Insulin Receptor Activation Research Articles

Pooled genome-wide CRISPR activation screening for rapamycin resistance genes in Drosophila cells.

Loss-of-function and gain-of-function genetic perturbations provide valuable insights into gene function. In Drosophila cells, while genome-wide loss-of-function screens have been extensively used to reveal mechanisms of a variety of biological processes, approaches for performing genome-wide gain-of-function screens are still lacking. Here, we describe a pooled CRISPR activation (CRISPRa) screening platform in Drosophila cells and apply this method to both focused and genome-wide screens to identify rapamycin resistance genes. The screens identified three genes as novel rapamycin resistance genes: a member of the SLC16 family of monocarboxylate transporters (CG8468), a member of the lipocalin protein family (CG5399), and a zinc finger C2H2 transcription factor (CG9932). Mechanistically, we demonstrate that CG5399 overexpression activates the RTK-Akt-mTOR signaling pathway and that activation of insulin receptor (InR) by CG5399 requires cholesterol and clathrin-coated pits at the cell membrane. This study establishes a novel platform for functional genetic studies in Drosophila cells.

Novel Isoform DTX3c Associates with UBE2N-UBA1 and Cdc48/p97 as Part of the EphB4 Degradation Complex Regulated by the Autocrine IGF-II/IRA Signal in Malignant Mesothelioma.

EphB4 angiogenic kinase over-expression in Mesothelioma cells relies upon a degradation rescue signal provided by autocrine IGF-II activation of Insulin Receptor A. However, the identity of the molecular machinery involved in EphB4 rapid degradation upon IGF-II signal deprivation are unknown. Using targeted proteomics, protein-protein interaction methods, PCR cloning, and 3D modeling approaches, we identified a novel ubiquitin E3 ligase complex recruited by the EphB4 C tail upon autocrine IGF-II signal deprivation. We show this complex to contain a previously unknown N-Terminal isoform of Deltex3 E3-Ub ligase (referred as "DTX3c"), along with UBA1(E1) and UBE2N(E2) ubiquitin ligases and the ATPase/unfoldase Cdc48/p97. Upon autocrine IGF-II neutralization in cultured MSTO211H (a Malignant Mesothelioma cell line that is highly responsive to the EphB4 degradation rescue IGF-II signal), the inter-molecular interactions between these factors were enhanced and their association with the EphB4 C-tail increased consistently with the previously described EphB4 degradation pattern. The ATPase/unfoldase activity of Cdc48/p97 was required for EphB4 recruitment. As compared to the previously known isoforms DTX3a and DTX3b, a 3D modeling analysis of the DTX3c Nt domain showed a unique 3D folding supporting isoform-specific biological function(s). We shed light on the molecular machinery associated with autocrine IGF-II regulation of oncogenic EphB4 kinase expression in a previously characterized IGF-II+/EphB4+ Mesothelioma cell line. The study provides early evidence for DTX3 Ub-E3 ligase involvement beyond the Notch signaling pathway.

Mutual connected IL-6, EGFR and LIN28/Let7-related mechanisms modulate PD-L1 and IGF upregulation in HNSCC using immunotherapy.

