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.     

Metabolic effects of intra-abdominal fat in GHRKO mice

Mice with targeted deletion of the growth hormone receptor (GHRKO mice) are growth hormone (GH) resistant, small, obese, hypoinsulinemic, highly insulin sensitive and remarkably long-lived. To elucidate the unexpected coexistence of adiposity with improved insulin sensitivity and extended longevity, we examined effects of surgical removal of visceral (epididymal and perinephric) fat on metabolic traits related to insulin signaling and longevity. Comparison of results obtained in GHRKO mice and in normal animals from the same strain revealed disparate effects of visceral fat removal (VFR) on insulin and glucose tolerance, adiponectin levels, accumulation of ectopic fat, phosphorylation of insulin signaling intermediates, body temperature, and respiratory quotient (RQ). Overall, VFR produced the expected improvements in insulin sensitivity and reduced body temperature and RQ in normal mice and had opposite effects in GHRKO mice. Some of the examined parameters were altered by VFR in opposite directions in GHRKO and normal mice, and others were affected in only one genotype or exhibited significant genotype × treatment interactions. Functional differences between visceral fat of GHRKO and normal mice were confirmed by measurements of adipokine secretion, lipolysis, and expression of genes related to fat metabolism. We conclude that in the absence of GH signaling, the secretory activity of visceral fat is profoundly altered and unexpectedly promotes enhanced insulin sensitivity. The apparent beneficial effects of visceral fat in GHRKO mice may also explain why reducing adiposity by calorie restriction fails to improve insulin signaling or further extend longevity in these animals.     

Neuronal control of lipid metabolism by STR‐2 G protein‐coupled receptor promotes longevity in Caenorhabditis elegans

The G protein‐coupled receptor (GPCR) encoding family of genes constitutes more than 6% of genes in Caenorhabditis elegans genome. GPCRs control behavior, innate immunity, chemotaxis, and food search behavior. Here, we show that C. elegans longevity is regulated by a chemosensory GPCR STR‐2, expressed in AWC and ASI amphid sensory neurons. STR‐2 function is required at temperatures of 20°C and higher on standard Escherichia coli OP50 diet. Under these conditions, this neuronal receptor also controls health span parameters and lipid droplet (LD) homeostasis in the intestine. We show that STR‐2 regulates expression of delta‐9 desaturases, fat‐5, fat‐6 and fat‐7, and of diacylglycerol acyltransferase dgat‐2. Rescue of the STR‐2 function in either AWC and ASI, or ASI sensory neurons alone, restores expression of fat‐5, dgat‐2 and restores LD stores and longevity. Rescue of stored fat levels of GPCR mutant animals to wild‐type levels, with low concentration of glucose, rescues its lifespan phenotype. In all, we show that neuronal STR‐2 GPCR facilitates control of neutral lipid levels and longevity in C. elegans.     

Lack of UCP1 stimulates fatty liver but mediates UCP1-independent action of beige fat to improve hyperlipidemia in Apoe knockout mice

Brown adipose tissue (BAT) plays a critical role in lipid metabolism and may protect from hyperlipidemia; however, its beneficial effect appears to depend on the ambient temperature of the environment. In this study, we investigated the effects of uncoupling protein 1 (UCP1) deficiency on lipid metabolism, including the pathophysiology of hyperlipidemia, in apolipoprotein E knockout (APOE-KO) mice at a normal (23 °C) and thermoneutral (30 °C) temperature. Unexpectedly, UCP1 deficiency caused improvements in hyperlipidemia, atherosclerosis, and glucose metabolism, regardless of an increase in hepatic lipid deposition, in Ucp1/Apoe double-knockout (DKO) mice fed a high-fat diet at 23 °C, with BAT hyperplasia and robust browning of inguinal white adipose tissue (IWAT) observed. Proteomics and gene expression analyses revealed significant increases in many proteins involved in energy metabolism and strong upregulation of brown/beige adipocyte-related genes and fatty acid metabolism-related genes in browned IWAT, suggesting an induction of beige fat formation and stimulation of lipid metabolism in DKO mice at 23 °C. Conversely, mRNA levels of fatty acid oxidation-related genes decreased in the liver of DKO mice. The favorable phenotypic changes were lost at 30 °C, with BAT whitening and disappearance of IWAT browning, while fatty liver further deteriorated in DKO mice compared with that in APOE-KO mice. Finally, longevity analysis revealed a significant lifespan extension of DKO mice compared with that of APOE-KO mice at 23 °C. Irrespective of the fundamental role of UCP1 thermogenesis, our results highlight the importance of beige fat for the improvement of hyperlipidemia and longevity under the atherogenic status at normal room temperature.     

