Longevity Papers

The increase in life expectancy over the past century has been accompanied by the recognition of age as the primary risk factor for a wide range of pathologies, including cardiovascular and neurodegenerative diseases, as well as cancer. Thus, the development of interventions that slow the underlying biological processes of aging could have beneficial effects on the prevention or progression of these diseases. To evaluate such geroprotective interventions, quantifying biological damage via aging clocks-particularly those based on transcriptomic biomarkers-has become highly relevant, as they provide estimations with strong biological interpretability. However, the limited availability of transcriptomic clocks for murine experimental models has hindered the implementation of these tools in aging research and their use in evaluating interventions that may have geroprotective effects. Here, we have developed an accurate, accessible, and ready-to-use murine transcriptomic clock that provides robust age predictions for healthy mice using hepatic RNA-seq data. Applying the clock to accelerated-aging models revealed an increased transcriptomic age in progeroid syndromes, demonstrating its ability to capture aging-related biological processes. Finally, considering the main interest of these tools, we show that well-established interventions with geroprotective potential, both genetic (Snell Dwarf, Ames Dwarf, and growth hormone receptor-deficient mice) and environmental (caloric, protein, or methionine restriction, as well as rapamycin supplementation in specific contexts), reduce the transcriptomic age estimated by the clock. These findings indicate that the proposed transcriptomic clock could be a valuable tool for studying the biology of aging and for designing and evaluating potential geroprotective interventions.

Physical exercise is a potent modifier of CNS aging, but its relationship with the cyclin-dependent kinase inhibitor p21 (CDKN1A) is neither uniformly suppressive nor equivalent to cellular senescence. Direct evidence remains sparse and preclinical. Sustained physiological exercise reduced hippocampal or cortical p21-associated signatures in models of metabolic distress, natural aging, and amyloid pathology. Conversely, acute treadmill exercise transiently increased hippocampal Cdkn1a, while spatial transcriptomics identified increased endothelial Cdkn1a during broader exercise-associated vascular rejuvenation. At the maladaptive extreme, excessive swimming produced sustained hippocampal p53-p21 activation accompanied by oxidative injury, apoptosis, senescence-associated β-galactosidase activity, and cognitive impairment. Genetic studies further indicate that endogenous p21 restrains exercise-responsive neural stem-cell activation, but they do not demonstrate that exercise itself reduces p21. To reconcile these findings, this review applies a four-axis interpretive matrix based on signal persistence, cellular and subcellular compartment, corroborating senescence-associated features, and functional consequence. The evidence is integrated into three states: physiological exercise associated with attenuation of persistent p21-related stress, transient or cell-specific adaptive p21 induction, and maladaptive exercise overload characterized by sustained p53-p21 signaling and tissue injury. Current findings therefore support exercise more strongly as a stress-preventive or senomorphic intervention than as a proven CNS senolytic or p21-targeted therapy. The central question is not simply whether exercise increases or decreases p21, but when, where, for how long, and within which cellular and molecular context the response reflects repair, quiescence, persistent senescence-like dysfunction, or progression toward cell death.

Cellular senescence contributes to aging and age-related diseases by driving chronic inflammation through the senescence-associated secretory phenotype (SASP), including interferon-stimulated genes (ISGs). Here we confirm and extend previous observations that cyclin D1 (CCND1), a key cell cycle regulator, is paradoxically upregulated across models of nonproliferating senescent cells. We show that CCND1 and its kinase partner CDK6 drive SASP and ISG expression in senescent cells by promoting DNA damage accumulation. This leads to the formation of cytoplasmic chromatin fragments that activate pro-inflammatory cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) signaling. In aged mouse livers, senescent hepatocytes show increased Ccnd1 expression. Hepatocyte-specific Ccnd1 knockout or treatment with the clinical grade CDK4/6 inhibitor palbociclib reduces DNA damage and ISGs in aged mouse liver. Further, palbociclib suppresses frailty and improves physical performance of aged mice. These findings demonstrate a role for CCND1/CDK6 in regulating DNA damage and inflammation in senescence and aging, highlighting it as a promising target for therapeutic repurposing.

While recent studies have established links between metabolic reprogramming and inflammatory senescence, the specific metabolic drivers in vascular aging remain incompletely defined. Here, we systematically characterized senescent phenotypes and targeted metabolomic profiles in primary aging endothelial cells, identifying a pyruvate dehydrogenase E1 component subunit alpha (PDHA1)-dependent metabolic shift as a hallmark of cellular senescence. Using a D-galactose-induced senescence model, we demonstrated that endothelial-specific

Insulin/IGF signaling (IIS) inhibits the nuclear localization of the DAF-16/FOXO transcription factor to regulate longevity and stress resistance in C. elegans. In the intestine, IIS promotes DAF-16 localization to endosomes and loss of TBC-2, a RAB-5 GAP, results in increased endomembrane localization of DAF-16 at the expense of nuclear localization, decreased DAF-16 target gene expression, longevity and stress resistance. Here we found that TBC-2 differentially regulates the localization of the IIS-regulated transcription factors PQM-1 and HLH-30/TFEB. Our results suggest a broader role for TBC-2 in negatively regulating IIS and that TBC-2 likely functions at an upstream point in the IIS pathway.