The development of techniques and immunotherapies are widely applied in cancer treatment such as checkpoint inhibitors, adoptive cell therapy, and cancer vaccines apart from radiation therapy, surgery, and chemotherapy give enduring anti-tumor effects. Minority people utilize single-agent immunotherapy, and most people adopt multiple-agent immunotherapy. The difficulties are resolved by including the biomarkers to choose the non-responders' and responders' potentials. The possibility of the potential complications and side effects are examined to improve cancer therapy effects. The Head and Neck Squamous Cell Carcinoma (HNSCC) is analyzed with the help of programmed cell death ligand 1 (PD-L1) and Insulin-like growth factor (IGF). But how IGF and PD-L1 upregulation depends on IL-6, EGFR, and LIN28/Let7-related mechanisms are poorly understood. Briefly, IL-6 stimulates gene expressions of IGF-1/2, and IL-6 cross-activates IGF-1R signaling, NF-κB, and STAT3. NF-κB, up-regulating PD-L1 expressions. IL-6/JAK1 primes PD-L1 for STT3-mediated PD-L1 glycosylation, stabilizes PD-L1 and trafficks it to the cell surface. Moreover, ΔNp63 is predominantly overexpressed over TAp63 in HNSCC, elevates circulating IGF-1 levels by repressing IGFBP3, and activates insulin receptor substrate 1 (IRS1).TP63 and SOX2 form a complex with CCAT1 to promote EGFR expression. EGFR activation through EGF binding extends STAT3 activation, and EGFR and its downstream signaling prolong PD-L1 mRNA half-life. PLC-γ1 binding to a cytoplasmic motif of elevated PD-L1 improves EGF-induced activation of inositol 1,4,5-tri-phosphate (IP3), and diacylglycerol (DAG) subsequently elevates RAC1-GTP. RAC1-GTP was convincingly demonstrated to induce the autocrine production and action of IL-6/IL-6R, forming a feedback loop for IGF and PD-L1 upregulation. Furthermore, the LIN28-Let7 axis mediates the NF-κB-IL-6-STAT3 amplification loop, activated LIN28-Let7 axis up-regulates RAS, AKT, IL-6, IGF-1/2, IGF-1R, Myc, and PD-L1, plays pivotal roles in IGF-1R activation and Myc, NF-κB, STAT3 concomitant activation. Therefore, based on a detailed mechanisms review, our article firstly reveals that IL-6, EGFR, and LIN28/Let7-related mechanisms mediate PD-L1 and IGF upregulation in HNSCC, which comprehensively influences immunity, inflammation, metabolism, and metastasis in the tumor microenvironment, and might be fundamental for overcoming therapy resistance.

CBL/CAP Is Essential for Mitochondria Respiration Complex I Assembly and Bioenergetics Efficiency in Muscle Cells.

CBL is rapidly phosphorylated upon insulin receptor activation. Mice whole body CBL depletion improved insulin sensitivity and glucose clearance; however, the precise mechanisms remain unknown. We depleted either CBL or its associated protein SORBS1/CAP independently in myocytes and assessed mitochondrial function and metabolism compared to control cells. CBL- and CAP-depleted cells showed increased mitochondrial mass with greater proton leak. Mitochondrial respiratory complex I activity and assembly into respirasomes were reduced. Proteome profiling revealed alterations in proteins involved in glycolysis and fatty acid degradation. Our findings demonstrate CBL/CAP pathway couples insulin signaling to efficient mitochondrial respiratory function and metabolism in muscle.

Loss of Slc12a2 specifically in pancreatic β-cells drives metabolic syndrome in mice.

The risk of type-2 diabetes and cardiovascular disease is higher in subjects with metabolic syndrome, a cluster of clinical conditions characterized by obesity, impaired glucose metabolism, hyperinsulinemia, hyperlipidemia and hypertension. Diuretics are frequently used to treat hypertension in these patients, however, their use has long been associated with poor metabolic outcomes which cannot be fully explained by their diuretic effects. Here, we show that mice lacking the diuretic-sensitive Na+K+2Cl-cotransporter-1 Nkcc1 (Slc12a2) in insulin-secreting β-cells of the pancreatic islet (Nkcc1βKO) have reduced in vitro insulin responses to glucose. This is associated with islet hypoplasia at the expense of fewer and smaller β-cells. Remarkably, Nkcc1βKO mice excessively gain weight and progressive metabolic syndrome when fed a standard chow diet ad libitum. This is characterized by impaired hepatic insulin receptor activation and altered lipid metabolism. Indeed, overweight Nkcc1βKO but not lean mice had fasting and fed hyperglycemia, hypertriglyceridemia and non-alcoholic steatohepatitis. Notably, fasting hyperinsulinemia was detected earlier than hyperglycemia, insulin resistance, glucose intolerance and increased hepatic de novo gluconeogenesis. Therefore, our data provide evidence supporting the novel hypothesis that primary β-cell defects related to Nkcc1-regulated intracellular Cl-homeostasis and β-cell growth can result in the development of metabolic syndrome shedding light into additional potential mechanisms whereby chronic diuretic use may have adverse effects on metabolic homeostasis in susceptible individuals.

