Oge Arum

Disruption of Growth Hormone Receptor Prevents Calorie Restriction from Improving Insulin Action and Longevity

Most mutations that delay aging and prolong lifespan in the mouse are related to somatotropic and/or insulin signaling. Calorie restriction (CR) is the only intervention that reliably increases mouse longevity. There is considerable phenotypic overlap between long-lived mutant mice and normal mice on chronic CR. Therefore, we investigated the interactive effects of CR and targeted disruption or knock out of the growth hormone receptor (GHRKO) in mice on longevity and the insulin signaling cascade. Every other day feeding corresponds to a mild (i.e. 15%) CR which increased median lifespan in normal mice but not in GHRKO mice corroborating our previous findings on the effects of moderate (30%) CR on the longevity of these animals. To determine why insulin sensitivity improves in normal but not GHRKO mice in response to 30% CR, we conducted insulin stimulation experiments after one year of CR. In normal mice, CR increased the insulin stimulated activation of the insulin signaling cascade (IR/IRS/PI3K/AKT) in liver and muscle. Livers of GHRKO mice responded to insulin by increased activation of the early steps of insulin signaling, which was dissipated by altered PI3K subunit abundance which putatively inhibited AKT activation. In the muscle of GHRKO mice, there was elevated downstream activation of the insulin signaling cascade (IRS/PI3K/AKT) in the absence of elevated IR activation. Further, we found a major reduction of inhibitory Ser phosphorylation of IRS-1 seen exclusively in GHRKO muscle which may underpin their elevated insulin sensitivity. Chronic CR failed to further modify the alterations in insulin signaling in GHRKO mice as compared to normal mice, likely explaining or contributing to the absence of CR effects on insulin sensitivity and longevity in these long-lived mice.

The growth hormone receptor gene‐disrupted mouse fails to respond to an intermittent fasting diet

The interaction of longevity‐conferring genes with longevity‐conferring diets is poorly understood. The growth hormone receptor gene‐disrupted (GHR‐KO) mouse is long lived; and this longevity is not responsive to 30% caloric restriction, in contrast to wild‐type animals from the same strain. To determine whether this may have been limited to a particular level of dietary restriction, we subjected GHR‐KO mice to a different dietary restriction regimen, an intermittent fasting diet. The intermittent fasting diet increased the survivorship and improved insulin sensitivity of normal males, but failed to affect either parameter in GHR‐KO mice. From the results of two paradigms of dietary restriction, we postulate that GHR‐KO mice would be resistant to any manner of dietary restriction; potentially due to their inability to further enhance insulin sensitivity. Insulin sensitivity may be a mechanism and/or a marker of the lifespan extending potential of an intervention.

Metabolic Alterations Due to Caloric Restriction and Every Other Day Feeding in Normal and Growth Hormone Receptor Knockout Mice

Mutations causing decreased somatotrophic signaling are known to increase insulin sensitivity and extend life span in mammals. Caloric restriction and every other day (EOD) dietary regimens are associated with similar improvements to insulin signaling and longevity in normal mice; however, these interventions fail to increase insulin sensitivity or life span in growth hormone receptor knockout (GHRKO) mice. To investigate the interactions of the GHRKO mutation with caloric restriction and EOD dietary interventions, we measured changes in the metabolic parameters oxygen consumption (VO2) and respiratory quotient produced by either long-term caloric restriction or EOD in male GHRKO and normal mice. GHRKO mice had increased VO2, which was unaltered by diet. In normal mice, EOD diet caused a significant reduction in VO2 compared with ad libitum (AL) mice during fed and fasted conditions. In normal mice, caloric restriction increased both the range of VO2 and the difference in minimum VO2 between fed and fasted states, whereas EOD diet caused a relatively static VO2 pattern under fed and fasted states. No diet significantly altered the range of VO2 of GHRKO mice under fed conditions. This provides further evidence that longevity-conferring diets cause major metabolic changes in normal mice, but not in GHRKO mice.

Specific suppression of insulin sensitivity in growth hormone receptor gene‐disrupted (GHR‐KO) mice attenuates phenotypic features of slow aging

In addition to their extended lifespans, slow‐aging growth hormone receptor/binding protein gene‐disrupted (knockout) (GHR‐KO) mice are hypoinsulinemic and highly sensitive to the action of insulin. It has been proposed that this insulin sensitivity is important for their longevity and increased healthspan. We tested whether this insulin sensitivity of the GHR‐KO mouse is necessary for its retarded aging by abrogating that sensitivity with a transgenic alteration that improves development and secretory function of pancreatic β‐cells by expressing Igf‐1 under the rat insulin promoter 1 (RIP::IGF‐1). The RIP::IGF‐1 transgene increased circulating insulin content in GHR‐KO mice, and thusly fully normalized their insulin sensitivity, without affecting the proliferation of any non‐β‐cell cell types. Multiple (nonsurvivorship) longevity‐associated physiological and endocrinological characteristics of these mice (namely beneficial blood glucose regulatory control, altered metabolism, and preservation of memory capabilities) were partially or completely normalized, thus supporting the causal role of insulin sensitivity for the decelerated senescence of GHR‐KO mice. We conclude that a delayed onset and/or decreased pace of aging can be hormonally regulated.

