Summary
The healthspan of mice is enhanced by killing senescent cells using a transgenic suicide gene. Achieving the same using small molecules would have a tremendous impact on quality of life and ...the burden of age‐related chronic diseases. Here, we describe the rationale for identification and validation of a new class of drugs termed senolytics, which selectively kill senescent cells. By transcript analysis, we discovered increased expression of pro‐survival networks in senescent cells, consistent with their established resistance to apoptosis. Using siRNA to silence expression of key nodes of this network, including ephrins (EFNB1 or 3), PI3Kδ, p21, BCL‐xL, or plasminogen‐activated inhibitor‐2, killed senescent cells, but not proliferating or quiescent, differentiated cells. Drugs targeting these same factors selectively killed senescent cells. Dasatinib eliminated senescent human fat cell progenitors, while quercetin was more effective against senescent human endothelial cells and mouse BM‐MSCs. The combination of dasatinib and quercetin was effective in eliminating senescent MEFs. In vivo, this combination reduced senescent cell burden in chronologically aged, radiation‐exposed, and progeroid Ercc1−/Δ mice. In old mice, cardiac function and carotid vascular reactivity were improved 5 days after a single dose. Following irradiation of one limb in mice, a single dose led to improved exercise capacity for at least 7 months following drug treatment. Periodic drug administration extended healthspan in Ercc1−/∆ mice, delaying age‐related symptoms and pathology, osteoporosis, and loss of intervertebral disk proteoglycans. These results demonstrate the feasibility of selectively ablating senescent cells and the efficacy of senolytics for alleviating symptoms of frailty and extending healthspan.
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DOBA, FZAB, GIS, IJS, IZUM, KILJ, NLZOH, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBMB, UILJ, UKNU, UL, UM, UPUK
Senescence is a tumor suppressor mechanism activated in stressed cells to prevent replication of damaged DNA. Senescent cells have been demonstrated to play a causal role in driving aging and ...age-related diseases using genetic and pharmacologic approaches. We previously demonstrated that the combination of dasatinib and the flavonoid quercetin is a potent senolytic improving numerous age-related conditions including frailty, osteoporosis and cardiovascular disease. The goal of this study was to identify flavonoids with more potent senolytic activity.
A panel of flavonoid polyphenols was screened for senolytic activity using senescent murine and human fibroblasts, driven by oxidative and genotoxic stress, respectively. The top senotherapeutic flavonoid was tested in mice modeling a progeroid syndrome carrying a p16INK4a-luciferase reporter and aged wild-type mice to determine the effects of fisetin on senescence markers, age-related histopathology, disease markers, health span and lifespan. Human adipose tissue explants were used to determine if results translated.
Of the 10 flavonoids tested, fisetin was the most potent senolytic. Acute or intermittent treatment of progeroid and old mice with fisetin reduced senescence markers in multiple tissues, consistent with a hit-and-run senolytic mechanism. Fisetin reduced senescence in a subset of cells in murine and human adipose tissue, demonstrating cell-type specificity. Administration of fisetin to wild-type mice late in life restored tissue homeostasis, reduced age-related pathology, and extended median and maximum lifespan.
The natural product fisetin has senotherapeutic activity in mice and in human tissues. Late life intervention was sufficient to yield a potent health benefit. These characteristics suggest the feasibility to translation to human clinical studies.
NIH grants P01 AG043376 (PDR, LJN), U19 AG056278 (PDR, LJN, WLL), R24 AG047115 (WLL), R37 AG013925 (JLK), R21 AG047984 (JLK), P30 DK050456 (Adipocyte Subcore, JLK), a Glenn Foundation/American Federation for Aging Research (AFAR) BIG Award (JLK), Glenn/AFAR (LJN, CEB), the Ted Nash Long Life and Noaber Foundations (JLK), the Connor Group (JLK), Robert J. and Theresa W. Ryan (JLK), and a Minnesota Partnership Grant (AMAY-UMN#99)-P004610401–1 (JLK, EAA).
