Reduction of insulin/insulin-like growth factor 1 (IGF1) signaling (IIS) extends the lifespan of various species. So far, several longevity mouse models have been developed containing mutations related to growth signaling deficiency by targeting growth hormone (GH), IGF1, IGF1 receptor, insulin receptor, and insulin receptor substrate. In addition, p70 ribosomal protein S6 kinase 1 (S6K1) knockout leads to lifespan extension. S6K1 encodes an important kinase in the regulation of cell growth. S6K1 is regulated by mechanistic target of rapamycin (mTOR) complex 1. The v-myc myelocytomatosis viral oncogene homolog (MYC)-deficient mice also exhibits a longevity phenotype. The gene expression profiles of these mice models have been measured to identify their longevity mechanisms. Here, we summarize our knowledge of long-lived mouse models related to growth and discuss phenotypic characteristics, including organ-specific gene expression patterns.
Insulin/insulin-like growth factor 1 (IGF1) signaling (IIS) is evolutionarily conserved from worms to human (1). Decreased IIS has been related to longevity in an evolutionarily conserved manner (2), showing consistent results in major model organisms, including worms (1), flies (3), and mice (4). Human centenarian studies have revealed that variations are positioned at proteins involved in the IIS system (5). In rodents, IGF1 is mainly produced from the liver by stimulation of growth hormone (GH, also known as somatotropin) secreted from the somatotrophs in the anterior pituitary (6). Circulating IGF1 binds to insulin/IGF1 receptors on the cell membrane of organs and primarily promotes tissue growth at an early developmental stage. Insulin, the other binding substrate of the insulin/IGF1 receptors, is produced by pancreatic β cells in response to postprandial nutrient influx (7). Insulin reduces high blood glucose levels after meals to maintain glucose homeostasis by suppressing glucose production in the liver, and stimulating glucose uptake in muscle and in fat by activation of their carbohydrate metabolism (8). This GH-IIS endocrine cascade axis is an excellent model for aging research to interpret how growth relates to metabolism. Insulin and IGF1 activate the IIS pathway through the insulin receptor (IR), IGF1 receptor (IGF1R), insulin receptor substrate 1 (IRS1), IRS2, phosphoinositide 3-kinase (PI3K), and protein kinase B (AKT) (7). Consequently, the IIS pathway changes gene expression through transcription factors, such as forkhead box O (FOXOs) and the v-myc myelocytomatosis viral oncogene homolog (MYC) (Fig. 1). In this review, we discuss physiological (Table 1) and global gene expression (Table 2) analyses in mice genetically modified to reduce GH-IIS axis in an effort to elucidate the longevity mechanism.
Several dwarf mouse models have been introduced in the aging research field, including Snell (disruption of the POU-domain transcription factor [Pit−/−]), Ames (defect of the homeobox protein prophet of PIT-1 [Prop 1−/−]), and Little (a missense mutation at growth hormone-releasing hormone receptor [Ghrhr−/−]). The Snell and Ames mice are hypopituitary dwarf deficient in GH, prolactin (PRL), and thyroid-stimulating hormone (TSH). These dwarf mice have been shown to live 37–68% longer than their wild-type (WT) littermates (9–11). The Little mice are another GH-deficient model. It was generated by deletion of the Ghrhr gene and the mice have been shown to live 23–25% longer than their WT littermates (9). The Snell, Ames, and Little GH deficient mice commonly have small body sizes and high adiposity, with low IGF1 levels.
The mouse GH receptor (Ghr) gene was disrupted (GHR−/−) to create a mouse model for human Laron syndrome (12). The GHR−/− mouse has been shown to have very low (less than 10% of normal) plasma IGF1 level and severely reduced IGF binding protein 3 (IGFBP3) level (13). A GH receptor antagonist (GHA) transgenic (Tg) mouse was generated by a single amino acid substitution in GH (14). A 50% reduction in plasma IGF1 has been reported in GHA tg mice, compared to their WT littermates. Both GHR−/− and GHA Tg mice have been shown to express obese and slow growth phenotypes (13, 15, 16), as well as small organs, including liver and kidney (17). GHR−/− mice have been reported to live longer (30–55%) than their WT littermates (13), but GHA Tg mice lived as long as their WT littermates (17).
Fibroblast growth factor-21 (FGF21) is an endocrine hormone related to energy metabolism and stress response (18). FGF21 binds to the transmembrane protein βKlotho, which has a protein sequence similar to Klotho (Kl). While studies have shown that Kl Tg mice overexpressing Kl lived longer compared to WT mice, βKlotho−/− mice lived as long as WT mice (19). Fasting has been shown to increase FGF21 levels, causing GH resistance by inactivation of downstream signal transducer and activator of transcription 5 (STAT5), and down-regulation of Igf1 gene (20). Indeed, FGF21-overexpressing Tg mice were shown to have small body sizes (60% of WT), decreased serum insulin and IGF1 levels, and a 36% extended lifespan (21). These results suggest that FGF21 is an upstream regulator of the IIS pathway.