Prevention of Neuromusculoskeletal Frailty in Slow-Aging Ames Dwarf Mice: Longitudinal Investigation of Interaction of Longevity Genes and Caloric Restriction

Ames dwarf (Prop1 df/df) mice are remarkably long-lived and exhibit many characteristics of delayed aging and extended healthspan. Caloric restriction (CR) has similar effects on healthspan and lifespan, and causes an extension of longevity in Ames dwarf mice. Our study objective was to determine whether Ames dwarfism or CR influence neuromusculoskeletal function in middle-aged (82 ± 12 weeks old) or old (128 ± 14 w.o.) mice. At the examined ages, strength was improved by dwarfism, CR, and dwarfism plus CR in male mice; balance/ motor coordination was improved by CR in old animals and in middle-aged females; and agility/ motor coordination was improved by a combination of dwarfism and CR in both genders of middle-aged mice and in old females. Therefore, extension of longevity by congenital hypopituitarism is associated with improved maintenance of the examined measures of strength, agility, and motor coordination, key elements of frailty during human aging, into advanced age. This study serves as a particularly important example of knowledge related to addressing aging-associated diseases and disorders that results from studies in long-lived mammals.

The slow-aging growth hormone receptor/binding protein gene-disrupted (GHR-KO) mouse is protected from aging-resultant neuromusculoskeletal frailty

Neuromusculoskeletal (physical) frailty is an aging-attributable biomedical issue of extremely high import, from both public health and individual perspectives. Yet, it is rarely studied in nonhuman research subjects and very rarely studied in animals with extended longevity. In an effort to address this relatively neglected area, we have conducted a longitudinal investigation of the neuromusculoskeletal healthspan in mice with two senescence-slowing interventions: growth hormone (GH) resistance, produced by GH receptor “knockout” (GHR-KO), and caloric restriction (CR). We report marked improvements in the retention of strength, balance, and motor coordination by the longevity-conferring GHR/BP gene disruption, CR regimen, or a combination of the two. Specifically, GHR-KO mice exhibit superior grip strength, balance, and motor coordination at middle age, and CR mice display superior grip strength at middle age. The advantageous effects established by middle-age are more pronounced in old-age, and these robust alterations are, generally, not gender-specific. Thus, we show that genetic and/or dietary interventions that engender longevity are also beneficial for the retention of neuromusculoskeletal health and functionality. The translational knowledge to be gained from subsequent molecular or histological investigations of these models of preserved functionality and decelerated senescence is potentially relevant to the efforts to reduce the specter of fear, falls, fracture, and frailty in the elderly.

Preservation of blood glucose homeostasis in slow-senescing somatotrophism-deficient mice subjected to intermittent fasting begun at middle or old age

Poor blood glucose homeostatic regulation is common, consequential, and costly for older and elderly populations, resulting in pleiotrophically adverse clinical outcomes. Somatotrophic signaling deficiency and dietary restriction have each been shown to delay the rate of senescence, resulting in salubrious phenotypes such as increased survivorship. Using two growth hormone (GH) signaling-related, slow-aging mouse mutants we tested, via longitudinal analyses, whether genetic perturbations that increase survivorship also improve blood glucose homeostatic regulation in senescing mammals. Furthermore, we institute a dietary restriction paradigm that also decelerates aging, an intermittent fasting (IF) feeding schedule, as either a short-term or a sustained intervention beginning at either middle or old age, and assess its effects on blood glucose control. We find that either of the two genetic alterations in GH signaling ameliorates fasting hyperglycemia; additionally, both longevity-inducing somatotrophic mutations improve insulin sensitivity into old age. Strikingly, we observe major and broad improvements in blood glucose homeostatic control by IF: IF improves ad libitum-fed hyperglycemia, glucose tolerance, and insulin sensitivity, and reduces hepatic gluconeogenesis, in aging mutant and normal mice. These results on correction of aging-resultant blood glucose dysregulation have potentially important clinical and public health implications for our ever-graying global population, and are consistent with the Longevity Dividend concept.Poor blood glucose homeostatic regulation is common, consequential, and costly for older and elderly populations, resulting in pleiotrophically adverse clinical outcomes. Somatotrophic signaling deficiency and dietary restriction have each been shown to delay the rate of senescence, resulting in salubrious phenotypes such as increased survivorship. Using two growth hormone (GH) signaling-related, slow-aging mouse mutants we tested, via longitudinal analyses, whether genetic perturbations that increase survivorship also improve blood glucose homeostatic regulation in senescing mammals. Furthermore, we institute a dietary restriction paradigm that also decelerates aging, an intermittent fasting (IF) feeding schedule, as either a short-term or a sustained intervention beginning at either middle or old age, and assess its effects on blood glucose control. We find that either of the two genetic alterations in GH signaling ameliorates fasting hyperglycemia; additionally, both longevity-inducing somatotrophic mutations improve insulin sensitivity into old age. Strikingly, we observe major and broad improvements in blood glucose homeostatic control by IF: IF improves ad libitum-fed hyperglycemia, glucose tolerance, and insulin sensitivity, and reduces hepatic gluconeogenesis, in aging mutant and normal mice. These results on correction of aging-resultant blood glucose dysregulation have potentially important clinical and public health implications for our ever-graying global population, and are consistent with the Longevity Dividend concept.

