Kirkland, James L.

AbstractFor many years, it was believed that the aging process was inevitable and that age‐related diseases could not be prevented or reversed. The geroscience hypothesis, however, posits that aging is, in fact, malleable and, by targeting the hallmarks of biological aging, it is indeed possible to alleviate age‐related diseases and dysfunction and extend longevity. This field of geroscience thus aims to prevent the development of multiple disorders with age, thereby extending healthspan, with the reduction of morbidity toward the end of life. Experts in the field have made remarkable advancements in understanding the mechanisms underlying biological aging and identified ways to target aging pathways using both novel agents and repurposed therapies. While geroscience researchers currently face significant barriers in bringing therapies through clinical development, proof‐of‐concept studies, as well as early‐stage clinical trials, are underway to assess the feasibility of drug evaluation and lay a regulatory foundation for future FDA approvals in the future.

Abstract The burden of senescent cells (SnCs), which do not divide but are metabolically active and resistant to death by apoptosis, is increased in older adults and those with chronic diseases. These individuals are also at the greatest risk for morbidity and mortality from SARS‐CoV‐2 infection. SARS‐CoV‐2 complications include cytokine storm and multiorgan failure mediated by the same factors as often produced by SnCs through their senescence‐associated secretory phenotype (SASP). The SASP can be amplified by infection‐related pathogen‐associated molecular profile factors. Senolytic agents, such as Fisetin, selectively eliminate SnCs and delay, prevent, or alleviate multiple disorders in aged experimental animals and animal models of human chronic diseases, including obesity, diabetes, and respiratory diseases. Senolytics are now in clinical trials for multiple conditions linked to SnCs, including frailty; obesity/diabetes; osteoporosis; and cardiovascular, kidney, and lung diseases, which are also risk factors for SARS‐CoV‐2 morbidity and mortality. A clinical trial is underway to test if senolytics decrease SARS‐CoV‐2 progression and morbidity in hospitalized older adults. We describe here a National Institutes of Health‐funded, multicenter, placebo‐controlled clinical trial of Fisetin for older adult skilled nursing facility (SNF) residents who have been, or become, SARS‐CoV‐2 rtPCR‐positive, including the rationale for targeting fundamental aging mechanisms in such patients. We consider logistic challenges of conducting trials in long‐term care settings in the SARS‐CoV‐2 era, including restricted access, consent procedures, methods for obtaining biospecimens and clinical data, staffing, investigational product administration issues, and potential solutions for these challenges. We propose developing a national network of SNFs engaged in interventional clinical trials.

Adipocytes accumulate within and around skeletal muscle with advancing age. Adipose progenitor cells, preadipocytes, are prone to cellular senescence, a state of stable growth arrest associated with the secretion of cytokines, chemokines and matrix remodeling proteins, collectively referred to as the senescence‐associated secretory phenotype (SASP). Considerable evidence implicates the SASP in age‐related tissue inflammation and degeneration; however, its impact on skeletal muscle is poorly understood. Herein, we examined the effects of conditioned media (CM) collected from healthy human (control CM) and senescent by irradiation (senescent CM) preadipocytes on human skeletal muscle cells (SkMCs). Compared to control CM, senescent CM had more than 10‐fold higher concentrations of IL‐6, IL‐1β, IL‐8, PAI‐1, MCP‐1 and TNFα as quantified by a multiplexed antibody‐based assay. Senescent CM increased expression of IL‐6 (5‐fold), IL‐1α (1.6‐fold), IL‐1β (3.5‐fold), MCP‐1 (2.5‐fold), PAI‐1 (1.7‐fold) and MMP‐3 (3‐fold) in differentiated SkMCs relative to control CM as measured by qPCR (all p < 0.05). Senescent CM increased proliferation of undifferentiated SKMCs by 20%; however, inhibited differentiation relative to control CM per the fusion index. Preliminary data further suggest senescent CM compromises insulin‐ and IGF‐1‐mediated activation of Akt. Collectively, our data suggest that the SASP of senescent preadipocytes may drive inflammation in SkMCs and compromise their differentiation and, potentially, response to metabolic an anabolic stimuli. These effects may contribute to age‐related changes in skeletal muscle health and regeneration. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .

