Lifespan extension by increased expression of the Drosophila homologue of the IGFBP7 tumour suppressor

Mammals possess multiple insulin-like growth factor (IGF) binding proteins (IGFBPs), and related proteins, that modulate the activity of insulin/IGF signalling (IIS), a conserved neuroendocrine signalling pathway that affects animal lifespan. Here, we examine if increased levels of an IGFBP-like protein can extend lifespan, using Drosophila as the model organism. We demonstrate that Imaginal morphogenesis protein-Late 2 (IMP-L2), a secreted protein and the fly homologue of the human IGFBP7 tumour suppressor, is capable of binding at least two of the seven Drosophila insulin-like peptides (DILPs), namely native DILP2 and DILP5 as present in the adult fly. Increased expression of Imp-L2 results in phenotypic changes in the adult consistent with down-regulation of IIS, including accumulation of eIF-4E binding protein mRNA, increase in storage lipids, reduced fecundity and enhanced oxidative stress resistance. Increased Imp-L2 results in up-regulation of dilp2, dilp3 and dilp5 mRNA, revealing a feedback circuit that is mediated via the fly gut and/or fat body. Importantly, over-expression of Imp-L2, ubiquitous or restricted to DILP-producing cells or gut and fat body, extends lifespan. This enhanced longevity can also be observed upon adult-onset induction of Imp-L2, indicating it is not attributable to developmental changes. Our findings point to the possibility that an IGFBP or a related protein, such as IGFBP7, plays a role in mammalian aging.     

SCREENING OF PHOTOSYNTHETIC O2‐EVOLVING PROKARYOTES FOR AN INSULIN‐LIKE ANTIGEN1

Diabetes mellitus (DM), a metabolic disorder, is becoming a major health problem worldwide. Insulin is the single hope for management of type 1 diabetes, but it is not always available or suitable. For finding additional bioresources, the present study was performed. ELISA‐based preliminary screening of cyanobacterial biomass using antihuman insulin antibody have detected an insulin‐like antigen in Spirulina platensis S‐5, Spirulina NCCU‐482, and Spirulina NCCU‐483. Their similarity with insulin‐like antigen was further confirmed by electrophoretic mobility using bovine insulin as marker.     

The association of phosphoinositide 3-kinase enhancer A with hepatic insulin receptor enhances its kinase activity

Dysfunction of hepatic insulin receptor tyrosine kinase (IRTK) causes the development of type 2 diabetes. However, the molecular mechanism regulating IRTK activity in the liver remains poorly understood. Here, we show that phosphoinositide 3-kinase enhancer A (PIKE-A) is a new insulin-dependent enhancer of hepatic IRTK. Liver-specific Pike-knockout (LPKO) mice display glucose intolerance with impaired hepatic insulin sensitivity. Specifically, insulin-provoked phosphoinositide 3-kinase/Akt signalling is diminished in the liver of LPKO mice, leading to the failure of insulin-suppressed gluconeogenesis and hyperglycaemia. Thus, hepatic PIKE-A has a key role in mediating insulin signal transduction and regulating glucose homeostasis in the liver.     

Human insulin A‐chain peptide analog(s) with in vitro biological activity

In a previous study, we showed that a synthetic human insulin 1‐chain analog, named analog ( 3 ) was capable of mimicking in vitro effects of native insulin, including stimulation of cell proliferation, glucose uptake and glycogen synthesis. Here, we have synthesized three new analogs ( 6, 9, 12 ) of the human A‐chain, bearing or not their N‐ or C‐terminal residue, to determine the structural features which are responsible for their biological properties. In vitro experiments clearly demonstrated that the N‐terminal part of the peptides is required for the biological activity of the molecules, suggesting its crucial role in the mechanism underlying the cellular effect. Our findings may help to better understand the mechanism of interaction between insulin and its receptor. In addition, the present data demonstrate that some mini‐insulin derived from the A‐chain can exert similar effects as native insulin. These small peptides may offer specific advantages over insulin in the definition of new strategies for diabetes treatment. Copyright © 2009 John Wiley & Sons, Ltd. This article was published online on 17 July 2009. An error was subsequently identified. This notice is included in the online and print versions to indicate that both have been corrected 4 August 2009.     

