Longevity Papers

Lysosomal dysfunction is a well-recognized feature of aging. Here, we used a suite of tools for rapid lysosomal isolation to construct a multitissue atlas of the metabolite changes lysosomes undergo during aging. Aged lysosomes in brain, heart, muscle, and white adipose tissue accumulated glycerophosphodiesters and cystine, metabolites that are causally linked to juvenile lysosomal storage disorders, Batten disease, and cystinosis. Levels of these metabolites increased linearly with age, preceding organismal decline. Caloric restriction, a lifespan-extending intervention, mitigated these changes in the heart and muscle but not the brain. Our findings link lysosomal storage disorders to aging-related dysfunction and open avenues for the mechanistic investigation of how lysosomal functions deteriorate during aging and in age-associated diseases.

Cardiac fibrosis, a major pathological hallmark of aging that leads to heart failure, is characterized by excessive collagen deposition. Our knowledge of what sustains collagen synthesis in the aging heart is still very preliminary. Here, we uncover a central role for chaperone-mediated autophagy (CMA), a selective lysosomal degradation pathway, in this process. We demonstrate that CMA is suppressed in the aging heart, which promotes collagen overproduction in fibroblasts, whereas enhancing CMA activity ameliorates fibrosis and diastolic dysfunction. Mechanistically, we identify SHMT2 (serine hydroxymethyltransferase 2) as a CMA substrate whose accumulation with aging drives collagen synthesis by increasing glycine availability. Integrative omics revealed a systemic downregulation of the ketone body β-hydroxybutyrate (BHB) in aged mice. BHB supplementation - via a cyclic ketogenic diet - restored CMA, attenuated fibrosis, and improved cardiac function. This recovery was mediated through BHB-induced activation of the HCAR2 receptor and subsequent phosphorylation of HSPA8/HSC70, which systemically reactivates the CMA machinery. Furthermore, we show that

Aging is characterized by progressive physiological decline and accumulation of senescent cells that drive chronic "inflammaging" through the senescence-associated secretory phenotype (SASP). The complement system, traditionally viewed as a systemic extracellular defense mechanism, is now recognized as an essential intracellular network (the complosome). This review synthesizes current research on how intracellular C3 (intC3), and intracellular C5 (intC5) in certain contexts engage in extensive crosstalk with the mTOR, NF-κB, and AMPK pathways to modulate core cellular processes. We detail how intC3 intersects with multiple canonical hallmarks of aging-including cellular senescence, mitochondrial dysfunction, proteostasis loss, genomic instability, epigenetic alterations, altered intercellular communication, stem cell exhaustion, and deregulated nutrient sensing. Furthermore, we examine organ-specific consequences of intC3 dysregulation across the aging brain, liver, eye, kidney, vasculature, lung, and immune system, as well as cancer. Finally, we discuss therapeutic strategies-including complement inhibitors, senolytics, senomorphics, RNA interference, proteolysis targeting chimeras (PROTACs), and nanotechnology-enabled delivery-and outline critical gaps in compartment-specific tools, conditional knockouts, and longitudinal models. Precision strategies that preserve systemic complement function while selectively modulating intracellular pools will be essential to translate complosome biology into durable interventions for age-related diseases.

The aging brain exhibits a decline in the regenerative populations of neural stem cells (NSCs). While mechanisms that restore old NSC function have started to be identified, the role of lipids-especially complex lipids-in NSC aging remains largely unclear. Using lipidomic profiling by mass spectrometry, we identify age-related changes in complex lipids in quiescent NSCs in vitro and in vivo. Moreover, several polyunsaturated fatty acids increase across lipid classes in quiescent NSCs during aging. Using spatial lipidomics, we find that some of the changes in complex lipids are also observed in situ. Several age-related changes in complex lipids and side chain composition are occurring at the plasma membrane, as revealed by lipidomic profiling of isolated plasma membrane vesicles. Experimentally, we show that aging is accompanied by a decrease in plasma membrane order, a key membrane biophysical property, in old quiescent NSCs in vitro and in vivo. To determine the functional role of plasma membrane lipids in aging NSCs, we performed genetic and supplementation studies. Knocking out the phospholipid acyltransferase MBOAT2 exacerbates age-related lipidomic changes in old quiescent NSCs and impedes their ability to activate.