The effect of calorie restriction on the presence of apoptotic ovarian cells in normal wild type mice and low-plasma-IGF-1 Laron dwarf mice

BackgroundIt is known that caloric restriction extends lifespan and can minimize age-related dysfunction of the reproductive system. We became interested in how caloric restriction influences apoptosis, which is a crucial process that maintains ovarian cell homeostasis.MethodsWe examined ovarian cells in: 2.5-year-old wild type mice on caloric restriction (CR) or fed ad libitum (AL) and Laron dwarf mice (GHR-KO) at the same ages on CR or fed AL. Apoptosis was assessed by histochemical analysis on paraffin sections of ovarian tissue.ResultsMorphological and histochemical analysis revealed that CR improved reproductive potential in 2.5-year-old WT littermates and GHR-KO female mice, as indicated by the increased number of ovarian follicles. The level of apoptosis in ovarian tissue was higher in WT mice on a CR diet compared with WT mice on the AL diet. In GHR-KO mice, the level of apoptosis in ovaries was similar for mice on CR and on AL diets and bigger than in WT mice on CR.ConclusionsMorphological and histochemical analysis revealed a younger biological age of the ovaries in 2-year-old WT littermates and GHR-KO female mice on CR compared with animals fed AL.

Resistance to the Beneficial Metabolic Effects and Hepatic Antioxidant Defense Actions of Fibroblast Growth Factor 21 Treatment in Growth Hormone-Overexpressing Transgenic Mice

Fibroblast growth factor 21 (FGF21) modulates a diverse range of biological functions, including glucose and lipid metabolism, adaptive starvation response, and energy homeostasis, but with limited mechanistic insight. FGF21 treatment has been shown to inhibit hepatic growth hormone (GH) intracellular signaling. To evaluate GH axis involvement in FGF21 actions, transgenic mice overexpressing bovine GH were used. Expectedly, in response to FGF21 treatment control littermates showed metabolic improvements whereas GH transgenic mice resisted most of the beneficial effects of FGF21, except an attenuation of the innate hyperinsulinemia. Since FGF21 is believed to exert its effects mostly at the transcriptional level, we analyzed and observed significant upregulation in expression of various genes involved in carbohydrate and lipid metabolism, energy homeostasis, and antioxidant defense in FGF21-treated controls, but not in GH transgenics. The resistance of GH transgenic mice to FGF21-induced changes underlines the necessity of normal GH signaling for the beneficial effects of FGF21.

Interaction of growth hormone receptor/binding protein gene disruption and caloric restriction for insulin sensitivity and attenuated aging

The correlation of physiological sensitivity to insulin ( vis-à-vis glycemic regulation) and longevity is extensively established, creating a justifiable gerontological interest on whether insulin sensitivity is causative, or even predictive, of some or all phenotypes of slowed senescence (including longevity). The growth hormone receptor/ binding protein gene-disrupted (GHR-KO) mouse is the most extensively investigated insulin-sensitive, attenuated aging model. It was reported that, in a manner divergent from similar mutants, GHR-KO mice fail to respond to caloric restriction (CR) by altering their insulin sensitivity. We hypothesized that maximized insulin responsiveness is what causes GHR-KO mice to exhibit a suppressed survivorship response to dietary (including caloric) restriction; and attempted to refute this hypothesis by assessing the effects of CR on GHR-KO mice for varied slow-aging-associated phenotypes. In contrast to previous reports, we found GHR-KO mice on CR to be less responsive than their ad libitum (A.L.) counterparts to the hypoglycemia-inducing effects of insulin. Further, CR had negligible effects on the metabolism or cognition of GHR-KO mice. Therefore, our data suggest that the effects of CR on the insulin sensitivity of GHR-KO mice do not concur with the effects of CR on the aging of GHR-KO mice.