Aging is characterized by persistent low-grade inflammation linked to impaired tissue homeostasis, yet the underlying molecular mechanisms remain poorly understood. The mammalian skin is a clinically relevant site of aging-driven inflammation associated with compromised barrier function, inefficient wound healing, elevated oxidative stress and DNA damage accumulation. Here we show that, in the murine epidermis, aging engages a previously uncharacterized BMAL1-YAP functional cooperation with enhanced binding at inflammation-related enhancers, amplifying target gene transcription. Independent of its circadian clock role, BMAL1 partners with the mechanosensitive cofactor YAP at enhancer regions to regulate epidermal identity genes. However, in aged skin, this cooperative binding undergoes a functional shift, enhancing the expression of inflammation-related genes, partially coregulated by NF-κB. In addition, aged pro-inflammatory IL-17 signaling activates YAP in a Hippo-independent manner. These findings unveil a transcriptional mechanism underlying epidermal aging, linking chromatin dynamics to inflammatory programs through rewiring of BMAL1-YAP-occupied enhancers, highlighting potential strategies to counteract chronic inflammation and restore tissue homeostasis during aging.

Understanding how osteoblasts build and remodel bone matrix in vivo remains a fundamental challenge because cellular metabolism and matrix turnover are difficult to resolve across time and space within mineralized tissues. Here, we developed an integrated imaging platform combining stable isotope labeling with correlative electron microscopy and nanoscale secondary ion mass spectrometry (NanoSIMS) to visualize bone cell metabolism and matrix dynamics at nanometer resolution in vivo. This approach revealed rapid incorporation of dietary amino acids into osteoblast subcellular compartments within minutes of oral administration, followed by deposition of newly labeled extracellular matrix within hours. By linking elemental composition, isotope incorporation, and ultrastructure, we further show that cellular phosphorus signal is associated with early osteoblast amino acid incorporation. Multiday labeling revealed that newly deposited matrix is spatially associated with local osteocyte process architecture. Long-term amino acid tracing uncovered localized matrix turnover at osteocyte and osteoclast interfaces, including osteocyte-associated pericellular matrix remodeling and osteoclast association with newly formed, old, and mixed matrix regions. Finally, aging was associated with reduced osteoblast amino acid incorporation, diminished matrix deposition, and impaired osteocyte process-associated activity. Together, this work establishes a high-resolution platform for linking bone cell metabolism with matrix deposition and turnover in vivo, providing a broadly adaptable strategy to investigate skeletal aging, tissue remodeling, and metabolic dysfunction in disease.

Air pollution is the leading environmental cause of disease burden worldwide and is increasingly recognized as a potent driver of accelerated biological aging. Rather than acting through organ-specific toxicity alone, air pollutants appear to affect the fundamental hallmarks of aging itself, including oxidative stress, chronic inflammation, epigenetic dysregulation, telomere attrition, mitochondrial dysfunction, and cellular senescence. The pollutants of greatest concern are fine and ultrafine particulate matter (PM

Cellular senescence is associated with profound alterations in cellular physiology, including reduced membrane fluidity, impaired endosomal trafficking, diminished endocytic capacity, and increased extracellular RNase activity, all of which hinder efficient mRNA delivery. These barriers have limited the application of RNA-based approaches in senescent cells, particularly for delivering large therapeutic transcripts. This protocol describes an optimized reverse-transfection method for the efficient delivery of modified messenger RNA (mRNA) into senescent human fibroblasts. Although human telomerase reverse transcriptase (hTERT) mRNA was used as the model transcript, the workflow is broadly applicable to other mRNAs. In contrast to conventional transfection methods, in which RNA-lipid complexes are added to the culture medium after cell attachment, reverse transfection deposits the complexes onto the culture surface before cell seeding, enabling direct interaction between attaching cells and transfection complexes. To maximize transfection efficiency, the protocol incorporates nucleoside-modified mRNA containing pseudouridine and 5-methylcytidine, extended poly(A) tails, optimized complex-formation timing, RNase inhibition, transient elevation of endosomal pH with chloroquine, increased cell-seeding density, and extended incubation periods. Using this approach, transfection efficiencies of approximately 50%-80% were achieved in senescent fibroblasts following delivery of a 5 kb hTERT mRNA transcript. Peak telomerase activity was detected 24-48 h after transfection. A single transfection cycle produced measurable telomere elongation, whereas three sequential transfections resulted in substantial but finite telomere extension. Partial reversal of senescence-associated phenotypes was detectable within 72-96 h, including reduced senescence-associated β-galactosidase activity, decreased p16 and p21 expression, restoration of cell morphology, and extension of replicative lifespan. The delivered hTERT mRNA was degraded within 72-96 h, and immortalization was not observed. This protocol provides a practical approach for transient mRNA delivery into senescent cells and may be adaptable to a wide range of cell types and species.

