17α-Estradiol alleviates high-fat diet-induced inflammatory and metabolic dysfunction in skeletal muscle of male and female mice.

17α-estradiol (17α-E2) is a naturally occurring nonfeminizing diastereomer of 17ß-estradiol that has life span-extending effects in rodent models. To date, studies of the systemic and tissue-specific benefits of 17α-E2 have largely focused on the liver, brain, and white adipose tissue with far less focus on skeletal muscle. Skeletal muscle has an important role in metabolic and age-related disease. Therefore, this study aimed to determine whether 17α-E2 treatment has positive, tissue-specific effects on skeletal muscle during a high-fat feeding. We hypothesized that male, but not female, mice, would benefit from 17α-E2 treatment during a high-fat diet (HFD) with changes in the mitochondrial proteome to support lipid oxidation and subsequent reductions in diacylglycerol (DAG) and ceramide content. To test this hypothesis, we used a multiomics approach to determine changes in lipotoxic lipid intermediates, metabolites, and proteins related to metabolic homeostasis. Unexpectedly, we found that 17α-E2 had marked, but different, beneficial effects within each sex. In male mice, we show that 17α-E2 alleviates HFD-induced metabolic detriments of skeletal muscle by reducing the accumulation of diacylglycerol (DAG), and inflammatory cytokine levels, and altered the abundance of most of the proteins related to lipolysis and ß-oxidation. Similar to male mice, 17α-E2 treatment reduced fat mass while protecting muscle mass in female mice but had little muscle inflammatory cytokine levels. Although female mice were resistant to HFD-induced changes in DAGs, 17α-E2 treatment induced the upregulation of six DAG species. In female mice, 17α-E2 treatment changed the relative abundance of proteins involved in lipolysis, ß-oxidation, as well as structural and contractile proteins but to a smaller extent than male mice. These data demonstrate the metabolic benefits of 17α-E2 in skeletal muscle of male and female mice and contribute to the growing literature of the use of 17α-E2 for multi tissue health span benefits.NEW & NOTEWORTHY Using a multiomics approach, we show that 17α-E2 alleviates HFD-induced metabolic detriments in skeletal muscle by altering bioactive lipid intermediates, inflammatory cytokines, and the abundance of proteins related to lipolysis and muscle contraction. The positive effects of 17α-E2 in skeletal muscle occur in both sexes but differ in their outcome.

Reproductive Ageing: Inflammation, immune cells, and cellular senescence in the aging ovary.

In brief: Recent reports suggest a relationship between ovarian inflammation and functional declines, although it remains unresolved if ovarian inflammation is the cause or consequence of ovarian aging. In this review, we compile the available literature in this area and point to several current knowledge gaps that should be addressed through future studies. Abstract: Ovarian aging results in reduced fertility, disrupted endocrine signaling, and an increased burden of chronic diseases. The factors contributing to the natural decline of ovarian follicles throughout reproductive life are not fully understood. Nevertheless, local inflammation may play an important role in driving ovarian aging. Inflammation progressively rises in aged ovaries during the reproductive window, potentially affecting fertility. In addition to inflammatory markers, recent studies show an accumulation of specific immune cell populations in aging ovaries, particularly lymphocytes. Other hallmarks of the aging ovary include the formation and accumulation of multinucleated giant cells, increased collagen deposition, and increased markers of cellular senescence. Collectively, these changes significantly impact the quantity and quality of ovarian follicles and oocytes. This review explores recent literature on the alterations associated with inflammation, fibrosis, cell senescence, and the accumulation of immune cells in the aging ovary.

17α-estradiol, a lifespan-extending compound, attenuates liver fibrosis by modulating collagen turnover rates in male mice.

Estrogen signaling is protective against chronic liver diseases, although men and a subset of women are contraindicated for chronic treatment with 17ß-estradiol (17ß-E2) or combination hormone replacement therapies. We sought to determine if 17α-estradiol (17α-E2), a naturally occurring diastereomer of 17ß-E2, could attenuate liver fibrosis. We evaluated the effects of 17α-E2 treatment on collagen synthesis and degradation rates using tracer-based labeling approaches in male mice subjected to carbon tetrachloride (CCl4)-induced liver fibrosis. We also assessed the effects of 17α-E2 on markers of hepatic stellate cell (HSC) activation, collagen cross-linking, collagen degradation, and liver macrophage content and polarity. We found that 17α-E2 significantly reduced collagen synthesis rates and increased collagen degradation rates, which was mirrored by declines in transforming growth factor ß1 (TGF-ß1) and lysyl oxidase-like 2 (LOXL2) protein content in liver. These improvements were associated with increased matrix metalloproteinase 2 (MMP2) activity and suppressed stearoyl-coenzyme A desaturase 1 (SCD1) protein levels, the latter of which has been linked to the resolution of liver fibrosis. We also found that 17α-E2 increased liver fetuin-A protein, a strong inhibitor of TGF-ß1 signaling, and reduced proinflammatory macrophage activation and cytokines expression in the liver. We conclude that 17α-E2 reduces fibrotic burden by suppressing HSC activation and enhancing collagen degradation mechanisms. Future studies will be needed to determine if 17α-E2 acts directly in hepatocytes, HSCs, and/or immune cells to elicit these benefits.