The effect of overexpression of GH in Tg mice has been studied by insertion of the GH gene from other species, including human, rat, bovine, and ovine. Most GH Tg mice showed a high plasma GH level and a giant body size (6, 22). Interestingly, bovine GH (bGH) Tg mice exhibited lean bodies (16) and a 45% shorter lifespan than WT mice (22). In addition, bGH Tg mice had kidney damage, namely nephropathy with glomerular enlargement (23, 24). Human patients having an excess of GH show large hands, feet, face and internal organs, in a condition called acromegaly (25). Acromegaly patients also have lean bodies, heart problems, a high risk of thyroid cancer and colorectal cancer, and often have insulin-resistance or some diabetes (25, 26).
GH stimulates its primary target, the liver, causing it to produce IGF1. A number of microarray analyses have identified genes for longevity mechanism using liver tissues taken from Ames and Snell dwarf mice (27–34). Although early studies identified only a few genes (27, 28), later studies using high-density oligonucleotide microarrays that can detect > 12,000 RNA transcripts identified hundreds of differentially expressed genes (DEGs) as candidates in the aging mechanism. Papaconstantinou
Using Ames and Little male mice, 547 common DEGs (P < 0.001) were identified in liver during the aging process in 3, 6, 12, and 24 month old mice (29). Gene ontology (GO) analysis showed that the up-regulated DEGs are involved in the oxidative stress response, xenobiotic metabolism, mitochondrial respiration and β-oxidation, and the down-regulated DEGs are involved in humoral defense, including the complement system regulatory genes.
Using GHR−/− male mice (1.5 months old), 330 DEGs were identified in the liver. Up-regulated DEGs were those for β-oxidation, oxidative phosphorylation (OXPHOS), tricarboxylic acid (TCA) cycle, and factors for transcription and translation. Serine protease inhibitors were down-regulated DEGs (38). Using two GHR site-specific truncated mutants, GHR-569 and GHR-391 mouse strains, phosphorylation status of STAT5 was either decreased or removed in the liver by bGH treatment, respectively (38). GHR-STAT5 signaling was severely weakened in these two mutant mice. Using these two mutant mice, 20 common DEGs (P < 0.0005 and FC > 1.5) were identified in the liver (38). Down-regulated DEGs included Igf1, Igfals, Epidermal growth factor receptor (Egfr), and Hsd3b5. Up-regulated DEGs included Sulfotransferase family 2A, dehydroepiandrosterone (DHEA)-preferring, member 2 (Sult2a2), Angiogenin, ribonuclease, RNase A family, 5 (Ang), and Hao2. Sult2a2 and Ang play roles in the sulfation of hydroxysteroid and host defense, respectively. Up-regulated DEGs also included several genes encoding CYPs and glutathione S-transferases (GSTs) (38). Additional studies of GHR-391 and GHR−/− mice livers showed that the loss of GHR-STAT5 signaling caused intracellular lipid accumulation and steatosis by increasing phosphorylation of STAT1 and STAT3 proteins (39). A recent study using young GHR−/− male and female mice (16–18 weeks old) showed a transcript regulatory network for steroid hormone biosynthesis in the liver, composed of several long-noncoding RNA and microRNAs (40).
Using 3 month old young Fgf21 Tg male mice, transcriptome analysis identified 33, 22, and 8 DEGs (FDR < 0.1 and FC > 2) from the liver, epididymal white adipose tissue (WAT), and gastrocnemius, respectively (21). The hepatic DEGs included up-regulation of Fgf21, Fmo3, Cytochrome P450, family 2, subfamily b, polypeptide 9 (Cyp2b9), and Metallothionein 1 (Mt1) genes; and down-regulation of Igfals, Hsd3b5, Major urinary protein 4 (Mup4), Glutathione S-transferase, pi 1 (Gstp1), and Sterol carrier protein 2 (Scp2) genes (21). The authors compared the hepatic DEGs they found to 43 common hepatic DEGs from the four long-lived dwarf strains, including Snell, Ames, Little, and GHR−/− mice (41). They found eight overlapped genes, including down-regulated Igfals, Hairy and enhancer of split 6 (Hes6), Aminolevulinic acid synthase 2, erythroid (Alas2), Cyp4a12b, Mup4, Serine (or cysteine) peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 12 (Serpina12), and Hsd3b5; as well as up-regulated Fmo3 (21). Although only a small number of DEGs were found in the liver, white fat and skeletal muscle, the young long-lived Fgf21 Tg male mice showed dramatic phenotypical and physiological changes, including low body weight and low levels of plasma IGF-1, insulin, and glucose, as well as high levels of plasma ketone bodies (21).
Swindell performed a meta-analysis using microarray data obtained from livers of Snell, Ames, Little, and GHR−/− mice. DEG selection (mean difference significance test P < 0.05) revealed different DEG numbers among these mice models. The GHR−/− mice showed the smallest number of DEGs (46 DEGs), and other mice strains showed hundreds of DEGs: 151–316 DEGs from Snell, 400–987 DEGs from Ames, and 475–650 DEGs from Little (41). The range in number of DEGs was due to different ages of the mice. The results revealed that these mice strains shared strong down-regulation of Igf1 (41). Comparison of gene expression across these four dwarf mice identified 13 common DEGs, including 3 up-regulated genes: Hao3, Sult2a2, and Serine peptidase inhibitor, Kazal type 3 (Spink3). It also identified 10 down-regulated genes: Mup3, Mup4, Igf1, Igfals, Egfr, Socs2, Leukemia inhibitory factor receptor alpha (Lifr), Carboxylesterase 3A (Ces3a: also known as Es31), Hsd3b5, and Kidney expressed gene 1 (Keg1) (41). One down-regulated common DEG, Lifr, has not yet been studied for its role in aging and longevity. The Lifr gene encodes a receptor for the leukemia inhibitory factor (LIF) cytokine that affects various biological functions, including stem cell self-renewal, the reproductive process, and bone modeling (42). Interestingly, LIF can affect the anterior pituitary gland to decrease somatotroph, lactotroph and gonadotroph cells (43).