Structural and biochemical characterization of native and recombinant single insulin-like growth factor-binding domain protein (SIBD-1) from the Central American hunting spider Cupiennius salei (Ctenidae)

Cupiennius salei single insulin-like growth factor binding domain protein (SIBD-1) is an 8.6 kDa Cys-, Pro-, and Gly-rich protein, discovered in the hemocytes of the Central American hunting spider Cupiennius salei. SIBD-1 exhibits high sequence similarity to the N-terminal domain of the insulin-like growth factor-binding protein superfamily and has been reported to play an important role in the spider's immune system. Here, the recombinant expression and the elucidation of the three-dimensional structure of recombinant SIBD-1 and the characterization of the sugar moiety at Thr2 of native SIBD-1 is described in detail.     

Synthesis of relaxin‐2 and insulin‐like peptide 5 enabled by novel tethering and traceless chemical excision

This report presents an entirely chemical, general strategy for the synthesis of relaxin‐2 and insulin‐like peptide 5. Historically, these two peptides have represented two of the more synthetically challenging members of the insulin superfamily. The key synthetic steps involve two sequential oxime ligations to covalently link the individual A‐chain and B‐chain, followed by disulfide bond formation under aqueous, redox conditions. This is followed by two chemical reactions that employ diketopiperazine cyclization‐mediated cleavage and ester hydrolysis to liberate the connecting peptide and the heterodimeric product. This approach avoids the conventional iodine‐mediated disulfide bond formation and enzyme‐assisted proteolysis to generate biologically active two‐chain peptides. This novel synthetic strategy is ideally suited for peptides such as relaxin and insulin‐like peptide 5 as they possess methionine and tryptophan that are labile under strong oxidative conditions. Additionally, these peptides possess multiple arginine and lysine residues that preclude the use of trypsin‐like enzymes to obtain biologically active hormones. This synthetic methodology is conceivably applicable to other two‐chain peptides that contain multiple disulfide bonds. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd.     

Normal Insulin Sensitivity in Lean Offspring of Obese Parents

Objective: Offspring of diabetic or hypertensive patients are insulin resistant at a prediabetic/prehypertensive stage. We tested the hypothesis that insulin action may be impaired in the offspring of obese nondiabetic parents. Research Methods and Procedures: Twenty‐one lean offspring of nonobese subjects [(OL) 22 ± 3 years of age] were matched to 23 lean offspring of obese subjects (OOb) by gender distribution, age, BMI, and waist circumference. Anthropometry, oral glucose tolerance, in vivo insulin sensitivity [by a euglycemic insulin clamp (6 pmol/min per kilogram_FFM; where FFM represents fat‐free mass)], and thermogenesis (by indirect calorimetry) were measured in each subject. The study subjects were from a population of 267 nuclear families (one offspring and both his/her parents) in which there was statistically significant (χ² = 30.2, p = 0.001) concordance of BMI between parents and offspring. Results: In comparing OOb with OL, no statistically significant difference or trend toward a difference was detected in fasting plasma glucose and insulin concentrations, glucose and insulin responses to oral glucose, insulin sensitivity [metabolism value = 45 ± 12 (OOb) vs. 47 ± 17 μmol/min per kilogram_FFM (OL)], insulin‐induced inhibition of protein and lipid oxidation, stimulation of glucose oxidation and nonoxidative glucose disposal, respiratory quotient, resting energy expenditure, and glucose‐induced thermogenesis. Discussion: The metabolic similarity between lean offspring of obese parents and those of nonobese parents suggests that insulin resistance and its correlates are not co‐inherited with the predisposition to develop obesity.     