Growth hormone receptor gene disruption in mature‐adult mice improves male insulin sensitivity and extends female lifespan

Studies in multiple species indicate that reducing growth hormone (GH) action enhances healthy lifespan. In fact, GH receptor knockout (GHRKO) mice hold the Methuselah prize for the world''s longest‐lived laboratory mouse. We previously demonstrated that GHR ablation starting at puberty (1.5 months), improved insulin sensitivity and female lifespan but results in markedly reduced body size. In this study, we investigated the effects of GHR disruption in mature‐adult mice at 6 months old (6mGHRKO). These mice exhibited GH resistance (reduced IGF‐1 and elevated GH serum levels), increased body adiposity, reduced lean mass, and minimal effects on body length. Importantly, 6mGHRKO males have enhanced insulin sensitivity and reduced neoplasms while females exhibited increased median and maximal lifespan. Furthermore, fasting glucose and oxidative damage was reduced in females compared to males irrespective of Ghr deletion. Overall, disrupted GH action in adult mice resulted in sexual dimorphic effects suggesting that GH reduction at older ages may have gerotherapeutic effects.     

Effects of pH,NH<Subscript>4</Subscript>-N concentration,temperature, and storage period on basidiospore germination in an ectomycorrhizal ammonia fungus <Emphasis Type="Italic">Hebeloma vinosophyllum</Emphasis>

Basidiospore germination in an ectomycorrhizal ammonia fungus Hebeloma vinosophyllum was stimulated by 10–500 mM NH₄Cl aqueous solution at pH 4.5–9.0, but not by pure water. The basidiospores germinated at 10°–35°C with an optimum at 25°–30°C. The highest germination percentage (83.0%) was observed in 100 mM NH₄Cl aqueous solution adjusted to pH 8.0 by KOH, when the basidiospores were incubated at a density of 10⁶ spores/ml at 30°C for 14 days. The percent germination value decreased with the increased duration of storage under both dry and wet conditions. Humidity and temperature affected the longevity of H. vinosophyllum basidiospores. The basidiospores maintained their germination ability longer under a dry condition than under a wet condition. The greatest longevity was accomplished by storage at 15°C under a dry condition.     

Longevity and resistance to cold stress in cold‐stress selected lines and their controls in Drosophila melanogaster

Thermal environments can influence many fitness‐related traits including life span. Here, we assess whether longevity in Drosophila melanogaster can experimentally evolve as a correlated response to cold‐stress selection, and whether genotype‐by‐temperature and sex‐by‐temperature interactions are significant components of variation in life span. Three replicated S lines were cold‐stress selected and compared with their respective unselected controls (Clines) in the 16^th generation of thermal selection. Cold‐stress resistance exhibited a substantial direct response to selection, and also showed a significant interaction between sex and type of line. Mean longevity exhibited a significant interaction between adult test temperature (14 and 25 °C) and line (with suggestive evidence for increased longevity of S lines when tested at 14 °C), but there was no evidence for increased longevity in S lines at normal temperatures (i.e. 25 °C). Another temperature‐dependent effect was sex‐specific, with males being the longer lived sex at 25 °C but the less long‐lived sex at 14 °C. Additionally, we tested in an exploratory way the relationship between longevity and cold‐stress resistance by also measuring resistance to a prefreezing temperature before and after one generation of longevity selection at 14 °C (selection intensity, i = 1.47 for S lines, and 1.42 for C lines). In this longevity selection, we found that cold‐stress resistance increased by about 6% in S lines and 18% in C lines. However, taken together, the results indicate no simple relationship between longevity and cold‐stress resistance, with genotype‐by‐sex interactions in both traits. Temperature dependent interaction in longevity is apparent between S and C lines, and sex‐specific variation in mean longevity also depends on temperature.     

The double face of cold in cancer

In a recent paper published in Nature, multiple evidence is provided that cold exposure causes tumor growth restriction in mice, by activating brown adipose tissue metabolism and by subsequent cancer cells’ glucose starvation. The paper shows a tumor growth inhibition by 80% for multiple cancer types in mice exposed to 4 °C in comparison with mice exposed to 30 °C. These results are very promising since cost effective protocols could be designed for future clinical trials, for several cancer forms. In this commentary, an extensive analysis is performed on the potential of these results. Some previous published studies are discussed as well, showing differences in tumor growth for mice housed in different external temperatures.     