The role of sex in the innate and adaptive immune environment of metastatic colorectal cancer.

BACKGROUND: Women with colorectal cancer (CRC) have a significant survival advantage over men. Sex influences on the tumour microenvironment (TME) are not well characterised, despite the importance of immune response in CRC. We hypothesised that sex-divergent immune responses could contribute to survival. METHODS: Using a murine model of metastatic CRC, we examined T cells, macrophages, and cytokines locally and systemically. TME and serum cytokines were measured by multiplex bead-based arrays, while FCA was used to identify cells and phenotypes. IHC provided spatial confirmation of T cell infiltration. RESULTS: Females had increased survival and T cell infiltration. CD8, CD4 and Th2 populations correlated with longer survival. Males had increased serum levels of chemokines and inflammation-associated cytokines. Within the TME, males had lower cytokine levels than females, and a shallower cytokine gradient to the periphery. Female tumours had elevated IL-10+ macrophages, which correlated with survival. CONCLUSIONS: These data demonstrate survival-associated differences in the immune response of males and females to metastatic CRC. Females showed changes in cytokine production accompanied by increased immune cell populations, biased toward Th2-axis phenotypes. Key differences in the immune response to CRC correlated with survival in this model. These differences support a multi-faceted shift across the TME.

Physiological Aging: Links Among Adipose Tissue Dysfunction, Diabetes, and Frailty.

Advancing age is associated with progressive declines in physiological function that lead to overt chronic disease, frailty, and eventual mortality. Importantly, age-related physiological changes occur in cellularity, insulin-responsiveness, secretory profiles, and inflammatory status of adipose tissue, leading to adipose tissue dysfunction. Although the mechanisms underlying adipose tissue dysfunction are multifactorial, the consequences result in secretion of proinflammatory cytokines and chemokines, immune cell infiltration, an accumulation of senescent cells, and an increase in senescence-associated secretory phenotype (SASP). These processes synergistically promote chronic sterile inflammation, insulin resistance, and lipid redistribution away from subcutaneous adipose tissue. Without intervention, these effects contribute to age-related systemic metabolic dysfunction, physical limitations, and frailty. Thus adipose tissue dysfunction may be a fundamental contributor to the elevated risk of chronic disease, disability, and adverse health outcomes with advancing age.

JAK inhibition alleviates the cellular senescence-associated secretory phenotype and frailty in old age.

Chronic, low grade, sterile inflammation frequently accompanies aging and age-related diseases. Cellular senescence is associated with the production of proinflammatory chemokines, cytokines, and extracellular matrix (ECM) remodeling proteases, which comprise the senescence-associated secretory phenotype (SASP). We found a higher burden of senescent cells in adipose tissue with aging. Senescent human primary preadipocytes as well as human umbilical vein endothelial cells (HUVECs) developed a SASP that could be suppressed by targeting the JAK pathway using RNAi or JAK inhibitors. Conditioned medium (CM) from senescent human preadipocytes induced macrophage migration in vitro and inflammation in healthy adipose tissue and preadipocytes. When the senescent cells from which CM was derived had been treated with JAK inhibitors, the resulting CM was much less proinflammatory. The administration of JAK inhibitor to aged mice for 10 wk alleviated both adipose tissue and systemic inflammation and enhanced physical function. Our findings are consistent with a possible contribution of senescent cells and the SASP to age-related inflammation and frailty. We speculate that SASP inhibition by JAK inhibitors may contribute to alleviating frailty. Targeting the JAK pathway holds promise for treating age-related dysfunction.

Retinal gene expression responses to aging are sexually divergent.

