Longevity In Mice Research Articles

Intermediate alleles of HTT: A new pathway in longevity

Centenarians are the best example of successful aging, reaching extreme longevity escaping age-related diseases. Genome sequencing studies provided evidence for genetic factors linked to heathy long life, including genes related to age-dependent diseases. HTT (Huntingtin) gene is linked to Huntington's Disease, but also associated to longevity in capuchins and mice. HTT Intermediate alleles (IAs) are defined as CAG repeat expansion between 27 and 35. According to recent data IAs might increase Alzheimer's Disease risk, but also might have a neuroprotective effect and can confer an advantage in brain development. Here, we investigated, for the first time, the possible implication of HTT IAs in extreme longevity and their possible association in cognitive decline. We analysed the distribution of IAs in Italian Centenarians (n = 143) and compared with pathological controls with cognitive decline (n = 232, including 80 Alzheimer's Disease, 78 Frontotemporal Dementia and 74 Subjective Cognitive Decline patients) and healthy controls (n = 104). Our data show a statistically significant higher frequency of IAs in Centenarians with respect to pathological controls with cognitive decline (p = .031; OR = 2.3097 95% CI 1.0591 to 5.0371), with a percentage of 11.2 respect to 5.4 respectively. The highest presence of IAs in Centenarians confirms and extends in humans a possible implication of HTT gene in exceptional lifespan and in brain development with a neuroprotective effect.

Multi-omic rejuvenation of naturally aged tissues by a single cycle of transient reprogramming.

The expression of the pluripotency factors OCT4, SOX2, KLF4, and MYC (OSKM) can convert somatic differentiated cells into pluripotent stem cells in a process known as reprogramming. Notably, partial and reversible reprogramming does not change cell identity but can reverse markers of aging in cells, improve the capacity of aged mice to repair tissue injuries, and extend longevity in progeroid mice. However, little is known about the mechanisms involved. Here, we have studied changes in the DNA methylome, transcriptome, and metabolome in naturally aged mice subject to a single period of transient OSKM expression. We found that this is sufficient to reverse DNA methylation changes that occur upon aging in the pancreas, liver, spleen, and blood. Similarly, we observed reversion of transcriptional changes, especially regarding biological processes known to change during aging. Finally, some serum metabolites and biomarkers altered with aging were also restored to young levels upon transient reprogramming. These observations indicate that a single period of OSKM expression can drive epigenetic, transcriptomic, and metabolomic changes toward a younger configuration in multiple tissues and in the serum.

Abolishing the prelamin A ZMPSTE24 cleavage site leads to progeroid phenotypes with near-normal longevity in mice.

Prelamin A is a farnesylated precursor of lamin A, a nuclear lamina protein. Accumulation of the farnesylated prelamin A variant progerin, with an internal deletion including its processing site, causes Hutchinson-Gilford progeria syndrome. Loss-of-function mutations in ZMPSTE24, which encodes the prelamin A processing enzyme, lead to accumulation of full-length farnesylated prelamin A and cause related progeroid disorders. Some data suggest that prelamin A also accumulates with physiological aging. Zmpste24-/- mice die young, at ∼20 wk. Because ZMPSTE24 has functions in addition to prelamin A processing, we generated a mouse model to examine effects solely due to the presence of permanently farnesylated prelamin A. These mice have an L648R amino acid substitution in prelamin A that blocks ZMPSTE24-catalyzed processing to lamin A. The LmnaL648R/L648R mice express only prelamin and no mature protein. Notably, nearly all survive to 65 to 70 wk, with ∼40% of male and 75% of female LmnaL648R/L648R mice having near-normal lifespans of 90 wk (almost 2 y). Starting at ∼10 wk of age, LmnaL648R/L648R mice of both sexes have lower body masses than controls. By ∼20 to 30 wk of age, they exhibit detectable cranial, mandibular, and dental defects similar to those observed in Zmpste24-/- mice and have decreased vertebral bone density compared to age- and sex-matched controls. Cultured embryonic fibroblasts from LmnaL648R/L648R mice have aberrant nuclear morphology that is reversible by treatment with a protein farnesyltransferase inhibitor. These novel mice provide a model to study the effects of farnesylated prelamin A during physiological aging.