Malfunction of genes for DNA repair, such as Excision repair cross-complementing rodent repair deficiency, complementation group 1 (Ercc1), Ercc6 (also known as CSB), and Xeroderma pigmentosum, complementation group A (Xpa), caused progeroid syndromes in DNA repair-deficient mouse models. Csbm/m::Xpa−/− and Ercc1−/− mice showed a severe phenotype, Ercc1−/Δ−7 showed intermediate, Csbm/m showed mild, and Xpa−/− showed no phenotype in mouse models (44, 45). Interestingly, Csbm/m::Xpa−/− progeria mice showed reduction of serum IGF1 and glucose levels, and suppression of the GH-IIS axis (45). Schumacher
At the GH signal-deficient models including Snell, Ames, GHR−/−, and Fgf21 Tg mice, female models showed a greater difference of lifespan than male models (Table 1). GH secretion from the pituitary gland is stimulated by sex-steroids (47), and there is a different pattern of secretion between males and females (48). This indicates sexual dimorphism, including global gene expression in the liver (49–51).
JAK2 phosphorylates GHR serving as a docking site for STAT transcription factors in GH signaling in the liver. There are seven mouse STAT proteins, and STAT5a and STAT5b proteins play key roles in sex-specific hepatic gene expression (52, 53). In young (1.4–1.8 month old) mice, STAT5a regulated 23% (89 out of 393 genes) of the sex-specific gene expression, including down-regulation of Y-linked Ddx3y, Eif2s3y, and Jarid1d genes, up-regulation of Sult (sulfotransferase) genes, and both up- and down-regulation of CYP genes in female livers (53). STAT5b regulated 90% (767 out of 950 genes) of down-regulated genes and 61% (461 out of 753) of up-regulated genes in females (52). STAT5b-dependent sex-specific genes were primarily involved in oxidative metabolism, tryptophan and fatty acid metabolism, steroid metabolism, signaling, xenobiotic metabolism, and serine protease inhibition (52).
The major proportion of circulating IGF1 is produced by the liver (54). In the liver, by the binding of GH, GHR signaling transmits through JAK-STAT, and consequently activates IGF1 transcription, production, and secretion into the blood stream (6). Circulating IGF1 bioavailability is mediated by IGF binding proteins (IGFBPs). Deletion of the Pappa gene encoding a metalloproteinase, Pappalysin-1 that cleaves IGF1 bound to IGFBP4 decreased bioavailability of IGF1 (55). Indeed, Pappa−/− mice showed 40% smaller body mass, lower cancer incidence, and a 24–38% longer lifespan than WT controls (56–58).
Igf1-deficient (Igf1−/−) mice showed variable neonatal lethality in several different genetic backgrounds such as 129/Sv mice (10% survival), C57BL/6J x 129/Sv F1 hybrids (16% survival), and MF1 x 129/Sv F1 hybrids (68% survival) (59). The surviving Igf1−/− mice showed 24–40% lower body weights and increased maximum lifespan, but no significant change in mean lifespan (60). Although their body mass was lower than WT (WT 39 g vs Igf1−/− 30 g for male; WT 28 g vs Igf1−/− 17 g for female), there was almost no impact on the size of major organs, including liver (Igf1−/− 1.37 g vs WT 1.39 g), kidney (Igf1−/− 0.21 g vs WT 0.21 g), brain (Igf1−/− 0.44 g vs WT 0.47 g), and heart (Igf1−/− 0.16 g vs WT 0.22 g) (60). The IGF1 receptor (IGF1R) deficient (Igf1r−/−) mice show respiratory failure at birth that is 100% lethal (59). Similar to Igf1r−/− mice, homozygous insulin receptor (InsR)-deficient mice suffered from hyperglycemia and died soon after birth (61). Heterozygous InsR knockout (IR+/−) male mice showed heteroinsufficiency, including insulin insensitivity and a 20% extended maximum lifespan, but no significant change in average lifespan (62).
Both inducible liver-specific Igf1-disruption mice (LI-Igf1−/−) (54), and liver-specific Igf1-deficient mice (LID mice) (63) showed a 75% decrease IGF1 levels and high GH levels in serum. Igf1 expression was shut off after ~28 days for LI-Igf1−/− mice (54) and after ~10 days in LID mice (63). LI-Igf1−/− female mice, but not male mice, lived 16% longer than WT mice (64), whereas male LID mice exhibited a shortened lifespan (65).