PURPOSE: Sex and age are critical factors in a variety of retinal diseases but have garnered little attention in preclinical models. The current lack of knowledge impairs informed decision making regarding inclusion and design of studies that incorporate both sexes and/or the effects of aging. The goal of this study was to examine normative mouse retina gene expression in both sexes and with advancing age. METHODS: Retinal gene expression in female and male C57BL/6JN mice at 3 months and 24 months of age were compared for sex differences and aging responses through whole transcriptome microarray analysis. Sex differences and age-related changes were examined in the context of cellular pathways and processes, regulatory patterns, and cellular origin, as well as for overlap with described changes in retinal disease models. Selected age and sex differences were confirmed with quantitative PCR. RESULTS: Age-related gene expression changes demonstrated commonalities and sexually divergent responses. Several cellular pathways and processes, especially inflammation-related, are affected and were over-represented in fibroblast, microglial, and ganglion cell-specific genes. Lifelong, and age-dependent, sex differences were observed and were over-represented in fibroblast-specific genes. Age and sex differences were also observed to be regulated in models of diabetic retinopathy, glaucoma, and other diseases. CONCLUSIONS: These findings demonstrate that most age-related changes in retinal gene expression are sexually divergent and that there are significant sex differences in gene expression throughout the lifespan. These data serve as a resource for vision researchers seeking to include sex and age as factors in their preclinical studies.

Development of a high-throughput method for real-time assessment of cellular metabolism in intact long skeletal muscle fibre bundles.

KEY POINTS: We developed a method that allows for real-time assessment of cellular metabolism in isolated, intact long skeletal muscle fibre bundles from adult mice. This method can be used to study changes in mitochondrial function and fuel utilisation in live skeletal muscle fibre bundles. Our method enables flexibility in experimental design and high-throughput assessment of mitochondrial parameters in isolated skeletal muscle fibre bundles. Extensor digitorum longus (EDL) fibre bundles obtained from chronic high-fat diet fed mice had lower basal oxygen consumption under FCCP-induced maximal respiration, when compared to control chow-fed mice. EDL fibre bundles obtained from chronic high-fat diet fed mice had enhanced mitochondrial oxidation capacity under FCCP-induced maximal respiration, when compared to control chow-fed mice. ABSTRACT: Metabolic dysfunction in skeletal muscle contributes to the aetiology and development of muscle diseases and metabolic diseases. As such, assessment of skeletal muscle cellular bioenergetics provides a powerful means to understand the role of skeletal muscle metabolism in disease and to identify possible therapeutic targets. Here, we developed a method that allows for the real-time assessment of cellular respiration in intact skeletal muscle fibre bundles obtained from the extensor digitorum longus (EDL) muscle of adult mice. Using this method, we assessed the contribution of ATP turnover and proton leak to basal mitochondrial oxygen consumption rate (OCR). Our data demonstrate that the mitochondria in EDL fibres are loosely coupled. Moreover, in the presence of carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP), we show that palmitate exposure induced comparable peak OCR and higher total OCR in EDL fibre bundles when compared to pyruvate exposure, suggesting that fatty acids might be a more sustainable fuel source for skeletal muscle when mitochondria are driven to maximal respiration. Application of this method to EDL fibre bundles obtained from chronic high-fat diet fed mice revealed lower basal OCR and enhanced mitochondrial oxidation capacity in the presence of FCCP when compared to the chow-diet fed control mice. By using a 96-well microplate format, our method provides a flexible and efficient platform to investigate mitochondrial parameters of intact skeletal muscle fibres obtained from adult mice.

TRAIL receptor deletion in mice suppresses the inflammation of nutrient excess.

BACKGROUND & AIMS: Low-grade chronic inflammation is a cardinal feature of the metabolic syndrome, yet its pathogenesis is not well defined. The purpose of this study was to examine the role of TRAIL receptor (TR) signaling in the pathogenesis of obesity-associated inflammation using mice with the genetic deletion of TR. METHODS: TR knockout (TR(-/-)) mice and their littermate wild-type (WT) mice were fed a diet high in saturated fat, cholesterol and fructose (FFC) or chow. Metabolic phenotyping, liver injury, and liver and adipose tissue inflammation were assessed. Chemotaxis and activation of mouse bone marrow-derived macrophages (BMDMϕ) was measured. RESULTS: Genetic deletion of TR completely repressed weight gain, adiposity and insulin resistance in FFC-fed mice. Moreover, TR(-/-) mice suppressed steatohepatitis, with essentially normal serum ALT, hepatocyte apoptosis and liver triglyceride accumulation. Gene array data implicated inhibition of macrophage-associated hepatic inflammation in the absence of the TR. In keeping with this, there was diminished accumulation and activation of inflammatory macrophages in liver and adipose tissue. TR(-/-) BMDMϕ manifest reduced chemotaxis and diminished activation of nuclear factor-κ B signaling upon activation by palmitate and lipopolysaccharide. CONCLUSIONS: These data advance the concept that macrophage-associated hepatic and adipose tissue inflammation of nutrient excess requires TR signaling.