Effects of Rhodiola rosea and its major compounds on insulin resistance in Caenorhabditis elegans

• Hyperglycemic insulin resistance model of C. elegans is conducted. • Major compounds are identified in Rhodiola rosea extact (EAE). • Effects of EAE on insulin resistance in C. elegans is evaluated. • EAE could activate IRS and AMPK pathways. By establishing insulin resistance model of Caenorhabditis elegans , the effects of Rhodiola rosea extract and its compounds on glucose level, insulin signal intensity, antioxidant enzymes and cell apoptosis of C. elegans were investigated in present study. The mRNA expression of DAF-2, DAF-16, AkT-1, SKN-1, AAK-2 and SOD-3 were detected by qRT-PCR. The results revealed that R. rosea extract contain salidroside, kaempferol, chlorogenic acid, kaemphenol-7- O -glucoside, caffeic acid, and quercetin, etc. At the concentration of 200 μg/mL, R. rosea extract treated C. elegans showed increased insulin signal intensity by 52.46% compared with the normal group, while, glucose content decreased by 72.13% and reactive oxygen species (ROS) level decreased by 36.84%. Compared with high glucose model group, the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) increased by 10.9, 14.2 and 27.9, respectively. After R. rosea extract intervention, the expression levels of DAF-2, AkT-1, AAK-2, SKN-1 and other genes significantly increased. These results also indicated that R. rosea extract could activate insulin receptor substrate (IRS) and adenosine 5′-monophosphate (AMP)-activated protein kinase (AMPK) pathways to enhance insulin signaling, promote nutrient metabolism, and achieve the effect for improving insulin resistance of C. elegans .

Development of Dementia in Type 2 Diabetes Patients: Mechanisms of Insulin Resistance and Antidiabetic Drug Development

Dementia is reported to be common in those with type 2 diabetes mellitus. Type 2 diabetes contributes to common molecular mechanisms and an underlying pathology with dementia. Brain cells becoming resistant to insulin leads to elevated blood glucose levels, impaired synaptic plasticity, microglial overactivation, mitochondrial dysfunction, neuronal apoptosis, nutrient deprivation, TAU (Tubulin-Associated Unit) phosphorylation, and cholinergic dysfunction. If insulin has neuroprotective properties, insulin resistance may interfere with those properties. Risk factors have a significant impact on the development of diseases, such as diabetes, obesity, stroke, and other conditions. Analysis of risk factors of importance for the association between diabetes and dementia is important because they may impede clinical management and early diagnosis. We discuss the pathological and physiological mechanisms behind the association between Type 2 diabetes mellitus and dementia, such as insulin resistance, insulin signaling, and sporadic forms of dementia; the relationship between insulin receptor activation and TAU phosphorylation; dementia and mRNA expression and downregulation of related receptors; neural modulation due to insulin secretion and glucose homeostasis; and neuronal apoptosis due to insulin resistance and Type 2 diabetes mellitus. Addressing these factors will offer clinical outcome-based insights into the mechanisms and connection between patients with type 2 diabetes and cognitive impairment. Furthermore, we will explore the role of brain insulin resistance and evidence for anti-diabetic drugs in the prevention of dementia risk in type 2 diabetes.