IRS-2 branch of IGF-1 receptor signaling is essential for appropriate timing of myelination

Insulin-like growth factor (IGF)-1 increases proliferation, inhibits apoptosis and promotes differentiation of oligodendrocytes and their precursor cells, indicating an important function for IGF-1 receptor (IGF-1R) signaling in myelin development. The insulin receptor substrates (IRS), IRS-1 and -2 serve as intracellular IGF-1R adaptor proteins and are expressed in neurons, oligodendrocytes and their precursors. To address the role of IRS-2 in myelination, we analyzed myelination in IRS-2 deficient (IRS-2(-/-)) mice and age-matched controls during postnatal development. Interestingly, expression of the most abundant myelin proteins, myelin basic protein and proteolipid protein was reduced in IRS-2(-/-) brains at postnatal day 10 (P10) as compared to controls. myelin basic protein immunostaining in P10-IRS-2(-/-) mice revealed a reduced immunostaining, but an unchanged regional distribution pattern. In cerebral myelin isolates at P10 unaltered relative expression of different myelin proteins was found, indicating quantitatively reduced but not qualitatively altered myelination. Interestingly, up-regulation of IRS-1 expression and increased IGF-1R signaling were observed in IRS-2(-/-) mice at P10-14, indicating a compensatory mechanism to overcome IRS-2 deficiency. Adult IRS-2(-/-) mice showed unaltered myelination and motor function. Furthermore, in neuronal/brain-specific insulin receptor knockout mice myelination was unchanged. Thus, our experiments reveal that IGF-1R/IRS-2 mediated signals are critical for appropriate timing of myelination in vivo.     

Relationship of plasma adiponectin with sex hormone and insulin-like growth factor levels

Objective: Recent studies have suggested that a relationship between adiponectin and sex hormone, prolactin, and insulin‐like growth factor levels could be important for breast cancer risk and insulin sensitivity. Therefore, we assessed the relationship of adiponectin with plasma concentrations of estrone; estradiol; estrone sulfate; testosterone; androstenedione; dehydroepiandrosterone (DHEA); dehydroepiandrosterone sulfate (DHEAS); sex hormone binding globulin (SHBG); prolactin; insulin‐like growth factor (IGF‐1); its binding protein, IGF binding protein 3 (IGFBP‐3); c‐peptide; and IGF binding protein 1 (IGFBP‐1) among 360 postmenopausal women not taking postmenopausal hormones from the Nurses’ Health Study. Research Methods and Procedures: Multivariate models were adjusted for physical activity, alcohol consumption, age at blood draw, age at first birth/parity, fasting status, and time of day of blood draw; a separate model was additionally adjusted for BMI at blood draw. Results: Estrogens were inversely associated with adiponectin levels; however, except for free estradiol, these associations were substantially attenuated after adjustment for BMI. Free estradiol levels were 27% lower among women in the top vs. bottom quartile of adiponectin levels. No consistent associations were observed for the androgens, prolactin, IGF‐1, and IGFBP‐3. However, SHBG, c‐peptide, and IGFBP‐1 were strongly and independently associated with adiponectin levels (r = 0.29, ?0.30, 0.24, respectively). Conclusion: With the exceptions of SHBG, c‐peptide, and IGFBP‐1, the studied analytes were modestly associated with adiponectin and the associations were, in large part, mediated by body fat.     