Systemic metabolic benefits of 17α-estradiol are not exclusively mediated by ERα in glutamatergic or GABAergic neurons.

17α-Estradiol (17αE2), a less-feminising enantiomer of 17ß-estradiol, has been shown to prolong lifespan and improve metabolic health in a sex-specific manner in male, but not in female mice. Recent studies have demonstrated the pivotal role of estrogen receptor α (ERα) in mediating the effects of 17αE2 on metabolic health. However, the specific tissues and/or neuronal signalling pathways that 17αE2 acts through remain to be elucidated. ERα expression in glutamatergic and GABAergic neurons (principal excitatory and inhibitory neurons respectively) in the hypothalamus is essential for estradiol signalling. Therefore, we hypothesised that knocking out ERα from one of these neuronal populations would attenuate the established beneficial metabolic effects of 17αE2 in male mice exposed to a high fat diet. To test this hypothesis we used two established brain specific ERα KO models, targeting either glutamatergic or GABAergic neurons (Vglut2/Vgat-ERαKO). We show that both of these ERα KO models exhibit a strong reduction in ERα expression in the arcuate nucleus of the hypothalamus, a control centre for metabolic regulation. Deletion of ERα from GABAergic neurons significantly diminished the effect of 17αE2 on body weight relative to controls, although these animals still show metabolic benefits with 17αE2 treatment. The response to 17αE2 was unaffected by ERα deletion in glutamatergic neurons. Our results support a benefit of 17αE2 treatment in protection against metabolic dysfunction, but these effects do not depend on exclusive ERα expression in glutamatergic and GABAergic neurons and persist when ERα expression is strongly reduced in the arcuate nucleus of the hypothalamus.

Aged mice exhibit faster acquisition of intravenous opioid self-administration with variable effects on intake.

Although opioid abuse is more prevalent in young individuals, opioid use, overdose, and use disorders continue to climb at a rapid rate among the elderly. Little is known about abuse potential in a healthy aged population, in part due to technical and logistical difficulties testing intravenous self-administration in aged rodents. The goal of this study was to address the critical gap in the literature regarding age-dependent differences in opioid (remifentanil and fentanyl) self-administration between old and young mice. Male and female mice were grouped into young (mean: 19 weeks) and old (mean: 101 weeks), and were trained to self-administer intravenous fentanyl or remifentanil in daily sessions. In both old and young mice, acquisition, intake, and cue-responding after forced abstinence were measured for both drugs, and a dose-response curve (remifentanil) and dose-escalation (fentanyl) were conducted. Surprisingly, old mice learned to self-administer both remifentanil and fentanyl faster and more accurately than young mice. Baseline intake of remifentanil was also substantially greater in old mice compared to their young counterparts; however, we did not see increased intake of fentanyl with age at either dose tested. Further, compared to young mice, the old mice showed a greater incubation of responding for cues previously associated with remifentanil after a forced abstinence, but again this was not observed with fentanyl. Together these data suggest that an aged population may have an increased drug-abuse vulnerability for opioids compared to young counterparts and underscore the importance of future work on mechanisms responsible for this increased vulnerability.

The role of cellular senescence in ovarian aging.

This review explores the relationship between ovarian aging and senescent cell accumulation, as well as the efficacy of senolytics to improve reproductive longevity. Reproductive longevity is determined by the age-associated decline in ovarian reserve, resulting in reduced fertility and eventually menopause. Cellular senescence is a state of permanent cell cycle arrest and resistance to apoptosis. Senescent cells accumulate in several tissues with advancing age, thereby promoting chronic inflammation and age-related diseases. Ovaries also appear to accumulate senescent cells with age, which might contribute to aging of the reproductive system and whole organism through SASP production. Importantly, senolytic drugs can eliminate senescent cells and may present a potential intervention to mitigate ovarian aging. Herein, we review the current literature related to the efficacy of senolytic drugs for extending the reproductive window in mice.

Age- and sex- divergent translatomic responses of the mouse retinal pigmented epithelium.

