aged mice, TRM-specific EP2 deletion restored mitochondrial fitness and immune homeostasis, and reversed cognitive decline, frailty, sarcopenia, adiposity, and cardiac dysfunction toward youthful states. Plasma proteomics identified the liver as a major source of age-associated immune changes. Single-cell RNA-seq of mouse liver and multiorgan flow cytometry revealed accumulation of senescent CXCR4+ neutrophils across efferocytic organs in aging. These cells exhibited the senescence-associated secretory phenotype (SASP), DNA damage response activation, cell cycle inhibitor induction, NETosis, and anti-apoptosis programs, and were efficiently cleared following EP2 deletion. Liver multiplex imaging localized paracrine stress to parenchymal cells neighboring senescent neutrophils. Ex vivo efferocytosis assays showed that aged TRMs were most impaired in clearing senescent neutrophils relative to apoptotic substrates, with both functions restored by EP2 deletion or pharmacologic antagonism. Mechanistically, EP2 signaling suppressed integrin-dependent stabilization of senescent neutrophils on TRMs and downstream engulfment. Analyses of human liver and heart datasets revealed conserved EP2 up-regulation in aged TRMs, enrichment of senescent neutrophils, and reduced TRM–neutrophil interactions.
CONCLUSION
This work identifies EP2 signaling in TRMs as a central regulator of organ-wide aging through its control of senescent neutrophil clearance, reframing aging as a failure of active cellular clearance rather than passive degeneration. With age, neutrophils acquire senescence-associated features, and their accumulation drives tissue injury through two converging mechanisms: intrinsic degranulation and NETosis, and extrinsic paracrine stress on neighboring parenchymal cells. Pharmacological inhibition of EP2 restores TRM efferocytic capacity and promotes clearance of senescent neutrophils, positioning EP2 antagonism as a tractable therapeutic strategy for age-related organ and functional decline.
Tissue-resident macrophage (TRM) clearance of senescent neutrophils is restored by EP2 deletion or inhibition to limit organ aging.
(Top) In aged TRMs, EP2 signaling suppresses (1) integrin engagement with senescent neutrophil (Neu) ligands required for clearance and (2) engulfment of senescent neutrophils into phagolysosomes, allowing senescent neutrophils to accumulate, (3) undergo degranulation and NETosis, and (4) exert paracrine stress on neighboring parenchymal cells, (5) collectively driving organ aging.
(Bottom) EP2 deletion or inhibition restores (1) integrin-mediated interaction and (2) phagolysosomal engulfment, limiting (3) degranulation and NETosis, (4) paracrine stress, and (5) organ aging.
NETosis, neutrophil extracellular trap (NET) formation; AC, adenylyl cyclase; PKA, protein kinase A; TFs, transcription factors.