Characterization of Five Collagenous Biomaterials by SEM Observations, TG-DTA, Collagenase Dissolution Tests and Subcutaneous Implantation Tests.

Collagenous biomaterials that are clinically applied in dentistry have dermis-type and membrane-type, both of which are materials for promoting bone and soft tissue formation. The properties of materials supplied with different types could affect their biodegradation periods. The purpose of this study was to characterize five of these products by four different methods: scanning electron microscopy (SEM) observation, thermogravimetry-differential thermal analysis (TG-DTA), 0.01 wt% collagenase dissolution test, and subcutaneous implantation test in vivo. SEM micrographs revealed that both dermis and membranous materials were fibrous and porous. The membranous materials had higher specific derivative thermal gravimetry (DTG) peak temperatures in TG-DTA at around 320 °C, longer collagenase dissolution time ranging from about 300 to 500 min, and more longevity in mice exceeding 9 weeks than the dermis materials. There existed a correlation between the peak temperature in TG-DTA and the collagenase dissolution time. It was considered that higher cross-link degree among collagen fibrils of the membrane-type collagenous materials might account for these phenomena. The experimental protocol and numerical results obtained could be helpful for selection and future development of fibrous collagenous biomaterials in clinical use.

Continuous and intermittent ketogenic diets initiated in late middle-aged mice improves cognition and motor endurance at advanced age.

Alzheimer's disease (AD) is the most common age-related neurodegenerative disease, and an effective therapeutic strategy that promotes healthy aging may lower age-related risks of AD and ameliorate AD associated cognitive dysfunctions. There has been substantial interest in the application of ketogenic diets (KD) to manage neurological disorders associated with aging, and studies have also shown KDs started from early middle age improved cognition and longevity in mice. Thus, KDs might be used to reduce risk of cognitive declines at old age. KDs started later in life or intermittently administered may be more feasible and promote compliance in an older population. Therefore, this study sought to test if continuous or intermittent KDs started in late-middle-aged mice would improve measures of cognitive and motor function at advanced ages. 18-month-old male C57BL/6J mice were randomly assigned to an isocaloric control (CD), ketogenic (KD), or intermittent ketogenic (IKD, 3 days of KD/week) diet. Continuous or intermittent ketosis was induced at a constant level of energy intake. At 20, 23, and 26 months of age, a panel of behavior tests were performed to assess cognitive (novel object recognition, Y-maze, and Barnes-maze) and motor (grid-wire hang, grip strength, open field, and rearing) functions. Y-maze alternation rate was significantly higher for both IKD and KD mice at 23 months of age and for KD mice at 26 months indicating an improved working memory. 26-month-old KD mice also showed better spatial learning memory as measured by time spent in target quadrant in Barnes-maze. Improved motor endurance and strength was observed in aged IK and KD mice as tested by grid wire hang. A significantly increased composite score of all the behavior parameters was observed in KD mice at 26 months of age, and IK mice showed a trend toward increased score compared to CD mice. KD and IKD initiated in late-middle-aged mice improved cognition and motor endurance in aged mice. KD had a more potent effect on overall heath span in aged mice shown as a higher composite score of all the tests performed with IKD showing results intermediate to other diet groups.

Apheresis Products from Patients with Multiple Myeloma Treated with G-CSF Are a Suitable Source of T Cells for the Production of BCMA-Targeting CAR-T Cells