Aging-associated loss of chromatin compaction is linked to derepression of retrotransposable elements (RTEs) in mouse and human tissues. Whether such RTE transcription contributes to the microglia activation that is common in aged brains is unknown. Here, we show that DAXX, a histone chaperone and RTE repressor, is downregulated during aging, preserves microglia homeostasis and inhibits cellular senescence. Loss of Daxx in young-adult microglia drives a reactive phenotype marked by chromatin decompaction at RTEs, loss of homeostatic markers, cell cycle re-entry and behavioral changes. This state leads to DNA damage and microglial depletion, followed by replacement with DAXX-deficient/Apoe

Aging remodels compartmentalized nicotinamide adenine dinucleotide (NAD⁺) circuits in ways that influence stress responses, senescence, tissue repair and susceptibility to fibrosis. Beyond its classical role as a redox cofactor, NAD⁺ fuels sirtuins, PARPs and CD38, linking cellular metabolism to chromatin remodeling, DNA repair, calcium signaling and cell fate. A central translational question is whether modulating NAD⁺ in a given tissue and time window will favor regeneration, restrain fibrotic remodeling, or support malignant adaptation. In this review, we define NAD⁺ as a compartmentalized, high-turnover metabolic circuit whose topology, timing and cell-type specificity shape tissue trajectories during aging, repair and fibrosis. We first describe how aging reshapes these circuits through CD38 upregulation, PARP-sirtuin competition, extracellular eNAMPT amplification, and altered mitochondrial NAD⁺ transport, which redistributes NAD⁺ between compartments without necessarily changing bulk tissue concentration. We then summarize how de novo, Preiss-Handler Handler (including niacin/GPR109a) and nicotinamide salvage pathways are organized across nuclear, cytosolic, mitochondrial and extracellular compartments, and how key consumers including sirtuins, PARPs, CD38 and SARM1 govern these pools. We then examine how these circuits operate across acute injury, chronic senescence-associated remodeling, and malignant microenvironments in heart, lung, liver, kidney, skin, muscle and brain. Finally, we integrate emerging human data on NAD⁺ precursors and enzyme-directed strategies and propose a framework for NAD⁺-based interventions that prioritizes timing, compartmental targeting and oncologic stratification.

Nuclear lamins provide structural integrity to the nuclear envelope through coiled-coil dimer meshworks. Lamin A contains a C-terminal immunoglobulin (Ig)-like domain and a cysteine-rich unstructured tail, whereas lamin C lacks the latter, retaining only one cysteine within the Ig-like domain. Mutations R435C and R471C in the Ig-like domain are linked to progeroid syndromes, fatal disorders characterized by premature aging. Here, we elucidate a pathogenic mechanism driven by aberrant disulfide cross-linking. We found that the R435C mutation, but not R471C, facilitates successive disulfide bond formation between Ig-like domains in vitro using purified recombinant proteins, causing nuclear deformation in lamin C-overexpressing cells. In lamin A-overexpressing cells, both R435C and R471C mutations induce additional intermolecular disulfide bonds involving the lamin A-specific cysteine residues in the C-terminal tail. Importantly, we demonstrate that glutathione and its precursor, N-acetyl cysteine, can disrupt these aberrant bonds. Using Caenorhabditis elegans as an in vivo model, we show that the orthologous cysteine mutation causes progeria phenotypes, which are suppressed by antioxidant treatment. These findings identify aberrant disulfide cross-linking as a key driver of progeria and suggest antioxidant therapies as a potential treatment strategy. Our study offers broader implications for vertebrate aging, suggesting that oxidative stress-mediated changes in lamin architecture are a conserved mechanism contributing to the loss of nuclear structural integrity and age-dependent nuclear aberration.

: Background: Mitochondrial dysfunction serves as a fundamental driver of the aging process, precipitating progressive functional decline through complex molecular cascades. Summary: This review elucidates the core mechanisms underpinning this dysfunction, including reactive oxygen species (ROS)-induced redox imbalance, mitochondrial DNA (mtDNA) damage accumulation, impaired mitophagy, and metabolic reprogramming. Crucially, we examine how mitochondria act as signaling hubs for inter-organ crosstalk. Through the secretion of mitokines (e.g., FGF21, GDF15) and the release of damage-associated molecular patterns (DAMPs), dysfunctional mitochondria trigger chronic inflammation via the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) and NLRP3 inflammasome (NLRP3) pathways, actively driving systemic aging within the skeletal muscle-brain and adipose/liver-cardiovascular axes. Additionally, this paper synthesizes current therapeutic interventions, ranging from lifestyle modifications and nicotinamide adenine dinucleotide (NAD+) precursors to frontier technologies like mitochondrial transplantation and gene editing. Key Messages: While promising in animal models, clinical translation of these interventions is currently hindered by limited long-term safety data and evidence gaps. Therefore, mitochondria-targeted studies incorporating integrated multi-organ phenotyping are urgently required to establish robust strategies for extending human healthspan.

Protein restriction extends lifespan across species and engages many hallmarks of aging. We propose that these diverse responses can be understood as components of a single coordinated physiological state. This response involves both cellular nutrient sensing and endocrine and neural coordination, with enhanced longevity emerging from this adaptive response.