Влияние ожирения на развитие инсулинорезистентности и окислительного стресса в эксперименте

Purpose. To study influence of abdominal obesity on the formation of peripheral insulin resistance and development of oxidative stress in rats. Materials and methods. Diabetes was developed in 90 white male rats and nonporous females of the Wistar line weighing 80–120 g. In the first group of rats, diabetes mellitus was induced by a single intraperitoneal injection of a 2.5% solution of the antibiotic streptozotocin at a dose of 60 mg/kg of animal weight during a 24–48-hours fasting. In the second group, newborn rats were given a 2.5% solution of streptozotocin once at a dose of 60 mg/kg of body weight, while these rats received a diet enriched with lipids. The composition of the diet included: 8% protein, 30% fat, 62% carbohydrates. The control group received an equivalent amount of saline. The dynamics of carbohydrate and lipid metabolism was assessed. Results. In rats with type 2 diabetes mellitus (DM2) with abdominal obesity and DM2 without obesity structural and functional disorders developed in the membranes of erythrocytes and eye lens fibers, which correlate with indicators of glucose-insulin homeostasis and the level of lipid peroxidation. However, in rats with DM2 and abdominal obesity, the changes in the studied parameters were more significant. Conclusion. Streptozotocin-induced diabetes in obese rats, along with dyslipidemia, leads to the formation of insulin resistance, as evidenced by an increase in the degree of hyperglycemia, a decrease in the activity of insulin receptors and glucose uptake by cells against the background of the development of oxidative stress. The initiating factor of eye lens clouding in cataracts in rats with DM and obesity is apparently a shift in osmolarity in the cells of lens fiber membranes because of the accumulation of insoluble forms of sorbitol and fructose in them. Key words: diabetes mellitus; obesity; streptozotocin; antioxidants; insulin receptors; diabetic cataract; senile cataract

PSUN123 Increased Expression of Insulin Signaling Intermediates After Six-Months Of Medical Care In Women With Obesity And Polycystic Ovary Syndrome

Abstract Background Recently an androgen-induced obesity-independent mechanism of insulin resistance in a mouse model mimicking lean polycystic ovary syndrome (PCOS) was discovered. To date there is little to no data on the mechanism of insulin resistance or molecular insulin signaling in adipose tissue of women with PCOS undergoing Vertical Sleeve Gastrectomy (VSG). Does the mechanism of androgen receptor (AR)-mediated regulation of insulin signaling seen in the lean PCOS mouse model occur in adipose tissue of women with obesity and PCOS? Objective We hypothesize that in women with obesity and PCOS, those with higher levels of free androgen index will display more dysregulation in the insulin signaling pathway in adipose tissue. More specifically, we believe that visceral more than subcutaneous adipose tissue in the women in the study will display more dysregulation; and lastly that VSG will alleviate insulin signaling dysregulation. Briefly, insulin activates insulin receptor (IR) which leads to activation of phospho-AKT which increases phosphorylated forkhead box 01 (p-FOXO1, gluconeogenesis regulation) and transcriptionally regulates acetyl co-A carboxylase (ACC, de novo lipogenesis (DNL) regulation). Methods This is a nested mechanistic study within a randomised controlled clinical trial. We performed molecular analyses (Western blots for protein and qPRC for mRNA expression, respectively, n = 3-4/group) exploring intermediates in the insulin signaling pathway (IR, p-AKT, p-FOXO1, and ACC) in (i) visceral and subcutaneous (SubQ) adipose tissue from women with obesity and PCOS before VSG bariatric surgery (n = 3/group); and (ii) SubQ WAT from women with obesity and PCOS at baseline and six-months after medical intervention with metformin (MTF) and lifestyle modification. Results Subcutaneous (SubQ) white adipose tissue (WAT) displayed similar levels of p-ACC compared to visceral WAT from women pre-VSG. Six months of medical care increased p-ACC in SubQ WAT by 5-fold compared to baseline. IR displayed similar expression in the visceral compared to SubQ WAT in women pre-VSG. IR expression increased by 20-fold at 6-months of medical care compared to baseline in SubQ WAT. p-FOXO1 displayed similar expression in the visceral compared to SubQ WAT. However, p-FOXO1 was decreased at 6-months of medical care compared to baseline. p-AKT levels were similar in visceral compared to SubQ WAT pre-VSG. However, p-AKT levels were significantly increased by 4-fold at 6-months of medical care compared to baseline. Conclusion None of the insulin signaling intermediates measured were different in visceral compared to SubQ in women with obesity and PCOS. Additionally, the findings indicated lowered DNL, lowered regulation of gluconeogenic genes and thus lowered gluconeogenesis, and increased insulin action in SubQ WAT in women with obesity and PCOS after 6-months of medical care. Presentation: Sunday, June 12, 2022 12:30 p.m. - 2:30 p.m.