Estimating reaction norms for predictive population parameters,age specific mortality,and mean longevity in temperature‐dependent cohorts of Culex quinquefasciatus Say (Diptera: Culicidae)

Culex quinquefasciatus plays a major role in the transmission of important parasites and viruses throughout the world. Because temperature is an important limiting factor on growth and longevity of all mosquito species, estimating the reaction norms provides very important basic information for understanding both plasticity and individual variations of the population. In the present study, Cx. quinquefasciatus were maintained at five different constant temperatures (15°, 20°, 23°, 27°, and 30°C) for two subsequent generations. Reproductive population parameters in blood‐fed mated females and longevities of virgin and blood‐fed mated adults reared at different temperatures were compared for the two generations. Longevity increased as temperature decreased within a range of 15° to 30°C for the unmated adults, and 15° to 27°C for the mated and blood‐fed adults. Generation times were as long as 124.07 and 106.76 days for two subsequent generations reared at 15°C, and the highest intrinsic rate of increase (r_m) values were estimated at 0.22 and 0.18, respectively, from the cohorts reared at 27°C. For survival rates, reproductive rates (R₀), and r_m values, 30°C was found to be a critical temperature for this species. These cohorts produced the smallest amount of eggs (R₀= 5.06), r_m values decreasing across generations (from 0.11 to 0.06), and the survival rates from egg to adult were found to be insufficient (16.1 and 10.8%). Additionally, the rate of exponential increase with age and age specific mortalities (b) were calculated for the virgin cohorts. Age specific mortality rates increased as temperature decreased. The increase in mortality rates started to accelerate at 27°C and was more pronounced at 30°C, for both females and males. We estimated the coefficients of variation for the b values in which females have smaller coefficients than those of the males at all temperatures.     

Bcl‐xL overexpression does not enhance specific erythropoietin productivity of recombinant CHO cells grown at 33°C and 37°C

Overexpression of bcl‐xL in recombinant Chinese hamster ovary (rCHO) cells has been known to suppress apoptotic cell death and thereby extend culture longevity during batch culture. However, its effect on specific productivity (q) of rCHO cells is controversial. This study attempts to investigate the effect of bcl‐xL overexpression on q of rCHO cells producing erythropoietin (EPO). To regulate the bcl‐xL expression level, the Tet‐off system was introduced in rCHO cells producing EPO (EPO‐off‐bcl‐xL). The bcl‐xL expression level was tightly controlled by doxycycline concentration. To evaluate the effect of bcl‐xL overexpression on specific EPO productivity (q_EPO) at different levels, EPO‐off‐bcl‐xL cells were cultivated at the two different culture temperatures, 33°C and 37°C. The q_EPO at 33°C and 37°C in the presence of 100 ng/mL doxycycline (without bcl‐xL overexpression) were 4.89 ± 0.21 and 3.18 ± 0.06 μg/10⁶cells/day, respectively. In the absence of doxycycline, bcl‐xL overexpression did not affect q_EPO significantly, regardless of the culture temperature, though it extended the culture longevity. Taken together, bcl‐xL overexpression showed no significant effect on the q_EPO of rCHO cells grown at 33°C and 37°C. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009     

Catalytic De/Hydrogenation in Mg by Co‐Doped Ni and VOx on Active Carbon: Extremely Fast Kinetics at Low Temperatures and High Hydrogen Capacity

A multi‐component catalyst Ni‐VO_x/AC (VO_x is comprised of V₂O₅ and VO₂, x = 2.18) was synthesized by a wet impregnation method. The synthesized Ni‐VO_x/AC shows a superior catalytic effect on de/hydrogenation of Mg. The MgH₂+Ni‐VO_x/AC composites can absorb 6.2 wt.‐% hydrogen within only 1 min at 150 °C under a hydrogen pressure of 2 MPa and desorb 6.5 wt.‐% hydrogen within 10 min at 300 °C under an initial hydrogen pressure of 1 KPa, which overcomes a critical barrier for practical use of Mg as a hydrogen storage material. A significant decrease of activation energy (E_a) indicates that Ni‐VO_x/AC catalyst is highly efficient for Mg de/hydrogenation, which may be ascribed to the synergistic effect of bimetals (metal oxides) and nanocarbon.     