Aging is the main risk factor for age-related macular degeneration (AMD), a retinal neurodegenerative disease that leads to irreversible blindness, particularly in people over 60 years old. Retinal pigmented epithelium (RPE) atrophy is an AMD hallmark. Genome-wide chromatin accessibility, DNA methylation, and gene expression studies of AMD and control RPE demonstrate epigenomic/transcriptomic changes occur during AMD onset and progression. However, mechanisms by which molecular alterations of normal aging impair RPE function and contribute to AMD pathogenesis are unclear. Here, we specifically interrogate the RPE translatome with advanced age and across sexes in a novel RPE reporter mouse model. We find differential age- and sex- associated transcript expression with overrepresentation of pathways related to inflammation in the RPE. Concordant with impaired RPE function, the phenotypic changes in the aged translatome suggest that aged RPE becomes immunologically active, in both males and females, with some sex-specific signatures, which supports the need for sex representation for in vivo studies.

A pilot study evaluating dosing tolerability of 17α-estradiol in male common marmosets (Callithrix jacchus).

17α-estradiol extends healthspan and lifespan in male mice without significant feminization or deleterious effects on reproductive function, making it a candidate for human translation. However, studies in animal models that more accurately replicate human physiology are necessary to establish 17α-estradiol dosing standards for clinical trials. This study evaluated the tolerability of 17α-estradiol treatment in the common marmoset over a short treatment duration. We found that male marmosets tolerated two dosing regimens (0.37-0.47 or 0.62-0.72 mg/kg/day) as evidenced by the absence of gastrointestinal distress, changes in vital signs, or overall health conditions. 17α-estradiol treatment mildly decreased body mass, adiposity, and glycosylated hemoglobin, although these changes were not statistically significant in most instances. However, neither dose of 17α-estradiol elicited feminization in our study, thereby suggesting that optimized dosing regimens may provide health benefits without feminization in primates. Additional studies are needed to determine if longer duration treatments would also be nonfeminizing and elicit significant health benefits, which would aid in developing dosing regimens targeting healthy aging in humans.

Senolytic treatment fails to improve ovarian reserve or fertility in female mice.

Senescent cell number increases with age in different tissues, leading to greater senescent cell load, proinflammatory stress, and tissue dysfunction. In the current study, we tested the efficacy of senolytic drugs to reduce ovarian senescence and improve fertility in reproductive age female mice. In the first experiment, 1-month-old C57BL/6 female mice were treated every other week with D + Q (n = 24) or placebo (n = 24). At 3 and 6 months of age, female mice were mated with untreated males to evaluate pregnancy rate and litter size. In the second experiment, 6-month-old C57BL/6 female mice were treated monthly with D + Q (n = 30), fisetin (n = 30), or placebo (n = 30). Females were treated once a month until 11 months of age, then they were mated with untreated males for 30 days to evaluate pregnancy rate and litter size. In the first experiment, D + Q treatment did not affect pregnancy rate (P = 0.68), litter size (P = 0.58), or ovarian reserve (P > 0.05). Lipofuscin staining was lower in females treated with D + Q (P = 0.04), but expression of senescence genes in ovaries was similar. In the second experiment, D + Q or fisetin treatment also did not affect pregnancy rate (P = 0.37), litter size (P = 0.20), or ovarian reserve (P > 0.05). Lipofuscin staining (P = 0.008) and macrophage infiltration (P = 0.002) was lower in fisetin treated females. Overall, treatment with D + Q or fisetin did not affect ovarian reserve or fertility but did decrease some senescence markers in the ovary.

A single-cell atlas of the aging mouse ovary.

Ovarian aging leads to diminished fertility, dysregulated endocrine signaling and increased chronic disease burden. These effects begin to emerge long before follicular exhaustion. Female humans experience a sharp decline in fertility around 35 years of age, which corresponds to declines in oocyte quality. Despite a growing body of work, the field lacks a comprehensive cellular map of the transcriptomic changes in the aging mouse ovary to identify early drivers of ovarian decline. To fill this gap we performed single-cell RNA sequencing on ovarian tissue from young (3-month-old) and reproductively aged (9-month-old) mice. Our analysis revealed a doubling of immune cells in the aged ovary, with lymphocyte proportions increasing the most, which was confirmed by flow cytometry. We also found an age-related downregulation of collagenase pathways in stromal fibroblasts, which corresponds to rises in ovarian fibrosis. Follicular cells displayed stress-response, immunogenic and fibrotic signaling pathway inductions with aging. This report provides critical insights into mechanisms responsible for ovarian aging phenotypes. The data can be explored interactively via a Shiny-based web application.