Heterozygous Igf1r+/− mice exhibited 8% smaller body sizes and ~40% higher serum IGF1 level than those of WT mice (4). Increased serum IGF1 level is a compensatory response to reduced IGF1 receptor levels. Igf1r+/− mice also showed a low cellular IGF1 signaling activity because of glucose tolerance in the liver (4). Igf1r+/− mice lived 26% longer than WT controls, 33% longer for female mice and 16% longer for male mice, though the male difference was not statistically significant (4). Another independent study using Igf1r+/− mice showed negligible changes in lifespan (66). Ladiges
Klotho (Kl) encodes a circulating hormone and functions as an antagonist against IR and IGF1R, increasing insulin resistance (68). Kl−/− mice die early at < 100 days of age and show systemic aging phenotypes: growth retardation (small body size, and atrophy of genital organs and thymus), hypokinesis, arteriosclerosis, ectopic calcification in various organs and arterial walls, osteoporosis, and skin atrophy (69). In contrast, Kl overexpression significantly extended lifespan (19–31%) of Kl Tg mice through GH-independent suppression of the IIS pathway (68). Inhibition of IIS by additional genetic interventions, like Kl−/−::IRS1+/− ameliorated aging-like phenotypes of Kl−/− mice and improved survival (68).
IRS1 and IRS2 are required for the IIS pathway and are evolutionarily-conserved mammalian lifespan regulators (70). IRS1 binds to the IR, activating the PI3K-AKT signaling cascade to regulate glucose metabolism through down-regulation of gluconeogenic enzymes and up-regulation of lipogenic enzymes (71). Irs1−/− mice showed insulin-resistance with defects in insulin signaling (72), and lived 14–16% longer than WT littermates (73). The Irs1−/− mice showed small body size, reduced fat mass, and protection from aging-induced insulin resistance (70). Heterozygous Irs1+/− mice had the same lifespan as WT (70). The lifespan extension seen in the Irs1−/− mice was shorter than that of Snell (29–51%), Ames (37–68%), Little (23–25%), or GHR−/− (16–55%) dwarf mice strains. These lifespan differences suggest that there are IRS1-independent, but GH-dependent mechanism for longevity.
IRS2 is known to play a central role in pancreatic β-cell function (74). Irs2−/− mice die early (maximum 3 months of age) due to their diabetic phenotypes that include defects in hepatic insulin signaling, glucose deregulation, and β-cell failure or apoptosis (70). Several studies have tried to restore the β-cell function of Irs2−/− mice, through mechanisms that included overexpression of transcription factor, Pancreatic and duodenal homeobox 1 (PDX1) to enhance β-cell functions (75), deletion of Protein tyrosine phosphatase, non-receptor type 1 (PTPN1) to reduce insulin sensitivity (76), and reduction of Phosphatase and tensin homolog (PTEN) expression, a potent inhibitor of insulin action (77). These trials achieved some improvements in lifespan, but created additional problems. The lifespan of Irs2−/−::Pdx1 Tg mice increased by 15 months, but they had severe spinal deformities. Irs2−/−::Ptpn1−/− mice had lifespan increases of 8–9 months, but β-cell function deterioration. And Irs2−/−::Pten+/− mice gained 10–12 months of life, but had lymph proliferative disease (78). Heterozygous Irs2+/− mice showed conflicting lifespan results. Taguchi
FOXO transcription factors (FoxOs) are inhibitory downstream targets of IIS through activation of AKT, TSC complex subunit 2 (TSC2) and Glycogen synthase kinase 3 (GSK3) (83). The FoxOs are known as evolutionarily conserved anti-aging effectors (84) with broad roles in glucose homeostasis, tumor suppression, autophagy, and resistance to oxidative stress (84, 85). Among the four mouse FoxO homologs (FoxO1, FoxO3, FoxO4 and FoxO6), FoxO3 was reported to be a longevity factor under dietary restriction condition (86). Lifespan extension by overexpression of FoxOs has not been reported in mice yet. In fresh water polyp Hydra, FoxO achieves immortality through maintaining self-renewal capacity of stem cells (87).
A meta-analysis by Webb
Ames, Snell, and GHR−/− dwarf mice showed an obese phenotype due to their high capacity for lipid storage and activation of pre-adipocyte differentiation (90). Stout
Fat-specific insulin receptor knockout (FIR−/−) mice showed adipocyte-specific insulin resistance, reduced body fat mass, protection from age- and hyperphagia-associated obesity, and protection from age-related insulin resistance (93). There was an interesting difference in adipocyte size in FIR−/− mouse, compared to WT. Adipocytes isolated from epididymal WAT of FIR−/− mice divided into two sizes, small (< 75 μm) and large (> 100 μm), compared to one medium size (75–100 μm) in WT mice (93). Bluher
As mentioned previously, the Igf1r−/− mice were lethal because of postnatal respiratory failure (59). Kappeler
Brain-specific Irs2+/− (bIrs2+/−) and bIrs2−/− mice showed lifespan extension by 18% and 14%, respectively (79). During the aging process, these mice showed hyperinsulinemia and insulin resistance, as well as high glucose oxidation, and stable superoxide dismutase 2 (SOD2) levels in the hypothalamus, with stable diurnal rhythm (79). These results indicate that the metabolic rate, but not the obese state or insulin sensitivity might be associated with longevity. Sadagurski
In contrast to the aforementioned studies, Li and Ren argued that aging-associated decreases of circulating IGF1 stimulates cardiac aging. They reported that cardiac-specific Igf1 (cIgf1) Tg male mice showed a 60–80% increase in plasma IGF1 levels and extended their lifespan by 23% (98). cIgf1 Tg mice exhibited improved cardiomyocyte function including increased Ca2+ homeostasis, reduction of protein damage, and decreased apoptosis (98). Indeed, long-lived Ames dwarf mice showed impairment of cardiac excitation-contraction coupling, compared to WT mice (99). Overall, these results suggest that the autocrine/paracrine role of IGF1 is important to cardiac-specific anti-aging effects.