Background:Autologous chimeric antigen receptor-T (CAR-T) cells that target BCMA (BCMA-CARs) have emerged as a promising treatment for multiple myeloma (MM). Current clinical protocols dictate that BCMA-CAR therapy is only used after patients have repeatedly relapsed. However, at this stage, the immunosuppressive nature of advanced MM and/or side-effects of the previous therapies cause T cell dysfunction and an unfavourable phenotype, such as exhaustion, senescence and loss of early memory cells.An alternative and convenient pool of ‘fitter’ T cells are apheresis products that are routinely collected to obtain progenitor cells for autologous stem cell transplantation (ASCT), an intervention that is often carried out early in MM treatment. However, to mobilise the progenitor cells, patients are treated with G-CSF, which could have negative effects on T cells such as reduce proliferation, impair CD8 + T cell function and induce regulatory T cell (Treg) expansion. Whether this has an effect on the BCMA-CARs generated from these T cells, however, is unknown. Therefore, we aimed to establish whether G-CSF treatment had detrimental effects on T cell phenotype, and moreover, to ascertain whether BCMA-CARs that are generated from these T cells were impaired compared to those produced from T cells prior to G-CSF infusion.Methods:T cells were isolated from the blood of 9 patients with MM before and after 4 days of subcutaneous G-CSF administration (PRE G-CSF and POST G-CSF, respectively) prior to peripheral blood CD34 + cell harvesting for an ASCT as consolidation after first-line induction treatment. Following stimulation with anti-CD3/anti-CD28 beads and IL-2, T cells were transduced with ARI2h, an anti-BCMA CAR produced at our institution that is currently being explored in a clinical trial for relapsed/refractory MM (NCT04309981). Freshly-isolated T cells or expanded ARI2h cells were analysed by flow cytometry for markers of cell identity, activation, dysfunction and memory, or alternatively, challenged with an MM cell line (ARP-1 or U266) and then tested for cytokine production and cytotoxic ability. In addition, PRE and POST G-CSF ARI2h CARs were injected into ARP-1 tumour-bearing mice to assess their in vivo function.Results:Firstly, the phenotype of PRE G-CSF and POST G-CSF T cells, before CAR production, was analysed to identify effects of G-CSF treatment. Interestingly, there were fewer POST G-CSF CD8 + T cells with a stem cell memory (CCR7 +CD45RA +CD95 +) phenotype, but the proportion of naïve (CCR7 +CD45RA +CD95 -) cells and other memory populations was not significantly different. Moreover, POST G-CSF T cells had a lower CD4:CD8 ratio, but did not contain more senescent-like cells or display evidence of pre-activation or increased expression of exhaustion markers. Due to the known effect of G-CSF on CD4 + Treg expansion, the percentage of Tregs was also compared between the PRE G-CSF and POST G-CSF samples, but no difference was observed.Following T-cell activation and CAR transduction, comparable transduction efficiencies and proliferation rates were obtained. Likewise, the in vitro function of PRE G-CSF and POST G-CSF ARI2h cells, as determined by assessing their cytotoxic response to MM cell lines and ability to produce effector molecules such as granzyme B, was similar. To test the in vivo function of ARI2h CAR-T cells expanded from PRE G-CSF and POST G-CSF samples, they were injected into a murine xenograft model of advanced MM. Mice administered with both PRE and POST G-CSF ARI2h cells survived longer than those given untransduced T cells (p=0.015 and p=0.039, respectively), but there was no difference in the longevity of mice between the PRE G-CSF and POST G-CSF groups (p=0.990) (Figure 1). The similarity of the in vitro and in vivo function of PRE and POST G-CSF ARI2h cells was reflected in the phenotype of the CAR-T cells after ex vivo expansion, with cells from both groups displaying equal levels of activation, exhaustion, and importantly for CAR-T cell activity, memory/effector phenotype.Conclusions:The in vitro and in vivo functions of ARI2h CAR-T cells when generated from either PRE G-CSF or POST G-CSF samples were comparable, despite G-CSF administration decreasing the CD8 + stem cell memory pool. Overall, we conclude that T cells from apheresis products, performed to collect G-CSF-mobilised peripheral blood progenitor cells for ASCT, are suitable for BCMA-CAR manufacture. [Display omitted] DisclosuresLozano: Grifols: Honoraria; Terumo BCT: Honoraria, Research Funding; Macopharma: Research Funding. Fernandez de Larrea: BMS: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Takeda: Honoraria, Research Funding; GSK: Honoraria; Sanofi: Consultancy; Janssen: Consultancy, Honoraria, Research Funding.

Hyper-Interleukin-6 Protects Against Renal Ischemic-Reperfusion Injury-A Mouse Model.