Aspalathin alleviates skeletal muscle insulin resistance and mitochondrial dysfunction.

Natural compounds may bear promising therapeutic benefits against metabolic diseases such as type 2 diabetes mellitus (T2DM), which are characterized by a state of insulin resistance and mitochondrial dysfunction. Here, we examined the cellular mechanisms by which aspalathin, a dihydrochalcone C-glucoside unique to rooibos, may ameliorate palmitate-induced insulin resistance and mitochondrial dysfunction in cultured C2C12 myotubules. This current study demonstrated that aspalathin remains effective in improving glucose uptake in insulin-resistant skeletal muscle cells, supported by the upregulation of insulin-dependent signaling that involves the activation of insulin receptor (IR) and direct phosphorylation of protein kinase B (AKT). Interestingly, aspalathin also improved mitochondrial respiration and function, which was evident by an increased expression of carnitine palmitoyltransferase 1 (Cpt1), fatty acid transport protein 1 (Fatp1), sirtuin 1 (Sirt1), nuclear respiratory factor 1 (Nrf1), and transcription factor A, mitochondrial (Tfam). Importantly, our results showed that aspalathin treatment was effective in ameliorating the devastating outcomes of insulin resistance and mitochondrial dysfunction that are linked with an undesired pro-inflammatory response, by reducing the levels of well-known pro-inflammatory markers such as interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), and protein kinase C-theta (PKC-theta). Thus, beyond improving glucose uptake and insulin signaling, the current study brings a new perspective in the therapeutic benefits of aspalathin in improving mitochondrial respiration and blocking inflammation to attenuate the detrimental effect of palmitate in skeletal muscle cells.

Functional selectivity of insulin receptor revealed by aptamer-trapped receptor structures

Activation of insulin receptor (IR) initiates a cascade of conformational changes and autophosphorylation events. Herein, we determined three structures of IR trapped by aptamers using cryo-electron microscopy. The A62 agonist aptamer selectively activates metabolic signaling. In the absence of insulin, the two A62 aptamer agonists of IR adopt an insulin-accessible arrowhead conformation by mimicking site-1/site-2’ insulin coordination. Insulin binding at one site triggers conformational changes in one protomer, but this movement is blocked in the other protomer by A62 at the opposite site. A62 binding captures two unique conformations of IR with a similar stalk arrangement, which underlie Tyr1150 mono-phosphorylation (m-pY1150) and selective activation for metabolic signaling. The A43 aptamer, a positive allosteric modulator, binds at the opposite side of the insulin-binding module, and stabilizes the single insulin-bound IR structure that brings two FnIII-3 regions into closer proximity for full activation. Our results suggest that spatial proximity of the two FnIII-3 ends is important for m-pY1150, but multi-phosphorylation of IR requires additional conformational rearrangement of intracellular domains mediated by coordination between extracellular and transmembrane domains.

Cerebrovascular insulin receptors are defective in Alzheimer's disease.