Both IIS and mechanistic target of rapamycin (mTOR) pathways could activate ribosomal protein S6 kinase 1 (S6K1), a regulator of growth and metabolism through activation of protein synthesis (100). Although male S6K1−/− mouse showed a similar lifespan to WT, female S6K1−/− mouse showed a 20% longer lifespan than WT. Other phenotypes of female S6K1−/− mice included lean and small body size, and improved insulin sensitivity (101). Interestingly, their total circulating IGF1, pituitary GH, TSH, and PRL levels were not changed compared to WT (101). Lamming
p66Shc, an isoform of SHC-transforming protein 1, binds to both IRS1 and S6 kinase and can bridge the two signalings (103). p66Shc−/− and p66Shc+/− mice showed 28% and 7% increased median lifespans, respectively (104). The p66Shc−/− mice showed less ROS production, high oxidative stress resistance, and prevention of obesity (104).
Myc transcription factor, known as a protooncogene regulates various downstream functions, including energy metabolism, ribosome biogenesis, cell cycle, apoptosis, differentiation, and stem cell maintenance (107). Recently, Hofmann
GH signal-deficient mice, including Snell, Ames, Little, GHR−/−, and Fgf21 Tg dwarf mice, showed increased lifespans and smaller body masses than WT mice. Therefore, body size was strongly dependent on GH action. This consistent trend suggests an inverse correlation between size and lifespan (109). However, small size can not be used as a general indicator of longevity, because Kl Tg, Irs2+/−, p66Shc−/−, and mtor+/−::mlst8+/− mice had normal body masses like WT mice, but showed longer lifespans than WT. In addition, GHA Tg mice had normal lifespans like WT mice, and Kl−/− and Irs2−/− mice showing dramatically shorter lifespans also had a dwarfism phenotype.
Snell, Ames, Little, and GHR−/− mice exhibited obesity, caused by loss of lipolytic and anti-lipogenic activities of GH. Especially, long-lived bIGF1R+/−, bIrs2+/−, and bIrs2−/− mice had high body fat. These results indicate brain-mediated endocrinological control of body fat and the IGF1 feedback loop. However, excess weight and obesity are known risk factors of aging and cause critical diseases, including diabetes and heart failure (110). Indeed, LI-Igf1−/−, FIR−/−, Irs1−/−, S6K1−/−, and p66Shc−/− mice showed less body fat than WT animals. In contrast, bGH Tg mice producing excess GH had lean bodies, but shortened lifespans. Surgical removal of visceral fat in GHR−/− mice decreased the fat-induced beneficial effects (92). Therefore, under the different endocrine circumstances, regardless of the body fat mass, adipose tissue itself contributes effects beneficial to longevity.
High insulin sensitivity could be associated with improved health. Long-lived GH signal-deficient mice, as well as FIR−/− and S6K1−/− mice, showed high insulin sensitivity. However, some long-lived mice with a mutation in IIS components, including LI-Igf1−/−, Igf1r+/−, bIGF1R+/−, and Irs1−/−, as well as the Kl Tg mice displayed insulin resistance. In the case of systemic Irs2 deficiency, Irs2−/− mice had diabetes, whereas brain-specific mutant bIrs2−/− mice showed both insulin resistance and longevity. Interestingly, R6/2::Irs2+/−::Irs2βTg mice, genetically modified to be Irs2+/− heterozygous except in β cells, showed a slow Huntington disease (HD) progression (97). In addition, insulin resistant (e.g., IR+/−) male mice showed a 20% extended maximum lifespan, but high insulin sensitivity mice, including PTP-1B−/− and PGC-1α Tg displayed shortened or similar lifespans compared to WT mice, respectively (62). It is clear that manipulation of insulin sensing signaling produces beneficial effects on longevity, however, either impaired or hyper-sensitive insulin signaling can lead to shortened lifespans.
Snell, Ames, Little, GHR−/−, and bIGF1R+/− mice showed both high-stress resistance and less tumor incidence. Pappa−/− mice also showed less tumor incidence, and Kl Tg, Igf1r+/− and p66Shc−/− mice showed high-stress resistance. Hepatic DEGs of Snell, Ames, Little, and GHR−/− mice showed up-regulation of DEGs for xenobiotic metabolism (e.g., Fmo3, CYPs, GSTs, and Sulfotransferases) and oxidative stress response (e.g., Hao2 and Hao3), representing improved oxidative stress response and toxin defense. In addition, similar hepatic DEG patterns were observed in DNA repair-deficient progeroid mice, and these changes might be a compensatory response (46). FoxO genes were up-regulated by inhibition of IIS signaling, and FoxOs regulated gene expression for stress response and DNA damage repair (111) (88). Genetically mTORC1-reduced mice, including mtor+/−;mlst8+/− and S6K1−/− showed extended lifespans with up-regulation of FoxOs (101). And p66Shc−/− mice showed less ROS production and high oxidative stress resistance (104). In addition, hepatic DEGs from Ames, Little, GHR−/−, and Irs1−/− mice commonly showed up-regulation of genes for mitochondrial respiration, including the TCA cycle, OXPHOS, and β-oxidation. Collectively, the gene expression analyses of IIS and mTORC1 reduction commonly represent improved stress response that is a signature of longevity. High resistance to oxidative stress and DNA damage in these mice might be contributing factors to low tumor incidence and increased longevity.