Background: Most of the ischemia-reperfusion injury (IR-I) occurs during reperfusion and is mediated by the immune system. In this study we determined whether immunomodulation with hyper-Interleukin-6 (a recombinant designer cytokine composed of interleukin-6 linked to its soluble receptor) is protective against IR-I in mice kidneys.Methods: Hyper-Interleukin-6 (HIL-6) was administered by in vivo plasmid DNA transfection to 10 male mice. Twenty-four hours later, unilateral nephrectomy was done. IR-I immediately followed by closure of the remaining kidney vascular pedicle for 40 min. Seven mice transfected with non-coding control plasmid served as the control group. The functional and morphological effects of IR-I and its effect on mice longevity were explored. This was done by serial blood tests and by histopathology done upon sacrifice of the animals at post-operative day 7.Findings: Mice pretreated with HIL-6 had a mean creatinine level at post-operative day 1 of 35.45 ± 4.03 μmol/l and mean Urea level was 14.18 ± 2.69 mmol/l, whereas mean creatinine was 89.33 ± 69.27 μmol/l (P = 0.025), and mean urea was 38.17 ± 20.77 mmol/l (P = 0.0024) in the control group. Histological changes in the control group included inflammatory infiltration, tubular damage, and architectural distortion. These were not seen in the treatment group. Seven days post-operatively the survival rate of treated mice was 100% compared to 50% in the control group (P = 0.015).Interpretation: In this single kidney mouse model, pretreatment with HIL-6 administration effectively protected against IR-I both morphologically and functionally. Further studies are needed to better understand the mechanism and feasibility of using this immunomodulator.

Young Adult LEW.1WR1 Rats, a Model of Liver FAT10 Overexpression, Develop Insulin Resistance and Fatty Liver With Age

As human lifespan increases, comorbid conditions that impact quality of life have become a serious problem. FAT10 has been identified as a gene that when knocked out, improves age associated metabolic dysfunctions and increased longevity in mice (1). There is increased Fat10 expression in the liver in obesity (2,5). Providing evidence that fat10 expression may be important for triggering the transition to metabolic dysfunction in aging. Adult LEW.1WR1(1WR1) rats have increased body mass without excess abdominal fat mass compared to control rats (3). Yet, it was unclear where the excess mass was stored. We hypothesized that the 1WR1 rat would develop increased liver fat mass as a product of increased insulin resistance in response to increased liver fat10 expression over time. To test this hypothesis, we did insulin tolerance tests(7 weeks & 15 weeks), triglyceride assays, and histological analysis of the liver(23 weeks), in 1WR1 rats(n=7) and Wistar Furth (WF) rats(n=7) on control diets. We analyzed images using histological scoring for nonalcoholic fatty liver disease from the literature (4). We also assessed the slides for Mallory Denk bodies (MBs). The body mass of 1WR1 rats were increased compared to WF rat groups starting from the age of 7 weeks (391.4∓8.572g vs. 271.8∓11.62g; p <0.0006). 1WR1 rats became more insulin resistant with age, the 1WR1 rat group has increased AUC of 7 and 15 week Insulin Tolerance Tests (401.5∓23.54 vs. 245.3∓10.20 7w ITT1; p= 0.0728, 15w ITT2 328.2∓14.86 vs. 217.8∓9.; p <0.0003) compared to WF rats. 1WR1 rats have increased liver mass (11.85g∓0.7699g vs. 7.235g∓0.3864g; p=0.0006) liver triglyceride levels compared to WF rats (192.8∓21.8 mg/mL vs. 130.1∓13.075 mg/mL; p=0.0728). 1WR1 rats have increased steatosis scores(1.857∓0.2608 vs. 1.143∓0.1429;p= 0.0862) yet significantly reduced inflammatory foci level (2∓0.8165 vs. 3∓0;p= 0.007), most 1WR1 hepatocytes are enlarged (ballooned) and contained MBs compared to WF rats suggesting 1WR1 rats have already passed the early inflammation stage. Adult 1WR1 rats developed reduced insulin sensitivity and lipid accumulation in the liver. These data support our hypothesis that 1WR1 rats would develop increased liver fat and impaired insulin resistance in response to aging and show that this process may be inflammation driven. (1) Canaan et al.,PNAS. April 2014; 111 (14): 5313-5318.(2)Vidal et al., FASEB.2020, 34: 1-1,(3) Collins et al., J Endocr Soc. 2019;3(1),(4).Kleiner et al., Hepatology, 2005; 41 (6): 1313–1321,(5).Dali-Youcef et al., Hepatol Commun. 2019;3(9):1205-1220.