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Senescent cells, which can release factors that cause inflammation and dysfunction, the senescence-associated secretory phenotype (SASP), accumulate with ageing and at etiological sites in multiple ...chronic diseases. Senolytics, including the combination of Dasatinib and Quercetin (D + Q), selectively eliminate senescent cells by transiently disabling pro-survival networks that defend them against their own apoptotic environment. In the first clinical trial of senolytics, D + Q improved physical function in patients with idiopathic pulmonary fibrosis (IPF), a fatal senescence-associated disease, but to date, no peer-reviewed study has directly demonstrated that senolytics decrease senescent cells in humans.
In an open label Phase 1 pilot study, we administered 3 days of oral D 100 mg and Q 1000 mg to subjects with diabetic kidney disease (N = 9; 68·7 ± 3·1 years old; 2 female; BMI:33·9 ± 2·3 kg/m2; eGFR:27·0 ± 2·1 mL/min/1·73m2). Adipose tissue, skin biopsies, and blood were collected before and 11 days after completing senolytic treatment. Senescent cell and macrophage/Langerhans cell markers and circulating SASP factors were assayed.
D + Q reduced adipose tissue senescent cell burden within 11 days, with decreases in p16INK4A-and p21CIP1-expressing cells, cells with senescence-associated β-galactosidase activity, and adipocyte progenitors with limited replicative potential. Adipose tissue macrophages, which are attracted, anchored, and activated by senescent cells, and crown-like structures were decreased. Skin epidermal p16INK4A+ and p21CIP1+ cells were reduced, as were circulating SASP factors, including IL-1α, IL-6, and MMPs-9 and −12.
“Hit-and-run” treatment with senolytics, which in the case of D + Q have elimination half-lives <11 h, significantly decreases senescent cell burden in humans.
NIH and Foundations.
ClinicalTrials.gov Identifier: NCT02848131. Senescence, Frailty, and Mesenchymal Stem Cell Functionality in Chronic Kidney Disease: Effect of Senolytic Agents.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Objective
The aim of this study was to determine the effect of age of obesity onset on senescence‐related markers in abdominal (AB) and femoral (FEM) subcutaneous adipose tissue (SAT) before and ...after moderate (~10%) weight loss.
Methods
AB and FEM SAT were collected from human females with childhood‐onset obesity (CO) or adult‐onset obesity (AO) before and after diet‐ and exercise‐induced weight loss. Immunofluorescence analysis of γH2AX/RAD51 (DNA damage/repair markers) and p53/p21 (senescence markers) was conducted in cultured preadipocytes, and senescence‐associated β‐galactosidase (SA‐β‐gal) activity was measured in SAT.
Results
CO had proportionately more AB and FEM preadipocytes with DNA damage (γH2AX+) and senescence markers (p53+ and/or p21+) than AO at baseline. The proportion of γH2AX+ FEM preadipocytes declined with weight loss in CO and was similar between groups after weight loss. The number of γH2AX foci in γH2AX+ preadipocytes decreased similarly between groups and regions with weight loss in parallel with an increase in RAD51. The proportion of p53+ and p21+ preadipocytes and SA‐β‐gal+ cells in SAT did not change with weight loss, but the total p21 intensity in p53+/p21+ FEM preadipocytes declined in AO.
Conclusions
These results provide preliminary evidence that females with CO have an accelerated preadipocyte aging state that improves with weight loss in terms of DNA damage but not senescence.