Reproduction and population parameters of the Nearctic predator Geocoris punctipes at constant and varying temperature regimes

The heteropteran predator Geocoris punctipes (Say) has been used in augmentative biological control since 2000 to control Lepidoptera. However, surprisingly, few data are available about the influence of temperature on its population development, which is of key importance to plan the number and moment of releases to obtain sufficient pest reduction. The objective of this study was to evaluate daily and total fecundity, longevity and life table parameters (m_x, l_x, r_m, R, λ, T and TD) of G. punctipes at constant (16.8°C, 21.5°C, 24.5°C and 28.3°C) and corresponding varying (day/night) (21/11°C, 24/18°C, 27/21°C and 30/26°C) temperatures. Pairs of adult predators aged 24 h and originating from nymphs exposed to the same temperature regimes were kept at the above‐mentioned temperature regimes in Petri dishes containing Anagasta kuehniella (Zeller) eggs and an oviposition substrate. Tests were conducted in climatic chambers at the different temperature regimes and a RH 70 ± 10% and a 14L: 10D photoperiod. Reproduction, longevity and life table parameters were significantly affected by temperature, with clear differences between treatments at low (16.8°C, 21/11°C, 21.5°C, 24/18°C) or a high (24.5°C, 27/21°C, 28.3°C, 30/26°C) temperature regimes. Highest reproduction and fastest population growth of G. punctipes took place at average temperatures ranging from 24.5°C to 30°C, and neither reproduction nor population growth was negatively influenced by varying temperatures at any of the temperature regimes.     

Anti‐aging drugs reduce hypothalamic inflammation in a sex‐specific manner

Aging leads to hypothalamic inflammation, but does so more slowly in mice whose lifespan has been extended by mutations that affect GH/IGF‐1 signals. Early‐life exposure to GH by injection, or to nutrient restriction in the first 3 weeks of life, also modulate both lifespan and the pace of hypothalamic inflammation. Three drugs extend lifespan of UM‐HET3 mice in a sex‐specific way: acarbose (ACA), 17‐α‐estradiol (17αE2), and nordihydroguaiaretic acid (NDGA), with more dramatic longevity increases in males in each case. In this study, we examined the effect of these anti‐aging drugs on neuro‐inflammation in hypothalamus and hippocampus. We found that age‐associated hypothalamic inflammation is reduced in males but not in females at 12 months of age by ACA and 17αE2 and at 22 months of age in NDGA‐treated mice. The three drugs blocked indices of hypothalamic reactive gliosis associated with aging, such as Iba‐1‐positive microglia and GFAP‐positive astrocytes, as well as age‐associated overproduction of TNF‐α. This effect was not observed in drug‐treated female mice or in the hippocampus of the drug‐treated animals. On the other hand, caloric restriction (CR; an intervention that extends the lifespan in both sexes) significantly reduced hypothalamic microglia and TNF‐α in both sexes at 12 months of age. Together, these results suggest that the extent of drug‐induced changes in hypothalamic inflammatory processes is sexually dimorphic in a pattern that parallels the effects of these agents on mouse longevity and that mimics the changes seen, in both sexes, of long‐lived nutrient restricted or mutant mice.     

Temperature dependence of O2 consumption; opposite effects of leptin and etomoxir on respiratory quotient in mice

Objectives : The aims were to compare the temperature dependence of the metabolic rate in young ob/ob mice with that in mature ob/ob and db/db mice and to examine the effect on the metabolic substrate preference of leptin and etomoxir in ob/ob, C57BL/6J (wild‐type), and db/db mice. Research Methods and Procedures : In vivo oxygen consumption and carbon dioxide production were continuously measured by indirect calorimetry, and body temperature and total locomotor activity were measured by an implanted transponder. Leptin, etomoxir, or vehicle was administered intraperitoneally. Results : The temperature dependence of the metabolic rate of mature ob/ob and db/db mice were similar to that in wild‐type mice. In young 6‐week‐old ob/ob mice, the metabolic rate was almost doubled at 15 °C. Leptin (2 × 3 mg/kg) decreased the respiratory quotient (RQ) and carbon dioxide production but did not alter oxygen consumption, body temperature, or locomotor activity in ob/ob and C57BL/6J mice and had no effect in the db/db mice. Etomoxir (2 × 30 mg/kg) enhanced RQ and decreased oxygen consumption, carbon dioxide production, and body temperature in ob/ob, C57BL/6J, and db/db mice. Total locomotor activity was reduced in ob/ob and C57BL/6J mice. Discussion : In young ob/ob mice, the temperature sensitivity was enhanced compared with mature mice. Leptin and etomoxir had opposite effects on metabolic substrate preference. Leptin and lowered environmental temperature increased the relative fat oxidation as indicated by decreased RQ, possibly through activation of the sympathetic nervous system.     