Snell, Ames, Little, GHR−/−, and Fgf21 Tg dwarf mice showed female-biased lifespan extension, and commonly showed STAT5-related gender-different hepatic gene expression. Moreover, female Igf1r+/−, LI-Igf1−/−, Irs1−/−, mtor+/−::mlst8+/−, S6K1−/−, and Myc+/− mice also lived longer than their male mice counterparts. These results suggest that the GH-IIS axis is regulated in a sex-specific manner. Gender is a critical factor in lifespan explained by genes that work differently in males and females in a concept known as sexual antagonistic pleiotropy (SAP) (112). STAT5-mediated liver gene expression analysis showed altered genes in xenobiotic metabolism (e.g., CYPs, GSTs, and sulfotransferases), oxidative stress response, and serine protease inhibitors. They overlap with the aforementioned longevity-associated liver gene expression data. In particular, Hsd3b5 encoding a ketosteroid reductase for steroid hormone biosynthesis, including testosterone was commonly down-regulated in Snell, Ames, Little, GHR−/−, and Fgf21 Tg mice. The expression of Hsd3b5 was associated with activation of GHR-STAT5 signaling, suggesting testosterone inactivation might contribute to the female-biased longevity.
On the other hand, Myc+/− mice showed up-regulated DEGs for xenobiotic metabolism and inhibition of mTOR and S6 kinases, however their tumor incidence was similar to WT mice (108). In addition, Kl Tg, bIGF1R+/−, and Irs1−/− mice showed similar lifespan extension in both males and females. Therefore, more work is needed to elucidate factors contributing to the lifespan of these mice.
This work was carried out with the support of Bio and Medical Technology Development Program through the National Research Foundation of Korea funded by the Ministry of Education, Science and Technology (NRF-2017R1D1A1A020 19355).
The authors have no conflicting interests.
Phenotypic characteristics of mice with mutations affecting the GH-IIS axis and lifespan
|Mice models||Key Physiology||Average Lifespan (%)||Body weight (%)||Body fat||Insulin sensitivity||Tumor incidence||Stress resistance||Plasma GH||Plasma IGF1||Plasma glucose||References|
|Snell (Pou1f1−/−)||GH, PRL and TSH deficiency||F+51 M+29 (11718806)||↓ (25–33)||↑||↑||↓||↑||↓||↓||↓||16577229 1371619|
|F and M+38 (15699523)||11718806|
|Ames (Prop1−/−)||GH, PRL and TSH deficiency||F+68 M+49 (8900272)||↓ (33)||↑||↑||↓||↑||↓||↓||↓||8900272 11719795|
|F and M+37 (11719795)|
|Little (Ghrhr−/−)||GH deficiency||F+25 M+23 (11371619)||↓ (50–67)||↑||↑||↓||↑||↓||↓||↓||1270792 11371619|
|Ghr−/−||GH resistance||F+38 M+55 (10875265)||↓ (41–50)||↑||↑||↓||↑||↑||↓||↓ or ↔||10875265|
|F+25 M+16 (12933651)||12933651|
|F+30 M+30 (16682650)||16682650|
|Fgf21 Tg*||GH resistance||F+40 M+30 (23066506)||↓ (60)||↔||↑||?||?||↑||↓||↓||18585098|
|LI-Igf1−/−||IGF1 reduction 75–85%||F+16 (21799924)||↓ (75–100)||↓||↓||?||?||↑||↓||↔||10359843|
|Heart Igf1 Tg*||IGF1 excessed||M+23a (17973971)||↔ (100)||?||?||?||?||?||↑||↔||17973971|
|Pappa−/−||IGF1 reduced||F and M+38 (17681037)||↓ (40)||?||↔||↓||?||↔||↔||↔||14973274|
|F+29 M+24 (20351075)||17681037|
|Klotho Tg||IIS inhibition||F+19 M+31 (16123266)||↔ (100)||?||F ↔||?||↑||?||?||↔||16123266|
|F+19 M+20 (16123266)||M↓|
|Igf1r+/−||IGF1 resistance||F+33 M NS (12483226)||↓ (90)||?||↓||↔||↑, F ↑ M||?||↔||↔||12483226|
|F and M NS (22132081)||↔||22132081|
|Brain IGF1R+/−*||Brain IIS reduction||F+8 M+13b (18959478)||↓ (90–92)||↑||↓||↓||↑||↓||↓||↔||18959478|
|FIR−/−||Fat-specific insulin resistance||F and M+11 (12543978)||↓ (70)||↓||↑||?||?||?||?||↔||12110165|
|Irs1−/−*||Insulin resistance or normal||F+16 M+14 (21283571)||↓ (70)||↓||↓||?||?||↔||↔||↔||7969452 17928362|