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FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Preadipocytes exposed to octanoate accumulate less lipid than cells exposed to long-chain fatty acids. This effect of octanoate involves significant attenuation of expression of key adipogenic ...transcription factors, including peroxisome proliferator-activated receptor (PPAR)γ, steroid regulatory binding element protein (SREBP)-1c and CCAAT element binding protein (C/EBPα) at both the mRNA and protein levels. Expression of differentiation markers, including adipocyte fatty acid binding protein (ALBP), glycerol-3-phosphate dehydrogenase (GPDH) and leptin, was also significantly diminished by octanoate. However, octanoate did not prevent the decrease in preadipocyte factor-1 (Pref-1) expression that occurs during adipogenesis, nor did it inhibit the early induction of C/EBPβ,δ. Treatment with synthetic PPARγ ligands partially offset the inhibitory effect of octanoate on differentiation. Ectopic expression of PPARγ2 in 3T3-L1 cells partially restored lipid accretion and GPDH activity in octanoate-treated cells. Adding octanoate together with troglitazone attenuated the effects of troglitazone on adipocyte differentiation in both normal 3T3-L1 cells and engineered 3T3-L1 cells that expressed ectopic PPARγ2, implying that octanoate might compete against troglitazone for its binding to PPARγ. These results suggest that octanoate may block adipogenesis at least in part by its influence on the expression/activation of PPARγ.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Senescent cells, which arise due to damage-associated signals, are apoptosis-resistant and can express a pro-inflammatory, tissue-destructive senescence-associated secretory phenotype (SASP). We ...recently reported that a component of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) surface protein, S1, can amplify the SASP of senescent cultured human cells and that a related mouse β-coronavirus, mouse hepatitis virus (MHV), increases SASP factors and senescent cell burden in infected mice. Here, we show that SARS-CoV-2 induces senescence in human non-senescent cells and exacerbates the SASP in human senescent cells through Toll-like receptor-3 (TLR-3). TLR-3, which senses viral RNA, was increased in human senescent compared to non-senescent cells. Notably, genetically or pharmacologically inhibiting TLR-3 prevented senescence induction and SASP amplification by SARS-CoV-2 or Spike pseudotyped virus. While an artificial TLR-3 agonist alone was not sufficient to induce senescence, it amplified the SASP in senescent human cells. Consistent with these findings, lung p16
senescent cell burden was higher in patients who died from acute SARS-CoV-2 infection than other causes. Our results suggest that induction of cellular senescence and SASP amplification through TLR-3 contribute to SARS-CoV-2 morbidity, indicating that clinical trials of senolytics and/or SASP/TLR-3 inhibitors for alleviating acute and long-term SARS-CoV-2 sequelae are warranted.
A key challenge in aging research is extending lifespan in tandem with slowing down functional decline so that life with good health (healthspan) can be extended. Here, we show that monthly ...clearance, starting from 20 months, of a small number of cells that highly express p21Cip1 (p21high) improves cardiac and metabolic function and extends both median and maximum lifespans in mice. Importantly, by assessing the health and physical function of these mice monthly until death, we show that clearance of p21high cells improves physical function at all remaining stages of life, suggesting healthspan extension. Mechanistically, p21high cells encompass several cell types with a relatively conserved proinflammatory signature. Clearance of p21high cells reduces inflammation and alleviates age-related transcriptomic signatures of various tissues. These findings demonstrate the feasibility of healthspan extension in mice and indicate p21high cells as a therapeutic target for healthy aging.
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•p21high cells, distinct from p16high cells, accumulate in various tissues with aging•Intermittent p21high cell clearance extends both median and maximum lifespans in mice•Intermittent p21high cell clearance results in sustained health improvement•Intermittent p21high cell clearance alleviates age-related transcriptomic signatures
Wang et al. identify a distinct senescent cell population (p21high cells) in various tissues with aging. p21high cells encompass numerous cell types with a conserved proinflammatory signature. Intermittent clearance of p21high cells reduces inflammation, extends lifespan, and results in sustained benefits to health and physical function in old mice.
In clinical organ transplantation, donor and recipient ages may differ substantially. Old donor organs accumulate senescent cells that have the capacity to induce senescence in naïve cells. We ...hypothesized that the engraftment of old organs may induce senescence in younger recipients, promoting age-related pathologies. When performing isogeneic cardiac transplants between age-mismatched C57BL/6 old donor (18 months) mice and young and middle-aged C57BL/6 (3- or 12- month-old) recipients , we observed augmented frequencies of senescent cells in draining lymph nodes, adipose tissue, livers, and hindlimb muscles 30 days after transplantation. These observations went along with compromised physical performance and impaired spatial learning and memory abilities. Systemic levels of the senescence-associated secretory phenotype factors, including mitochondrial DNA (mt-DNA), were elevated in recipients. Of mechanistic relevance, injections of mt-DNA phenocopied effects of age-mismatched organ transplantation on accelerating aging. Single treatment of old donor animals with senolytics prior to transplantation attenuated mt-DNA release and improved physical capacities in young recipients. Collectively, we show that transplanting older organs induces senescence in transplant recipients, resulting in compromised physical and cognitive capacities. Depleting senescent cells with senolytics, in turn, represents a promising approach to improve outcomes of older organs.
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BFBNIB, FZAB, GIS, IJS, KILJ, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
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