Central response to insulin is suspected to be defective in Alzheimer's disease. As most insulin is secreted in the bloodstream by the pancreas, its capacity to regulate brain functions must, at least partly, be mediated through the cerebral vasculature. However, how insulin interacts with the blood-brain barrier and whether alterations of this interaction could contribute to Alzheimer's disease pathophysiology both remain poorly defined. Here, we show that human and murine cerebral insulin receptors (INSRs), particularly the long isoform INSRα-B, are concentrated in microvessels rather than in the parenchyma. Vascular concentrations of INSRα-B were lower in the parietal cortex of subjects diagnosed with Alzheimer's disease, positively correlating with cognitive scores, leading to a shift towards a higher INSRα-A/B ratio, consistent with cerebrovascular insulin resistance in the Alzheimer's disease brain. Vascular INSRα was inversely correlated with amyloid-β plaques and β-site APP cleaving enzyme 1, but positively correlated with insulin-degrading enzyme, neprilysin and P-glycoprotein. Using brain cerebral intracarotid perfusion, we found that the transport rate of insulin across the blood-brain barrier remained very low (<0.03 µl/g·s) and was not inhibited by an insulin receptor antagonist. However, intracarotid perfusion of insulin induced the phosphorylation of INSRβ that was restricted to microvessels. Such an activation of vascular insulin receptor was blunted in 3xTg-AD mice, suggesting that Alzheimer's disease neuropathology induces insulin resistance at the level of the blood-brain barrier. Overall, the present data in post-mortem Alzheimer's disease brains and an animal model of Alzheimer's disease indicate that defects in the insulin receptor localized at the blood-brain barrier strongly contribute to brain insulin resistance in Alzheimer's disease, in association with β-amyloid pathology.

E2F1 transcription factor mediates a link between fat and islets to promote β cell proliferation in response to acute insulin resistance.

SUMMARYPrevention or amelioration of declining β cell mass is a potential strategy to cure diabetes. Here, we report the pathways utilized by β cells to robustly replicate in response to acute insulin resistance induced by S961, a pharmacological insulin receptor antagonist. Interestingly, pathways that include CENP-A and the transcription factor E2F1 that are independent of insulin signaling and its substrates appeared to mediate S961-induced β cell multiplication. Consistently, pharmacological inhibition of E2F1 blocks β-cell proliferation in S961-injected mice. Serum from S961-treated mice recapitulates replication of β cells in mouse and human islets in an E2F1-dependent manner. Co-culture of islets with adipocytes isolated from S961-treated mice enables β cells to duplicate, while E2F1 inhibition limits their growth even in the presence of adipocytes. These data suggest insulin resistance-induced proliferative signals from adipocytes activate E2F1, a potential therapeutic target, to promote β cell compensation.

Activation of the human insulin receptor by non-insulin-related peptides

The human insulin receptor signalling system plays a critical role in glucose homeostasis. Insulin binding brings about extensive conformational change in the receptor extracellular region that in turn effects trans-activation of the intracellular tyrosine kinase domains and downstream signalling. Of particular therapeutic interest is whether insulin receptor signalling can be replicated by molecules other than insulin. Here, we present single-particle cryoEM structures that show how a 33-mer polypeptide unrelated to insulin can cross-link two sites on the receptor surface and direct the receptor into a signalling-active conformation. The 33-mer polypeptide engages the receptor by two helical binding motifs that are each potentially mimicable by small molecules. The resultant conformation of the receptor is distinct from—but related to—those in extant three-dimensional structures of the insulin-complexed receptor. Our findings thus illuminate unexplored pathways for controlling the signalling of the insulin receptor as well as opportunities for development of insulin mimetics.

Differential regulation of insulin signalling by monomeric and oligomeric amyloid beta-peptide.