|Irs2+/−||IRS2 reduced||F and M+17 (17641201)||↔ (100)||?||↑ or ↔||?||?||↔||↔||↓||17641201|
|F and M NS (17928362)||17928362|
|Brain Irs2+/−||Brain IRS2 reduced||F and M (+/−)+18 (17641201)||↔ (100)||↑||↓||?||?||↔||↔||?||17641201|
|Brain Irs2−/−||F and M (−/−)+14 (17641201)|
|mtor+/−::mlst8+/−*||mTORC1 reduction 50%||F+14 M NS (22461615)||↔ (100)||?||↔||?||?||?||?||↔||22461615|
|S6K1−/−*||S6K1 deficient||F+20 M NS (19797661)||↓ (48)||↓||↑||?||?||↔||↔||↔||19797661|
|p66Shc+/−||IRS inhibition, S6K1 inhibition||F and M (+/−)+7a (10580504)||↔ (100)||↓||↔||?||↑||?||?||↔||10580504|
|p66Shc−/−||F and M (−/−)+28a (10580504)||20624962|
|Myc+/−*||IGF1 reduction and MYC-regulated genes||F+21 M+11a (25619689)||↓ (81)||↔||?||↔||?||?||↓||?||25619689|
|Normal or short-lived models|
|GHA Tg||GH resistance||NS (12933651)||↓ (55–70)||↑||↑||↓||?||?||↓||↓ or ↔||1791834 11487276|
|Bovine GH Tg||GH excessed||M-45 (14583653)||↑ (200)||↓||↓||↑||?||↑||↑||↓||1791834 11701438|
|LID||75% reduced IGF1||M-? (22421704)||↔ (100)||↑||↓||↓||?||↑||↓||↔||10377413|
|Kl−/−*||IIS activation, growth retardation||60.7 daysc (9363890)||↓ (44–47)||?||↓||?||?||?||?||?||9363890|
|Irs2−/−||Irs2 deletion||F-26 M-84 (9495343)||↓ (90)||?||↓||?||?||?||?||↑||9495343|
↑ up-regulated; ↓ down-regulated; ↔ no difference; ?, no data available.
NS, not significant.
Transcriptome studies from long-lived mouse models for the GH-IIS axis
|Genotype||Publication||Design (age: month)||Study Focus||Sex||Back-ground||Tissue||Platform (probesets)||DEG number (statistical criteria)|
|Snell||Dozmorov et al. 2002 11867646||WT vs Snell (6)||Systemic GH effects||Male||(DW × C3H)F1||Liver||(2,352) Clontech Mouse Atlas 1.2 array||17 (t-test FDR < 0.05)|
|Snell||Boylston et al. 2004 15379852||WT vs Snell (3–5, 24–28)||Systemic GH effects||Male||C3H/HeJ × DW/J F1 or C3DW||Liver||(12,422) Affymetrix MG U74Av2 array||> 50 (t-test, P < 0.05 and FC > 2)|
|Snell||Papaconstantinou et al. 2005 15888324||Snell ctrl(+/?)vs Snell (3–6, 20–28)||IGF-1 axis protein levels||Male||C3H/HeJ × DW/J F1 or C3DW||Liver||(12,422) Affymetrix MG U74Av2 array||17 (t-test P < 0.05)|
|Snell||Boylston et al. 2006 19943135||Snell ctrl(+/?)vs Snell (4–6, 24–26)||Systemic GH effects||Male||C3H/HeJ × DW/J F1 or C3DW||Liver||(12,422) Affymetrix MG U74Av2 array||205 (t-test FDR < 0.05)|
|Ames||Dozmorov et al. 2001 11213270||WT vs Ames (5, 13, 22)||Systemic GH effects||?||?||Liver||(265) early version of spotting array||13 (t-test P < 0.01)|
|Ames||Tsuchiya et al. 2004 15039484||WT vs Ames (2)||Systemic GH effects||Female||?||Liver||(12,422) Affymetrix MG U74Av2 array||212 (two-way ANOVA FDR < 0.05)|
|Ames||Amador-Noguez et al. 2004 15569359||WT littermate vs Ames (3, 6, 12, 24)||Systemic GH effects agaist wt aging patterns||Male||C57Bl/6||Liver||(> 14,000) Affymetrix MOE340A array||122 (t-test Bonferroni <0.05 and FC > 2)|
|Ames||Boylston et al. 2006 19943135||Ames ctrl(+/?)vs Ames (4–6, 12–14, 24–27)||Systemic GH effects||Male||C57Bl/6||Liver||(> 34,000) Affymetrix MG 430 2.0 arrays||785 (t-test FDR < 0.05)|
|Little||Amador-Noguez et al. 2004 15569359||WT vs Little mice (3,6,12,24)||Systemic GH effects agaist wt aging patterns||Male||C57Bl/6||Liver||(> 14,000) Affymetrix MOE340A array||104 (t-test Bonferroni <0.05 and FC > 2)|
|bGH Tg||Olsson et al. 2003 12736163||WT vs bGH-Tg (6)||Systemic GH effects||Male||?||Liver||(11,000) Affymetrix mouse 11ka and 11kb arrays||45 (difference call parameter 3–4)|
|Ghr||Miller et al. 2002 12403853||WT vs Ghr−/− (8–9)||Systemic GH effects||?||?||Liver||(2,352) Clontech Mouse Atlas 1.2 array||10 (t-test FDR < 0.05)|
|Ghr−/−||Rowland et al. 2005 15601831||WT vs Ghr−/− (1.4)||Systemic GH effects||Male||B6.C3H-6T||Liver||(12,422) Affymetrix MG U74Av2 array||398 (t-test P < 0.0005 and FC > 1.5)|