Inflammation is a hallmark of aging and a likely contributor to skeletal muscle loss ( sarcopenia ). We investigated whether the inflammatory phenotype of differentiated primary skeletal muscle‐derived cells (SkMDCs) from a 17‐year‐old (young) differed from that of a 68‐year‐old (older) donor. Under basal conditions, expression of genes encoding pro‐inflammatory cytokines was in fact lower in older compared to younger SkMDCs. However, in response to lipopolysaccharide (LPS), these factors were more highly upregulated in older cells. Specifically, in young compared to older SkMDCs, interleukin‐6 (IL‐6) increased 3‐ vs. 9‐fold, tumor necrosis factor‐alpha (TNF‐alpha) increased 11‐ vs. 80‐fold, and IL‐1alpha increased 14‐ vs. 21‐fold (all differences were p < 0.01). Given adipose tissue is a primary source of pro‐inflammatory stimuli and infiltrates skeletal muscle with age, we hypothesized that secreted factors from senescent preadipocytes drive a similar molecular phenotype to LPS. Exposure of SkMDCs to conditioned medium from senescent‐preadipocytes increased expression IL‐6 and TNF‐alpha in young and even more so in older cells (both p < 0.05). Thus, there are inherent differences in the molecular inflammatory phenotype and its response between young and older SkMDCs. These data also suggest that the age‐related accumulation of senescent preadipocytes may contribute to sarcopenia.

Significance Statement Renal artery stenosis (RAS) engenders stenotic-kidney ischemia, dysfunction, and injury, but whether these are mediated by cellular senescence has not been elucidated. INK-ATTAC transgenic mice, high-resolution imaging, and unbiased single-cell RNA sequencing of murine kidneys demonstrated cellular senescence as an important mechanism of progressive injury to epithelial/stromal cells within poststenotic kidneys. Both p16-specific and broad quercetin/dasatinib interventions to blunt senescence improved renal function and structure, underscoring the central role of senescence in the pathogenesis. Furthermore, this mechanism was conserved in human subjects with RAS. These observations reveal new mechanisms that contribute to the pathogenesis of chronic ischemic renal injury, and support the development of senolytic therapy to reduce senescent cell burden and delay renal injury. Background Peripheral vascular diseases may induce chronic ischemia and cellular injury distal to the arterial obstruction. Cellular senescence involves proliferation arrest in response to stress, which can damage neighboring cells. Renal artery stenosis (RAS) induces stenotic-kidney dysfunction and injury, but whether these arise from cellular senescenceand their temporal pattern remain unknown. Methods Chronic renal ischemia was induced in transgenic INK-ATTAC and wild type C57BL/6 mice by unilateral RAS, and kidney function ( in vivo micro-MRI) and tissue damage were assessed. Mouse healthy and stenotic kidneys were analyzed using unbiased single-cell RNA-sequencing. To demonstrate translational relevance, cellular senescence was studied in human stenotic kidneys. Results Using intraperitoneal AP20187 injections starting 1, 2, or 4 weeks after RAS, selective clearance of cells highly expressing p16 Ink4a attenuated cellular senescence and improved stenotic-kidney function; however, starting treatment immediately after RAS induction was unsuccessful. Broader clearance of senescent cells, using the oral senolytic combination dasatinib and quercetin, in C57BL/6 RAS mice was more effective in clearing cells positive for p21 ( Cdkn1a ) and alleviating renal dysfunction and damage. Unbiased, single-cell RNA sequencing in freshly dissociated cells from healthy and stenotic mouse kidneys identified stenotic-kidney epithelial cells undergoing both mesenchymal transition and senescence. As in mice, injured human stenotic kidneys exhibited cellular senescence, suggesting this process is conserved. Conclusions Maladaptive tubular cell senescence, involving upregulated p16 ( Cdkn2a ), p19 ( Cdkn2d ), and p21 ( Cdkn1a ) expression, is associated with renal dysfunction and injury in chronic ischemia. These findings support development of senolytic strategies to delay chronic ischemic renal injury.

Introduction: Senescent cells with essentially irreversible loss of replicative potential accumulate in various tissues with aging. Cellular senescence is associated with a pro‐inflammatory senescence‐associated secretory phenotype which the Janus‐associated kinase (JAK) pathway plays a role in. We hypothesized that senescent preadipocytes contribute to age‐related dysfunction of healthy cells. Methods: Preadipocytes from lean subjects were X‐irradiated to induce senescence. We co‐cultured non‐senescent cells with senescent cells or cultured them in conditioned medium (CM) from senescent preadipocytes to test the inflammation, differentiation and insulin sensitivity. Macrophage migration in response to CM was assayed as well. Moreover, we treated old or obese mice with Ruxolitinib, a JAK1/2 inhibitor, to test the beneficial effect. Results: The adipogenesis of preadipocytes was impaired by exposure to senescent cells. Macrophage migration was greater in response to senescent preadipocytes. Inflammation was increased in both adipose tissue and preadipocytes exposed to CM from senescent cells. All these adverse effect could be partially rescued by JAK inhibitor. In addition, senescent preadipocytes inhibited insulin‐induced phosphor‐AKT in adipose tissue and HepG2 cells. Insulin‐induced glycogen synthesis was also inhibited in HepG2 cells. Two months of Ruxolitinib treatment reduced inflammation, reversed age‐related lipodystrophy and improved physical function in frail, 24 month old mice. It also improved insulin sensitivity in diet‐induced obese mice. Conclusion: Senescent preadipocytes contribute to age‐related impairment in adipogenesis, fat tissue inflammation, and insulin resistance. Grant Funding Source : supported by NIH AG13925 and Robert and Arlene Kogod Center on Aging in Mayo Clinic.