Impaired IGF1R signaling in cells expressing longevity-associated human IGF1R alleles

Dampening of insulin/insulin-like growth factor-1 (IGF1) signaling results in the extension of lifespan in invertebrate as well as murine models. The impact of this evolutionarily conserved pathway on the modulation of human lifespan remains unclear. We previously identified two IGF1R mutations (Ala-37-Thr and Arg-407-His) that are enriched in Ashkenazi Jewish centenarians as compared to younger controls and are associated with the reduced activity of the IGF1 receptor as measured in immortalized lymphocytes. To determine whether these human longevity-associated IGF1R mutations affect IGF1 signaling, we engineered mouse embryonic fibroblasts (MEFs) expressing the different human IGF1R variants in a mouse Igf1r null background. The results indicate that MEFs expressing the human longevity-associated IGF1R mutations attenuated IGF1 signaling, as demonstrated by significant reduction in phosphorylation of both IGF1R and AKT after IGF1 treatment, in comparison with MEFs expressing the wild-type IGF1R. The impaired IGF1 signaling caused by the IGF1R mutations resulted in the reduced induction of the major IGF1-activated genes in MEFs, including EGR1, mCSF, IL3Rα, and TDAG51. Furthermore, the IGF1R mutations caused a delay in cell cycle progression after IGF1 treatment, indicating a dysfunctional physiological response to a cell proliferation signal. These results demonstrate that the human longevity-associated IGF1R variants are reduced-function mutations, implying that dampening of IGF1 signaling may be a longevity mechanism in humans.     

Serum insulin-like growth factor-I, insulin-like growth factor binding protein-3, and the pubertal growth spurt in the female rhesus monkey

While there is good evidence suggesting IGF-I links to pubertal development and crown-rump length growth among rhesus monkeys, linkages between IGF-I and other measures of morphological growth have not been established. In this study, the pubertal growth spurt in a number of morphological characteristics of female rhesus monkeys is related to serum endocrine status of insulin-like growth factor-I (IGF-I) and its binding protein, insulin-like growth factor binding protein-3 (IGFBP-3), to test the hypothesis that elevations in IGF-I and IGFBP-3 coincide with the time of greatest growth rate of different morphological characteristics. A longitudinal study of pubertal growth among four female rhesus monkeys was carried out across a 3-year period. Morphometric measurements included weight, crown-rump length, foot-length, and skinfolds at five sites (biceps, triceps, abdominal, subscapular, and suprailiac). These measures were taken as being representative of total mass, skeletal growth of the trunk and head, limb length, and body fatness, respectively. Measurements were carried out as closely as possible to 3-monthly, with interpolations being performed to standardise the data to exactly 3-monthly intervals for all individuals. Blood samples were taken at time of morphometry. Elevations in serum IGF-I and IGFBP-3 took place in a manner similar to that of humans, and across the period associated with onset of puberty. Mean 3-monthly gain in crown-rump length and foot length showed significant peaks across the measurement period, while mean 3-monthly gains in weight and sum of five skinfolds did not. Greatest foot length gain occurred on average between 3-3.5 years of age, while crown-rump length gain was greatest between 3.75-4 years of age. Periods of greatest gain in crown-rump length and foot length took place across the period of elevated serum IGF-I levels, which was between 3-4.5 years of age. Significant elevations in IGF-I and IGFBP-3 were not coincident with greatest gains in foot length or crown-rump length. Thus the hypothesis does not hold true for the two measures showing significant peaks in 3-monthly gain across the measurement period. The nature of the endocrine impact on macaque morphology remains unclear, although this may be fundamental to the understanding of the variation in the pubertal growth spurt and its influence on morphology at maturity both within and across primate species.     

Expression of insulin‐like growth factor‐1 receptor in metastatic uveal melanoma and implications for potential autocrine and paracrine tumor cell growth