Placental Ischemia increases Seizure Susceptibility through Reduced ASIC2a Expression

Introduction Eclampsia is diagnosed in pregnant women and women with preeclampsia, a hypertensive pregnancy disorder, who experience new-onset seizures. Previous studies from our laboratory showed that the rat model of preeclampsia, induced by reducing uterine perfusion pressure (RUPP), displays reduced latency to drug-induced seizures. The factors that contribute to increased seizure sensitivity in pregnancy and preeclampsia are unknown. While acid sensing ion channels (ASIC1a and 3) were shown to be important for reducing seizure longevity and severity, the role of ASIC2a in seizures during pregnancy or RUPP has not been investigated. Thus, we hypothesized that RUPP in mice reduces seizure latency and hippocampal ASIC2a expression. We further hypothesize that mice with reduced ASIC2a (ASIC2a+/-) will display increased seizure severity and duration. Methods Hippocampi from 5 RUPP and 6 Sham C57BL/6 pregnant mice were harvested on gestational day 18.5 and processed for Western blot analysis to assess ASIC2a expression. Seizures were induced using pentylenetetrazol (40 mg/kg, i.p) in RUPP and sham mice (n=5 per group), as well as pregnant wild type (WT) and ASIC2a heterozygous (HET) mice (n=7 or 14 per genotype) and video recorded for 30 minutes. Seizure behavior was scored using a modified Racine scale ranging from 0-7 (0 = no seizure, 7 = respiratory arrest). Seizure severity was separated into two generalized behaviors: petite-mal (1-3), grand-mal (4-7). Data are shown as Mean ± SEM. Results RUPP mice show a trend for reduced latency to seizure activity (63.4 ± 21.3 vs 103.3 ± 5.6s; p=0.068) and had a significant reduction in hippocampal ASIC2a expression (0.5 ± 0.1 in RUPP vs 1 ± 0.2 in Sham; p=0.037). ASIC2a HET mice displayed reduced latency to seizure activity (101 ± 18 vs 248 ± 85s, p=0.033), increased seizure duration (1253 ± 467s vs 823 ± 137s; p=0.019), decreased seizure-free duration (552 ± 125s vs 950 ± 145s; p=0.023), increased grand-mal seizure duration (713 ± 188 vs 18 ± 13s; p=0.004), and higher maximum seizure scores (WT= 3 ± 0; HET= 5 ± 1, p=0.011). ASIC2a HET mice display seizure behavior suggesting increased seizure susceptibility. Conclusion Our findings of reduced hippocampal ASIC2a expression in RUPP mice and increased seizure severity and longevity in pregnant mice with reduced ASIC2a, support the hypothesis that reduced ASIC2a increases seizure sensitivity in the context of placental ischemia.

Symbiotic polyamine metabolism regulates epithelial proliferation and macrophage differentiation in the colon

Intestinal microbiota-derived metabolites have biological importance for the host. Polyamines, such as putrescine and spermidine, are produced by the intestinal microbiota and regulate multiple biological processes. Increased colonic luminal polyamines promote longevity in mice. However, no direct evidence has shown that microbial polyamines are incorporated into host cells to regulate cellular responses. Here, we show that microbial polyamines reinforce colonic epithelial proliferation and regulate macrophage differentiation. Colonisation by wild-type, but not polyamine biosynthesis-deficient, Escherichia coli in germ-free mice raises intracellular polyamine levels in colonocytes, accelerating epithelial renewal. Commensal bacterium-derived putrescine increases the abundance of anti-inflammatory macrophages in the colon. The bacterial polyamines ameliorate symptoms of dextran sulfate sodium-induced colitis in mice. These effects mainly result from enhanced hypusination of eukaryotic initiation translation factor. We conclude that bacterial putrescine functions as a substrate for symbiotic metabolism and is further absorbed and metabolised by the host, thus helping maintain mucosal homoeostasis in the intestine.

Deletion of Nrip1 delays skin aging by reducing adipose-derived mesenchymal stem cells (ADMSCs) senescence, and maintaining ADMSCs quiescence.