Nutrient excess and physical inactivity are primary drivers of numerous age‐related chronic diseases. A growing body of evidence has implicated cellular senescence, a process in which cells lose the ability to divide and concurrently damage neighboring cells by the factors they secrete, as an underlying mechanism of aging and its associated conditions. However, the impact of lifestyle choices on the age‐associated accumulation of senescent cells remains poorly understood. Thus, we examined the effects of a fast food diet (FFD) and a voluntary exercise intervention on multiple parameters of healthspan and cellular senescence using a transgenic mouse model that expresses green fluorescent protein (GFP) in p16 Ink4a ‐positive senescent cells. As expected, 12‐month old male mice that had received the FFD diet for 5 months demonstrated significantly greater gains in body weight and fat mass compared to mice fed a standard chow diet. Moreover, mice fed the FFD exhibited hyperinsulinemia, impaired glucose tolerance, reduced exercise capacity, and diastolic dysfunction. Impressively, providing an exercise wheel (26.9 ± 6.9 km/wk) in parallel with the FFD nullified the deleterious effects of the FFD on body weight and composition, whole‐body metabolism, physical performance and cardiac function. Strikingly, our data further suggest that exercise markedly lowers the number of cells in visceral fat depots that express GFP and stain positively for senescent‐associated β‐galactosidase; markers of senescence, that are highly abundant in sedentary mice fed the FFD. Correspondingly, FFD‐induced expression of senescence‐associated inflammatory factors was also diminished by the exercise intervention. Collectively, our data suggest that lifestyle choices may affect the abundance of senescence cells with advancing age, and exercise may partly mediate its salutary effects on multiple organ systems by preventing their accumulation and/or mediating their removal.

AbstractObjectiveAdipose tissue (AT) senescence is associated with AT dysfunction in rodents, but little is known about human AT senescence. The study goal was to define the distribution of senescent cells in two subcutaneous depots and understand relationships with adiposity and inflammation.MethodsSixty‐three volunteers (48 females) underwent abdominal and femoral subcutaneous fat biopsies. Fat cell size, senescent cells using senescence‐associated β‐galactosidase staining per 100 nucleated cells (percentage), and mRNA expression of four cytokines were measured.ResultsThere was a larger proportion of senescent cells in femoral than abdominal subcutaneous AT (mean difference 1.6% [95% CI: 0.98%‐2.3%], p < 0.001), and the percentage of femoral AT senescent cells was greater in women than men (median 3.9% vs. 2.1%, p < 0.01). There was a positive correlation between senescence and fat cell size in abdominal (rs = 0.44, p < 0.001) and femoral (rs = 0.35, p = 0.007) AT depots. Abdominal AT tumor necrosis factor alpha (rs = 0.49, p < 0.01) and interleukin‐1β (rs = 0.44, p = 0.01) expression was positively correlated with abdominal, but not femoral, AT senescence.ConclusionsIn human subcutaneous AT, there are more senescent cells in femoral than abdominal depots; abdominal AT senescent cells are more associated with inflammatory signals than femoral AT senescent cells.

Senescent cells exacerbate COVID-19 Cellular senescence is a state elicited in response to stress signals and is associated with a damaging secretory phenotype. The number of senescent cells increases with advanced age and this in turn drives age-related diseases. Camell et al. show that senescent cells have an amplified inflammatory response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (see the Perspective by Cox and Lord). This response is communicated to nonsenescent cells, suppressing viral defense mechanisms and increasing the expression of viral entry proteins. In old mice infected with a SARS-CoV-2–related virus, treatment with senolytics to reduce the senolytic cell burden reduced mortality and increased antiviral antibodies. Science , abe4832, this issue p. eabe4832 ; see also abi4474, p. 281