We investigated the importance of the insulin‐like growth factor‐1 receptor (IGF‐1R) in hepatic metastases of uveal melanoma. The expression pattern of IGF‐1R in archival tissue samples of hepatic metastasis from 24 patients was analyzed by immunohistochemistry. All the samples of hepatic metastases stained positive for IGF‐1R. To investigate the biological role of IGF‐1R on the growth of metastatic uveal melanoma, a long‐term cell line obtained from a hepatic metastasis (TJU‐UM001) was evaluated. TJU‐UM001 expressed cell surface IGF‐1R (>90%) and proliferated in response to exogenous and endogenous insulin‐like growth factor‐1 (IGF‐1). Correlatively, anti‐IGF‐1R antibody completely blocked IGF‐1‐induced growth of TJU‐UM001 cells. IGF‐1 preferentially induced phosphorylation of Akt (S473) in quiescent TJU‐UM001 cells, and this was blocked by anti‐IGF‐1R antibody. This study suggests that autocrine and paracrine mechanisms underlie IGF‐1‐induced growth of metastatic uveal melanoma and underscore the potential benefit of IGF‐1 or IGF‐1R antagonism in treatment for metastatic uveal melanoma.     

Biotin,coenzyme Q10, and their combination ameliorate aluminium chloride‐induced Alzheimer's disease via attenuating neuroinflammation and improving brain insulin signaling

Insulin is important for brain function and neuronal survival. Insulin signaling is initiated by the phosphorylation of insulin receptor substrate‐1 (IRS‐1) at tyrosine (pTyr) residue. However, IRS‐1 is inhibited by phosphorylation at serine (pSer). In Alzheimer's disease (AD), oxidative stress and accumulation of amyloid beta (Aβ) induce neuroinflammation, which augments pSer‐IRS‐1 and reduces pTyr‐IRS‐1 disturbing insulin signaling pathway. Coenzyme Q10 (CoQ10) and biotin possess antioxidant and anti‐inflammatory properties, and, in this study, their impact on insulin signaling is investigated in an aluminium chloride (AlCl₃) model of AD. AD was induced by oral administration of AlCl₃ (75 mg/kg) for 60 days. Biotin (2 mg/kg), CoQ10 (10 mg/kg), and their combination were supplemented concomitantly with AlCl₃ for 60 days. Memory test and histological examination were performed. Brain levels of lipid peroxides, antioxidants (reduced glutathione and superoxide dismutase), inflammatory markers (tumor necrosis factor‐α, interleukin‐6 [IL‐6], IL‐1, and nuclear factor κB), and phosphorylated Akt (survival kinase) as well as protein levels of Aβ, IRS‐1 (pTyr and pSer), and caspase‐3 (apoptotic marker) were determined. AlCl₃ resulted in impaired memory, significant increase in Aβ, lipid peroxides, inflammatory markers, caspase‐3, and pSer‐IRS‐1, with significant reduction of the antioxidants, pTyr‐IRS‐1, and p‐Akt reflecting Aβ‐induced inflammation and defective insulin signaling. Histological examination revealed focal aggregations of inflammatory cells and neuronal degeneration. The biochemical deviations and histological changes were attenuated by the concomitant treatment with biotin and, to greater extent, with CoQ10 and the combination. In conclusion, biotin and CoQ10 could protect against AD via attenuating inflammatory response and enhancing insulin signaling.     

Effects of citraconylation on enzymatic modification of human proinsulin using trypsin and carboxypeptidase B

Insulin is a polypeptide hormone which is produced by the β‐cell of pancreas and controls the blood glucose level in the human body. Enzymatic modification of human proinsulin using trypsin and carboxypeptidase B generally causes high accumulation of insulin derivatives, leading to more complicated purification processes. A simple method including citraconylation and decitraconylation in the enzymatic modification process was developed for the reduction of a major derivative, des‐threonine human insulin. Addition of 3.0 g citraconic anhydride per g protein into the reaction solution led to the citraconylation of lysine residues in human proinsulin and reduction of relative des‐threonine insulin content from 13.5 to 1.0%. After the enzymatic hydrolysis of the citraconylated proinsulin, 100% of lysine residues can be decitraconylated and restored by adjusting pH to 2–3 at 25 °C. Combination of hydrogen peroxide addition and citraconylation of proinsulin expressed in recombinant Escherichia coli remarkably improved the conversion yield of insulin from 52.7 to 77.7%. Consequently, citraconylation of lysine residues blocked the unexpected cleavage of human proinsulin by trypsin, minimized the formation of des‐threonine insulin and hence increased the production yield of active insulin. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009     