Influences of temperature and humidity on the life history parameters and prey consumption of <Emphasis Type="Italic">Anthocoris minki</Emphasis> Dohrn (Heteroptera: Anthocoridae)

Anthocoris minki Dohrn is a promising indigenous Anthocoris species for the biological control of Agonoscena pistaciae Burck. and Laut. (Homoptera: Psyllidae) in pistachio orchards in Turkey. The adult longevity, fecundity, life table parameters and prey consumption of A. minki fed on Ephestia kuehniella Zeller (Lepidoptera: Pyralidae) eggs were studied at combinations of three constant temperatures (20, 25 and 30 ± 1°C) with two relative humidity (RH) levels (40 and 65 ± 5%). Studies indicated that temperature and RH significantly affected adult longevity, fecundity and prey consumption of A. minki. The greatest adult female longevity was 116.0 days at 20°C and 65% RH; the shortest adult female longevity was 27.5 days at 30°C and 40% RH. At all tested temperatures, the oviposition period and prey consumption of both females and males significantly decreased at low RH compared to high RH. The highest and lowest total fecundities were 276.0 eggs (at 20°C and 65% RH) and 42.4 eggs (at 25°C and 40% RH), respectively. The intrinsic rates of natural increase (r _m) at 40 and 65% RH were 0.049 and 0.076 at 20°C, 0.072 and 0.096 at 25°C and 0.076 and 0.112 at 30°C, respectively. The highest mean numbers of E. kuehniella eggs consumed by females and males were 859.6 (at 20°C) and 515.3 (at 25°C) at 65% RH, respectively; the lowest were 183.3 (at 20°C) and 95.5 (at 25°C) at 40% RH, respectively.     

What does not kill it does not always make it stronger: High temperatures in pyriproxyfen treatments produce Aedes aegypti adults with reduced longevity and smaller females

Aedes aegypti control in Brazil comprises integrated actions, in which larvicide application is a supplementary measure. Despite the importance of analyzing the effects of temperature on the efficiency of larvicides to control mosquito populations, there is still a lack of information regarding the sublethal effect of larvicides. We hypothesized that mosquitoes which survived pyriproxyfen exposure, during their immature development, have small body sizes and live less than mosquitoes that were not exposed to this larvicide. We investigated the sublethal effects of five different pyriproxyfen concentrations (0.0001; 0.001; 0.01; 0.1 and 1 mg.L⁻¹), under three different temperatures (20, 25 and 30 °C). As we increased the larvicide concentration, less larvae survived and developed into adults. However, at 30 °C the survival was higher than at 25 °C and 20 °C comparing the concentrations of 0.001 mg.L⁻¹ and 0.01 mg.L⁻¹ (10% and 16% high, respectively). Mosquito survivors to pyriproxyfen exposure, in all thermal conditions, had shorter adult life spans than mosquitoes not exposed to pyriproxyfen during the larval stage. The females exposed at 30 °C showed smaller wings than females from experiments at 20 °C. These findings provide evidence that the biological parameters of the adult lifespan and wing centroid size are impaired due to larval exposure to pyriproxyfen, even at high temperatures. Reduced longevity and small wing size are fitness costs to survive this larvicide exposure. These findings provide support to assess resistance development to larvicides through future generations and contribute to the discussion on improving the rational application of larvicides.     

Effects of temperature on biology and life table parameters of the Asian citrus psyllid, Diaphorina citri Kuwayama (Homoptera: Psyllidae)

The development, survivorship, longevity, reproduction, and life table parameters of the Asian citrus psyllid, Diaphorina citri Kuwayama were evaluated at 10°C, 15°C, 20°C, 25°C, 28°C, 30°C and 33°C. The populations reared at 10°C and 33°C failed to develop. Between 15°C and 30°C, mean developmental period from egg to adult varied from 49.3 days at 15°C to 14.1 days at 28°C. The low‐temperature developmental thresholds for 1st through 5th instars were estimated at 11.7°C, 10.7°C, 10.1°C, 10.5°C and 10.9°C, respectively. A modified Logan model was used to describe the relationship between developmental rate and temperature. The survival of the 3rd through 5th nymphal instars at 15–28°C was essentially the same. The mean longevity of females increased with decreasing temperature within 15–30°C. The maximal longevity of individual females was recorded 117, 60, 56, 52 and 51 days at 15°C, 20°C, 25°C, 28°C and 30°C, respectively. The average number of eggs produced per female significantly increased with increasing temperature and reached a maximum of 748.3 eggs at 28°C (P<0.001). The population reared at 28°C had the highest intrinsic rate of increased (0.199) and net reproductive rate (292.2); and the shortest population doubling time (3.5 days) and mean generation time (28.6 days) compared with populations reared at 15–25°C. The optimum range of temperatures for D. citri population growth was 25–28°C.