Our previous studies found that deletion of nuclear receptor interacting protein 1 (Nrip1) extended longevity in female mice and delayed cell senescence. The current study investigates the role of NRIP1 in regulating functions of adipose-derived mesenchymal stem cells (ADMSCs) and explores the mechanisms of NRIP1 in skin aging. We first verified the skin aging phenotypes in young (6 months) and old (20 months) C57BL/6J (B6) mice and found deletion of Nrip1 can delay skin aging phenotypes, including reduced thickness of dermis and subcutaneous white adipose tissue (sWAT), as well as the accumulation of senescent cells in sWAT. In ADMSCs isolated from sWAT, we found that deletion of Nrip1 could decrease cell proliferation, prevent cell apoptosis, and suppress adipogenesis. Interestingly, deletion of Nrip1 also reduced cell senescence and maintain cell quiescence of ADMSCs. Moreover, the expressions of genes associated with senescence (p21, and p53), inflammation (p65, IL6, and IL1a), and growth factor (mTOR, Igf1) were reduced in Nrip1 knockout ADMSCs, as well as in siNrip1-treated ADMSCs. Suppression of Nrip1 by siNrip1 also decreased the expressions of mTOR, p-mTOR, p65, and p-p65 in ADMSCs. Reduced expressions of p65 and p-p65 were also confirmed in the skin of Nrip1 knockout mice. These findings suggest that NRIP1 plays an important role in delaying skin aging by reducing ADMSCs senescence and maintaining ADMSCs quiescence.

Inflammasome in ALS Skeletal Muscle: NLRP3 as a Potential Biomarker.

Since NLRP3 inflammasome plays a pivotal role in several neurodegenerative disorders, we hypothesized that levels of inflammasome components could help in diagnosis or prognosis of amyotrophic lateral sclerosis (ALS). Gene and protein expression was assayed by RT-PCR and Western blot. Spearman’s correlation coefficient was used to determine the linear correlation of transcriptional expression levels with longevity throughout disease progression in mice models. Kaplan-Meier analysis was performed to evaluate MCC950 effects (NLRP3 inhibitor) on lifespan of SOD1G93A mice. The results showed significant alterations in NLRP3 inflammasome gene and protein levels in the skeletal muscle of SOD1G93A mice. Spearman’s correlation coefficient revealed a positive association between Nlrp3 transcriptional levels in skeletal muscle and longevity of SOD1G93A mice (r = 0.506; p = 0.027). Accordingly, NLRP3 inactivation with MCC950 decreased the lifespan of mice. Furthermore, NLRP3 mRNA levels were significantly elevated in the blood of ALS patients compared to healthy controls (p = 0.03). In conclusion, NLRP3 could be involved in skeletal muscle pathogenesis of ALS, either through inflammasome or independently, and may play a dual role during disease progression. NLRP3 gene expression levels could be used as a biomarker to improve diagnosis and prognosis in skeletal muscle from animal models and also to support diagnosis in clinical practice with the blood of ALS patients.

Effect of Metformin on Cardiac Metabolism and Longevity in Aged Female Mice.

The antidiabetic drug metformin exerts pleiotropic effects on multiple organs, including the cardiovascular system. Evidence has shown that metformin improves healthspan and lifespan in male mice, yet its lifespan lengthening effect in females remains elusive. We herein demonstrated that metformin fails to extend the lifespan in female mice. Compared to 2-month-old young controls, 20-month-old female mice showed a spectrum of degenerative cardiac phenotypes alongside significant alterations in the extracellular matrix composition. Despite lowered reactive oxygen species production, long-term metformin treatment did not improve cardiac function in the aged female mice. In contrast, RNA sequencing analyses demonstrated that metformin treatment elevated the extracellular matrix-related gene while lowering oxidative phosphorylation-related gene expression in the heart. In addition, metformin treatment induced metabolic reprogramming that suppressed mitochondrial respiration but activated glycolysis (i.e., Warburg effect) in cultured primary cardiomyocytes and macrophages, thereby sustaining an inflammatory status and lowering ATP production. These findings suggest the unexpected detrimental effects of metformin on the regulation of cardiac homeostasis and longevity in female mice, reinforcing the significance of comprehensive testing prior to the translation of metformin-based novel therapies.

Extending lifespan by modulating the growth hormone/insulin-like growth factor-1 axis: coming of age.