Serum concentrations of insulin‐like growth factor‐i and insulin‐like growth factor binding protein‐2 and ‐3 in eight hoofstock species

The somatotropic axis, which includes growth hormone, insulin‐like growth factor (IGF)‐I, and IGF binding proteins (IGFBP), is involved in the regulation of growth and metabolism. Measures of the somatotropic axis can be predictive of nutritional status and growth rate that can be utilized to identify nutritional status of individual animals. Before the somatotropic axis can be a predictive tool, concentrations of hormones of the somatotropic axis need to be established in healthy individuals. To begin to establish these data, we quantified IGF‐I, IGFBP‐2, and IGFBP‐3 in males and females of eight threatened hoofstock species at various ages. Opportunistic blood samples were collected from Bos javanicus (Java banteng), Tragelaphus eurycerus isaaci (bongo), Gazella dama ruficollis (addra gazelle), Taurotragus derbianus gigas (giant eland), Kobus megaceros (Nile lechwe), Hippotragus equines cottoni (roan antelope), Ceratotherium simum simum (white rhinoceros), and Elephas maximus (Asian elephant). Serum IGF‐I and IGFBPs were determined by radioimmunoassay and ligand blot, respectively. Generally, IGF‐I and IGFBP‐3 were greater in males, and IGFBP‐2 was greater in females. In banteng (P = 0.08) and male Nile lechwe (P<0.05), IGF‐I increased with age, but decreased in rhinoceros (P = 0.07) and female Nile lechwe (P<0.05). In banteng, IGFBP‐3 was greater (P<0.01) in males. In elephants (P<0.05) and antelope (P = 0.08), IGFBP‐2 were greater in females. Determination of concentrations of hormones in the somatotropic axis in healthy animals makes it possible to develop models that can identify the nutritional status of these threatened hoofstock species. Zoo Biol 30:275–284, 2011. © 2010 Wiley‐Liss, Inc.     

Regulation of adenosine receptors expression in rat B lymphocytes by insulin

Development of diabetes is associated with altered expression of adenosine receptors (ARs). Some of these alterations might be attributed to changes in insulin concentration. This study was undertaken to investigate the possible insulin effect on ARs level, and to determine the signaling pathway utilized by insulin to regulate the expression of ARs in rat B lymphocytes. Western blot analysis of B lymphocytes protein extracts indicated that all four ARs were present at detectable levels in the cells cultured for 24 h without insulin (≤10^?11 M), although the protein band of A_2A‐AR was barely visible. Inclusion of insulin (10^?8 M) in the culture medium resulted in an increase of A₁‐AR and A_2A‐AR protein levels and a significant decrease of A_2B‐AR protein, whereas the protein level of A₃‐AR remained unchanged. Alterations in the ARs protein content were accompanied by changes in the ARs mRNA levels. Increase of the insulin concentration from 10^?11 to 10^?8 M resulted in 50% decrease of A_2B‐AR mRNA level and two‐, and threefold increase of A₁‐AR and A_2A‐AR mRNA levels, respectively. Pretreatment of B cells with cycloheximide completely blocked the insulin action on A₁‐AR and A_2A‐AR mRNA, but not on A_2B‐AR expression. Detailed pharmacological analysis demonstrated that insulin‐induced A₁‐AR and A_2A‐AR mRNA expression through the Ras/Raf‐1/MEK/ERK pathway. The insulin effect on A_2B‐AR expression was blocked by p38 MAP kinase inhibitor (SB 203580). Concluding, elevated insulin concentration differentially affects the expression of ARs in B lymphocytes in a fashion that might enhance the various immunomodulatory effects of adenosine. J. Cell. Biochem. 109: 396–405, 2010. © 2009 Wiley‐Liss, Inc.