|GHR-569, GHR-391||Rowland et al. 2005 15601831||WT vs GHR-569, GHR-391 (1.4)||Systemic STAT5 effects||Male||129/SVJ||Liver||(12,422) Affymetrix MG U74Av2 array||20 (t-test P < 0.0005 and FC > 1.5)|
|Ghr−/−, GHR-391||Barclay et al. 2011 21084450||WT vs Ghr−/−, GHR-391 (4)||STAT5-mediated effects||Male||C57BL/6J||Liver||(46,632) Illumina MouseWG-6 v1.1 array||55 common genes out of 466 (t-test P < 0.05, FC > 2)|
|Ghr−/−||Stout et al. 2015 26436954||WT vs Ghr−/− (6)||Systemic GH effects on WAT and BAT||?||?||WAT and BAT||(28,853) Affymetrix GeneChip Mouse Gene 1.0 ST Array||346 in SubQ WAT, 319 in PERI WAT, 192 in EPI WAT, 280 in BAT (t-test FDR < 0.05 and FC >1.5)|
|Ghr−/−||Chang et al. 2016 27064376||WT vs Ghr−/− (3.7–4.2)||Systemic GH effects on lncRNA and miRNA||Male and Female||C57Bl/b5||Liver||(25,376 transcript, 31,423 lncRNAs, 3,1000 miRNAs) Agilent Mouse LncRNA Array v2.0||3,442 lncRNAs, 1,750 mRNAs, 30 miRNAs in female, 2,642 lncRNAs, 1,489 mRNAs, 57 miRNAs in male (FC≥2)|
|Fgf21-Tg||Zhang et al. 2012 23066506||WT vs FGF21−/− (3)||Systemic FGF21 effects||Male||C57Bl/6J||Liver, WAT, Muscle||(> 25,600) Illumina mouseRef-8 v2.0 BeadChip||33 in liver, 8 in muscle, 22 in fat (t-test FDR < 0.1 and FC > 2)|
|Sex-specific liver gene expression|
|Ames||Amador-Noguez et al. 2005 15925325||WT vs Ames (3–6)||Sexual dimorphism of GH effects||Male vs Female||C57Bl/6||Liver||(> 14,000) Affymetrix MOE430A array||381 sex-independent, 110 sex-dependent (two-way ANOVA P < 0.001 and FC > 1.5)|
|STAT5b−/−||Clodfelter et al. 2006 16469768||WT vs STAT5b−/− (1.8–2.1)||Sexual dimorphism of STAT5b effects||Male vs Female||?||Liver||(23,574) Agilent Sosetta/Merck Mouse TOE 75k array||85 in female (t-test P <0.05, FC > 1.5)|
|STAT5a−/−||Clodfelter et al. 2007 17536022||WT vs STAT5a−/− (1.4–2.1)||Sexual dimorphism of STAT5a effects||Male vs Female||129 x Black/Swiss||Liver||(23,574) Agilent Sosetta/Merck Mouse TOE 75k array||769 in male, 461 in female (t-test P < 0.05, FC > 1.5)|
|IGF1- and IIS-reduced mice|
|FIR−/−||Bluher et al. 2004 15131119||WT vs WAT (3)||Fat-specific IR & Adipocyte size effect||Male||C57BL/6 x 129/Sv||WAT||(12,488) Affymetric MG-U74A-v2 array||48 for adipocyte size, 63 for IR signaling (t-test P <0.001 and µ1-µ2 > 2∑SD)|
|FIR−/−||Katic et al. 2007 18001293||WT vs FIR−/− (6, 18, 30–36)||Fat-specific IR effect||Male||129sv/C57Bl6/J||WAT||(22,690) Affymetrix mouse 430A array||? (ANOVA P < 0.01)|
|Irs1−/−||Selman et al. 2008 17928362||WT vs Irs1−/− (2.6, 15, 22)||Systemic IIS effects||Female||C57BL/6||Liver||(> 34,000) Affymetrix MG 430 2.0 array||? (t-test P < 0.05)|
|Irs1−/−||Page et al. 2018 29779018||WT vs Irs1−/− (15)||Systemic IIS effects||Female||C57BL/6||Liver, Muscle, Brain, WAT||RNA-sequencing by Thermo Fisher Ion Proton system||124 (101, specific) in liver, 185 (137) in muscle, 111 (94) in brain, 344 (319) in WAT (t-test FDR < 0.1)|
|mTORC1- and MYC-reduced mice|
|S6K1−/−||Selman et al. 2009 19797661||WT vs S6K1−/− (20)||Systemic S6K1 effect||Female||C57BL/6||Liver, Muscle, WAT||(> 38,000) Affymetrix MG 430 2.0 array||1,843 in liver, 2,309 in muscle, 1,970 in WAT (q-value < 0.01)|
|p66Shc−/−||Tomilov et al. 2010 19892704||WT vs p66Shc−/− (3, 12)||Systemic p66Shc effect||Male and Female||?||Liver, Spleen, Lungs, WAT, peritoneal macrophage||(> 38,000) Affymetrix MG 430 2.0 array
||top 250 genes (t-test P <0.05)|
|Myc+/−||Hofmann et al. 2015 25619689||WT vs Myc+/− (5, 24)||Systemic MYC effects||Male||C57BL/6NCrl||Liver, Muscle, WAT||(28,853) Affymetrix Mouse 1.0 Gene ST arrays||307 in liver, 160 in muscle, 412 in WAT (t-test FDR < 0.05 and FC > 1.5)|