Progress made in theyears of aging research have allowed the opportunity to explore potential interventions to slow aging and extend healthy lifespan. Studies performed in yeast, worms, flies and mice subjected to genetic and pharmacological interventions have given insight into the cellular and molecular mechanisms associated with longevity. Furthermore, it is now possible to effectively modulate pathways that slow aging at different stages of life (early life or at an adult age). Interestingly, interventions that extend longevity in adult micehave had sex-specific success, suggesting a potential link between particular pathways that modulate aging and sex. For example, reduction of the growth hormone (GH)/insulin-like growth factor-1 (IGF-1) axis at an adult age extends lifespan preferentially in females. Moreover, several postnataldietary interventions tested by the 'Intervention Testing Program (ITP)' from the National Institute of Aging (NIA) have shown that while pharmacological interventions like rapamycin affect the IGF-1/insulin pathway and preferentially extend lifespan in females; dietary compounds that target other cellular pathways are effective only in male mice-indicating mutually exclusive sex-specific pathways. Therefore, a combination of interventions that targetnon-overlapping aging-related pathways appears to be an effectiveapproach to further extend healthy lifespan in both sexes. Here, we review the germline and postnatal mouse lines that target the GH/IGF-1 axis as a mechanism to extend longevity as well as the dietary compounds that tested positive in the NIA program to increase lifespan. We believe that the interventions reviewed in this paper could constitute feasible combinations for an extended healthy lifespan in both male and female mice.

Lifelong restriction of dietary branched-chain amino acids has sex-specific benefits for frailty and lifespan in mice.

Protein restricted (PR) diets promote health and longevity in many species. While the precise components of a PR diet that mediate the beneficial effects to longevity have not been defined, we recently showed that many metabolic effects of PR can be attributed to reduced dietary levels of the branched-chain amino acids (BCAAs) leucine, isoleucine, and valine. Here, we demonstrate that restricting dietary BCAAs increases the survival of two different progeroid mouse models, delays frailty and promotes the metabolic health of wild-type C57BL/6J mice when started in midlife, and leads to a 30% increase in lifespan and a reduction in frailty in male, but not female, wild-type mice when fed lifelong. Our results demonstrate that restricting dietary BCAAs can increase healthspan and longevity in mice, and suggest that reducing dietary BCAAs may hold potential as a translatable intervention to promote healthy aging.

Longevity-Extending MetAP2 Inhibitors Induce Caloric Restriction Through P53-Dependent Induction of GDF-15

MetAP2 is a 67kDa protein which sits at the translation initiation complex and cleaves N-terminal methionine off of nascent peptides. Inhibitors of MetAP2 cause profound weight loss secondary to decreased food intake. These inhibitors also significantly extend longevity in mice in late-life intervention. However, the exact mechanism of action causing decreased food intake is not known. Here we investigated the molecular mechanism and target tissue of a MetAP2 inhibitor’s (Zgn1062) anorectic effects. First we identified the target tissue by testing targeted Zgn1062 delivery to specific brain regions. Delivery to the medio-basal hypothalamus did not have a significant effect but delivery to the lateral ventricle resulted in significantly decrease food intake and body weight after 2 and 14 hours. When we delivered a neuron-targeted AAV encoding MetAP2 shRNA we saw decreased efficacy of MetAP2 confirming the required for neuronal MetAP2 for anorectic effects. To determine the molecular mechanisms we performed RNAseq of wildtype and MetAP2 KO HT1080 cells across a timecourse of Zgn1062 treatment. The main pathway activated across timepoints in MetAP2-dependent manner was P53 signaling. A main P53 target that was upregulated was the known anorectic peptide GDF15. We confirmed GDF15 increases in vivo at both mRNA (liver and intestines) and protein level (serum) in response to Zgn1062. We also found that Zgn1062 treatment reduces senescent cell burden in visceral adipose tissue in vivo and reduces SASP gene expression in fat explants ex vivo. We hypothesize that Zgn1062’s potent P53 activation may play a role in clearance of senescent cells.