Importance: Preclinical and human observational studies suggest that metformin may decrease age-related pathology, including frailty. Objective: Determine whether metformin reduces frailty progression and biological age in older adults with glucose intolerance, a population at increased risk of becoming frail. Design, Setting and Participants: Randomized, double-blind, placebo-controlled trial of metformin in 145 non-frail or pre-frail older adults. Participants (72 +/-5 years, 48% female, 94% White, 35% Hispanic) were randomized to metformin vs. placebo for two years. Main Outcomes and Measures: Effect on frailty was primarily determined using generalized estimating equations by change in the Fried frailty phenotype score (based on weight loss, exhaustion, physical activity, gait speed, and grip strength). Because metformin can cause significant weight loss, effects on the Fried score were assessed with and without the weight loss criterion. Frailty also was assessed by change in the frailty index (composite of 95 deficits). Biological age was estimated by DNA methylation-based epigenetic clocks in blood. Results: Metformin led to a non-linear response in the Fried score rate of change, with an upward trajectory in year 1 (0.72 +/-0.22 per year vs. placebo, p=0.0011) and stabilization in year 2 (-0.33 +/-0.17 per year vs. placebo, p=0.056). Metformin led to more weight loss than placebo (-5.7 +/-5.2 vs. -2.3 +/-5.4 kg, p=0.0002); thus, when assessing effect on Fried score without the weight loss criterion, no difference was observed, indicating that weight loss in year 1 accounted for the change in Fried score. Notably, metformin caused a steady improvement in the frailty index (-0.006 +/-0.0026 per year vs. placebo, p=0.0222) that persisted with covariates adjustment including body mass index. Metformin reduced biological age estimated by PC-Horvath2 (-0.40 +/-0.16 per year, p=0.014) and PC-Hannum (-0.33 +/-0.16 per year, p=0.047) clocks. Metformin was well tolerated. Conclusions and Relevance: Metformin halts the progression of the deficit accumulation frailty index and reduces biological age, suggesting potential benefit for extending healthspan. Trial Registration: ClinicalTrials.gov: NCT02570672.

Cellular senescence contributes to neurodegeneration in Alzheimer's disease (AD), yet brain-penetrant senotherapeutic strategies remain limited. Here, we identify long interspersed nuclear element 1 (LINE-1) retrotransposons as key regulators of neuronal senescence and the senescence-associated-secretory-phenotype (SASP) in AD. Using transdifferentiated induced neurons (iNs) that preserve donor-specific aging-associated molecular signatures, we show that pharmacological inhibition of LINE-1 with nucleoside reverse transcriptase inhibitors (nRTIs) or antisense oligonucleotides reduces p16 expression, suppresses SASP and interferon-stimulated gene programs, and attenuates paracrine induction of reactive astrogliosis. Spatial transcriptomic analysis of human AD brain tissue further supports that senescent neurons with high LINE-1 expression are localized to inflammatory niches in the brain. Although bulk analysis finds no significant differences in LINE-1 expression between AD and control neurons, long-read single-cell RNA sequencing of iNs identifies a subset of neurons with elevated LINE-1 activity which display transcriptional signatures of neurodegeneration, immune activation, and senescence are enriched in AD relative to controls. RNA velocity analysis indicates that LINE-1 activation precedes the induction of canonical senescence markers, supporting a causal rather than consequential role. Mechanistically, LINE-1-derived cytoplasmic DNA activates the cGAS-STING innate immune pathway in post-mitotic neurons, and inhibition of cGAS phenocopies the effects of LINE-1 suppression. Together, these findings establish a LINE-1/cGAS-STING axis as a driver of neuronal senescence in AD and highlight LINE-1 inhibition as a tractable senomorphic strategy for neurodegenerative disease.

Cellular senescence is accompanied by profound lysosomal alterations, yet whether lysosome-associated factors actively drive aging remains unclear. Through a focused CRISPR/Cas9 screen in human mesenchymal progenitor cells (hMPCs), we identified N-acetylglucosamine-1-phosphotransferase subunits alpha and beta (GNPTAB), an enzyme responsible for lysosomal hydrolase targeting, as a potent regulator of cellular senescence. Genetic ablation of GNPTAB attenuated senescence, whereas its overexpression accelerated senescence. This pro-senescent function occurred independently of GNPTAB's canonical enzymatic role. Instead, GNPTAB binds to the innate immune adaptor stimulator of interferon genes (STING) via a specific interface (E1119), leading to activation of STING and its downstream TANK-binding kinase 1 (TBK1), as well as pro-inflammatory gene expression. A STING-binding-deficient GNPTAB mutant (E1119A) preserved canonical lysosomal functions but failed to induce senescence, while STING depletion abolished GNPTAB-driven senescence. Together, these findings uncover a new signaling pathway wherein GNPTAB engages STING to facilitate its activation, nominating this interface as a potential target for mitigating age-related cellular dysfunction.