Alzheimer’s disease and Type 2 diabetes are pathological processes associated to ageing. Moreover, there are evidences supporting a mechanistic link between Alzheimer’s disease and insulin resistance (one of the first hallmarks of Type 2 diabetes). Regarding Alzheimer’s disease, amyloid β-peptide aggregation into β-sheets is the main hallmark of Alzheimer’s disease. At monomeric state, amyloid β-peptide is not toxic but its function in brain, if any, is unknown. Here we show, by in silico study, that monomeric amyloid β-peptide 1-40 shares the tertiary structure with insulin and is thereby able to bind and activate insulin receptor. We validated this prediction experimentally by treating human neuroblastoma cells with increasing concentrations of monomeric amyloid β-peptide 1-40. Our results confirm that monomeric amyloid β-peptide 1-40 activates insulin receptor autophosphorylation, triggering downstream enzyme phosphorylations and the glucose Transporter 4 translocation to the membrane. On the other hand, neuronal insulin resistance is known to be associated to Alzheimer’s disease since early stages. We thus modelled the docking of oligomeric amyloid β-peptide 1-40 to insulin receptor. We found that oligomeric amyloid β-peptide 1-40 blocks insulin receptor, impairing its activation. It was confirmed in vitro by observing the lack of insulin receptor autophosphorylation, and also the impairment of insulin-induced intracellular enzyme activations and the glucose Transporter 4 translocation to the membrane. By biological system analysis, we have carried out a mathematical model recapitulating the process that turns amyloid β-peptide binding to insulin receptor from the physiological to the pathophysiological regime. Our results suggest that monomeric amyloid β-peptide 1-40 contributes to mimic insulin effects in the brain, which could be good when neurons have an extra requirement of energy beside the well-known protective effects on insulin intracellular signalling, while its accumulation and subsequent oligomerization blocks the insulin receptor producing insulin resistance and compromising neuronal metabolism and protective pathways.

Insulin increases placental triglyceride as a potential mechanism for fetal adiposity in maternal obesity

ObjectiveMaternal obesity increases the incidence of excess adiposity in newborns, resulting in lifelong diabetes risk. Elevated intrauterine fetal adiposity has been attributed to maternal hyperglycemia; however, this hypothesis does not account for the increased adiposity seen in newborns of mothers with obesity who have euglycemia. We aimed to explore the placental response to maternal hyperinsulinemia and the effect of insulin-like growth factor 2 (IGF-2) in promoting fetal adiposity by increasing storage and availability of nutrients to the fetus. MethodsWe used placental villous explants and isolated trophoblasts from normal weight and obese women to assess the effect of insulin and IGF-2 on triglyceride content and insulin receptor signaling. Stable isotope tracer methods were used ex vivo to determine effect of hormone treatment on de novo lipogenesis (DNL), fatty acid uptake, fatty acid oxidation, and esterification in the placenta. ResultsHere we show that placentae from euglycemic women with normal weight and obesity both have abundant insulin receptor. Placental depth and triglyceride were greater in women with obesity compared with normal weight women. In syncytialized placental trophoblasts and villous explants, insulin and IGF-2 activate insulin receptor, induce expression of lipogenic transcription factor SREBP-1 (sterol regulatory element-binding protein 1), and stimulate triglyceride accumulation. We demonstrate elevated triglyceride is attributable to increased esterification of fatty acids, without contribution from DNL and without an acceleration of fatty acid uptake. ConclusionsOur work reveals that obesity-driven aberrations in maternal metabolism, such as hyperinsulinemia, alter placental metabolism in euglycemic conditions, and may explain the higher prevalence of excess adiposity in the newborns of obese women.

Making Liquid Sugar Preparations from Lombok-Traditional Sugarcane Juice and Its Potential as Anti-Diabetic Sugar

The prevalence of Patients with Diabetes Mellitus remains on the rise worldwide. Diabetes is a multifactorial disorder that emboils genetic and environmental factors such as an unhealthy diet by consuming white sugar crystals. White crystal sugar consists of less chromium content that induces glucose in the blood to the cells, which can trigger hyperglycemia or high blood sugar levels. Chromium works by increasing the activity of insulin receptors to enhance glucose translocation to the cell. Previous studies showed sugarcane Nira had a high level of chromium compared with those processed into white crystalline sugar. However, it could not be used as a sweetener substitute since its storage age was only 2 to 3 days. This study identified that the treatment process of Lombok's Nira sugarcane into local liquefied sugar should be addressed to maintain chromium levels compared to the processing of Nira into the white crystalline sugar requiring optimum sucrose and omitting several minerals. Hence, sugarcane Nira as a refined sugar product can substitute for white sugar consumption in diabetes patients.