Topologically selective islet vulnerability and self-sustained downregulation of markers for \u03b2-cell maturity in streptozotocin-induced diabetes

Mouse models of Streptozotocin (STZ) induced diabetes represent the most widely used preclinical diabetes research systems. We applied state of the art optical imaging schemes, spanning from single islet resolution to the whole organ, providing a first longitudinal, 3D-spatial and quantitative account of β-cell mass (BCM) dynamics and islet longevity in STZ-treated mice. We demonstrate that STZ-induced β-cell destruction predominantly affects large islets in the pancreatic core. Further, we show that hyperglycemic STZ-treated mice still harbor a large pool of remaining β-cells but display pancreas-wide downregulation of glucose transporter type 2 (GLUT2). Islet gene expression studies confirmed this downregulation and revealed impaired β-cell maturity. Reversing hyperglycemia by islet transplantation partially restored the expression of markers for islet function, but not BCM. Jointly our results indicate that STZ-induced hyperglycemia results from β-cell dysfunction rather than β-cell ablation and that hyperglycemia in itself sustains a negative feedback loop restraining islet function recovery.

Dietary magnesium supplementation improves lifespan in a mouse model of progeria

Aging is associated with redox imbalance according to the redox theory of aging. Consistently, a mouse model of premature aging (LmnaG609G/+) showed an increased level of mitochondrial reactive oxygen species (ROS) and a reduced basal antioxidant capacity, including loss of the NADPH‐coupled glutathione redox system. LmnaG609G/+ mice also exhibited reduced mitochondrial ATP synthesis secondary to ROS‐induced mitochondrial dysfunction. Treatment of LmnaG609G/+ vascular smooth muscle cells with magnesium‐enriched medium improved the intracellular ATP level, enhanced the antioxidant capacity, and thereby reduced mitochondrial ROS production. Moreover, treatment of LmnaG609G/+ mice with dietary magnesium improved the proton pumps (complexes I, III, and IV), stimulated extramitochondrial NADH oxidation and enhanced the coupled mitochondrial membrane potential, and thereby increased H+‐coupled mitochondrial NADPH and ATP synthesis, which is necessary for cellular energy supply and survival. Consistently, magnesium treatment reduced calcification of vascular smooth muscle cells in vitro and in vivo, and improved the longevity of mice. This antioxidant property of magnesium may be beneficial in children with HGPS.

Resolving Peto's paradox: Modeling the potential effects of size-related metabolic changes, and of the evolution of immune policing and cancer suppression.

The intrinsic risk of cancer increases with body size and longevity; however, big long‐lived species do not exhibit this increase, a contradiction named Peto's paradox. Five hypotheses potentially resolving this paradox were modeled using the multistage model of carcinogenesis. The five hypotheses were based on (1) intrinsic changes in metabolic rate with body size; adaptive increase in immune policing of (2) cancer cells or (3) cells with driver mutations; or adaptive increase in cancer suppression via (4) decreased somatic mutation rate, or (5) increased genetic control. Parameter changes needed to stabilize cancer risk in three types of cancer were estimated for tissues scaled from mouse size and longevity to human and blue whale levels. The metabolic rate hypothesis alone was rejected due to a conflict between the required interspecific effect with the observed intraspecific effect of size on cancer risk, but some metabolic change was optionally incorporated in the other models. Necessary parameter changes in immune policing and somatic mutation rate far exceeded values observed; however, natural selection increasing the genetic suppression of cancer was generally consistent with data. Such adaptive increases in genetic control of cancers in large and/or long‐lived animals raise the possibility that nonmodel animals will reveal novel anticancer mechanisms.

Preclinical evaluation of multi stimuli responsive core-plasmonic nanoshell for photo-triggered tumor ablation: A disintegrable nanohybrid

Photothermal therapy (PTT) has gained tremendous interest as a mode of localized cancer therapy. Herein, we report a stable NIR absorbing gold modified dual responsive polymer core, chitosan-g-PNVCL (Au PNVCL NS). The in vitro studies reveal that Au PNVCL NS is a thermo-labile, non-immunogenic, biocompatible material possessing excellent photothermal transduction potential. The acute single-dose toxicity testing in mice reveals no abnormalities, indicating good tolerability of formulation. Au PNVCL NS shows excellent tumor ablation efficacy in vivo upon irradiation with 750 nm laser, thus improving the disease-free longevity of mice. Importantly, keeping our focus on the fate of this material, a significant surge of gold nanoparticles of size ~5 nm in the urine samples of treated mice assures rapid elimination of the material soon after PTT. These results, along with numerous gold nanoparticles deposition in the kidney of mice receiving PTT affirms the disintegrable nature of this nanoformulation while maintaining the desired PTT efficacy.