Effects of calorie, protein, and branched chain amino acid restriction on ovarian aging in mice.

Calorie restriction (CR) is an intervention that promotes longevity and preserves the ovarian reserve. Some studies have observed that the positive impacts of CR can be linked to restriction of protein (PR) and branched-chain amino acids (BCAAs) independent of calorie intake. The aim of this study was to compare the effects of protein and BCAA restriction to 30% CR on the ovarian reserve of female mice. For this, 3 month-old C57BL/6 female mice (n = 35) were randomized into four groups for four months dietary interventions including: control group (CTL; n = 8), 30% CR (CR; n = 9), protein restriction (PR; n = 9) and BCAA restriction (BCAAR; n = 9). Body mass gain, body composition, food intake, serum levels of BCAAs, ovarian reserve and estrous cyclicity were evaluated. We observed that CR, protein and BCAA restriction prevented weight gain and changed body composition compared to the CTL group. The BCAA restriction did not affect the ovarian reserve, while both PR and CR prevented activation of primordial follicles. This prevention occurred in PR group despite the lack of reduction of calorie intake compared to CTL group, and CR did not reduce protein intake in levels similar to the PR group. BCAA restriction resulted in increased calorie intake compared to CTL and PR mice, but only PR reduced serum BCAA levels compared to the CTL group. Our data indicates that PR has similar effects to CR on the ovarian reserve, whereas BCAA restriction alone did not affect it.

Metformin treatment results in distinctive skeletal muscle mitochondrial remodeling in rats with different intrinsic aerobic capacities.

The rationale for the use of metformin as a treatment to slow aging was largely based on data collected from metabolically unhealthy individuals. For healthspan extension metformin will also be used in periods of good health. To understand potential context specificity of metformin treatment on skeletal muscle, we used a rat model (HCR/LCR) with a divide in intrinsic aerobic capacity. Outcomes of metformin treatment differed based on baseline intrinsic mitochondrial function, oxidative capacity of the muscle (gastroc vs soleus), and the mitochondrial population (IMF vs SS). Metformin caused lower ADP-stimulated respiration in LCRs, with less of a change in HCRs. However, a washout of metformin resulted in an unexpected doubling of respiratory capacity in HCRs. These improvements in respiratory capacity were accompanied by mitochondrial remodeling that included increases in protein synthesis and changes in morphology. Our findings raise questions about whether the positive findings of metformin treatment are broadly applicable.

Metformin treatment results in distinctive skeletal muscle mitochondrial remodeling in rats with different intrinsic aerobic capacities.

The rationale for the use of metformin as a treatment to slow aging was largely based on data collected from metabolically unhealthy individuals. For healthspan extension metformin will also be used in periods of good health. To understand the potential context specificity of metformin treatment on skeletal muscle, we used a rat model (high-capacity runner/low-capacity runner [HCR/LCR]) with a divide in intrinsic aerobic capacity. Outcomes of metformin treatment differed based on baseline intrinsic mitochondrial function, oxidative capacity of the muscle (gastroc vs soleus), and the mitochondrial population (intermyofibrillar vs. subsarcolemmal). Metformin caused lower ADP-stimulated respiration in LCRs, with less of a change in HCRs. However, a washout of metformin resulted in an unexpected doubling of respiratory capacity in HCRs. These improvements in respiratory capacity were accompanied by mitochondrial remodeling that included increases in protein synthesis and changes in morphology. Our findings raise questions about whether the positive findings of metformin treatment are broadly applicable.

The retrotransposon-derived capsid genes PNMA1 and PNMA4 maintain reproductive capacity.

The human genome contains 24 gag-like capsid genes derived from deactivated retrotransposons conserved among eutherians. Although some of their encoded proteins retain the ability to form capsids and even transfer cargo, their fitness benefit has remained elusive. Here we show that the gag-like genes PNMA1 and PNMA4 support reproductive capacity during aging. Analysis of donated human ovaries shows that expression of both genes declines normally with age, while several PNMA1 and PNMA4 variants identified in genome-wide association studies are causally associated with low testosterone, altered puberty onset, or obesity. Six-week-old mice lacking either Pnma1 or Pnma4 are indistinguishable from wild-type littermates, but by six months the mutant mice become prematurely subfertile, with precipitous drops in sex hormone levels, gonadal atrophy, and abdominal obesity; overall they produce markedly fewer offspring than controls. These findings expand our understanding of factors that maintain human reproductive health and lend insight into the domestication of retrotransposon-derived genes.