Minimizing Mitogenic Potency of Insulin Analogues Through Modification of a Disulfide Bond.

The mechanisms by which insulin activates the insulin receptor to promote metabolic processes and cellular growth are still not clear. Significant advances have been gained from recent structural studies in understanding how insulin binds to its receptor. However, the way in which specific interactions lead to either metabolic or mitogenic signalling remains unknown. Currently there are only a few examples of insulin receptor agonists that have biased signalling properties. Here we use novel insulin analogues that differ only in the chemical composition at the A6–A11 bond, as it has been changed to a rigid, non-reducible C=C linkage (dicarba bond), to reveal mechanisms underlying signaling bias. We show that introduction of an A6-A11 cis-dicarba bond into either native insulin or the basal/long acting insulin glargine results in biased signalling analogues with low mitogenic potency. This can be attributed to reduced insulin receptor activation that prevents effective receptor internalization and mitogenic signalling. Insight gained into the receptor interactions affected by insertion of an A6-A11 cis-dicarba bond will ultimately assist in the development of new insulin analogues for the treatment of diabetes that confer low mitogenic activity and therefore pose minimal risk of promoting cancer with long term use.

Caveolae Microdomains Mediate STAT5 Signaling Induced by Insulin in MCF-7 Breast Cancer Cells.

Caveolae are small plasma membrane invaginations constituted for membrane proteins namely caveolins and cytosolic proteins termed cavins, which can occupy up to 50% of the surface of mammalian cells. The caveolae have been involved with a variety of cellular processes including regulation of cellular signaling. Insulin is a hormone that mediates a variety of physiological processes through activation of insulin receptor (IR), which is a tyrosine kinase receptor expressed in all mammalian tissues. Insulin induces activation of signal transducers and activators of transcription (STAT) family members including STAT5. In this study, we demonstrate, for the first time, that insulin induces phosphorylation of STAT5 at tyrosine-694 (STAT5-Tyr(P)694), STAT5 nuclear accumulation and an increase in STAT5-DNA complex formation in MCF-7 breast cancer cells. Insulin also induces nuclear accumulation of STAT5-Tyr(P)694, caveolin-1, and IR in MCF-7 cells. STAT5 nuclear accumulation and the increase of STAT5-DNA complex formation require the integrity of caveolae and microtubule network. Moreover, insulin induces an increase and nuclear accumulation of STAT5-Tyr(P)694 in MDA-MB-231 breast cancer cells. In conclusion, results demonstrate that caveolae and microtubule network play an important role in STAT5-Tyr(P)694, STAT5 nuclear accumulation and STAT5-DNA complex formation induced by insulin in breast cancer cells.

Intra-NAc insulin reduces the motivation for food and food intake without altering cue-triggered food-seeking

Insulin receptors are expressed throughout the adult brain, and insulin from the periphery reaches the central nervous system. In humans and rodents, actions of insulin in the brain decrease food intake. Furthermore, insulin receptor activation alters dopamine and glutamate transmission within mesolimbic regions that influence food-seeking and feeding including the nucleus accumbens (NAc). Here we determined how intra-NAc insulin affects conditioned approach (a measure of cue-triggered food-seeking), free food intake, and the motivation to obtain food in hungry rats using Pavlovian and instrumental approaches. Intra-NAc insulin did not affect conditioned approach but did reduce home cage chow intake immediately following conditioned approach testing. Consistent with reduced chow intake, intra-NAc insulin also reduced the motivation to work for flavored food pellets (assessed by a progressive ratio procedure). This effect was partially reversed by insulin receptor blockade and was not driven by insulin-induced sickness or malaise. Taken together, these data show that insulin within the NAc does not alter behavioral responses to a food cue, but instead reduces the motivation to work for and consume food in hungry animals. These data are discussed in light of insulin's role in the regulation of feeding, and its dysregulation by obesity.