Insulin Binding Research Articles

1320-P: Insulin Receptors and Their Signal Transduction Pathways in Choroid Plexus

The insulin receptor (IR) is a tyrosine kinase receptor present in cell membranes. It is highly expressed in hepatocytes (where it promotes glycogenesis and inhibits gluconeogenesis) , and skeletal muscle and fat (where it facilitates glucose uptake by translocation of the glucose transporter GLUT4) . IR expression is also found throughout the brain, both in neurons and all glial cell types, and for decades impaired insulin signaling has been implicated in the onset and development of Alzheimer’s disease. IR expression pattern in the brain parenchyma is heterogeneous, with highest levels of expression found in olfactory bulb, hypothalamus, and hippocampus. However, the choroid plexus (CP) , a structure present in all 4 ventricles of the brain and responsible for the secretion of cerebrospinal fluid, has the highest affinity for insulin binding compared to any other brain region. This work aims to characterize the insulin signaling in the epithelial cells of the choroid plexus (EChP) and its possible implications in diseases of the brain. Relative expression levels of specific splicing variants of the Insr gene in dissected CP of adult mice were evaluated by qPCR and revealed predominant expression of the variant coding for isoform IR-A when compared to isoform IR-B. In vitro stimulation of EChP with insulin elicits a classical tyrosine kinase cascade, with activation of both the mitogenic (ERK) and metabolic (Akt) arms of the insulin signaling pathway, in a time-and concentration-dependent manner. Using pharmacological (S961, an insulin receptor antagonist) and genetic (insulin receptor shRNA, and Insr cre-loxP) approaches, we show that downregulation of IR in these cells not only impairs insulin signaling, but it also decreases IGF2’s ability to activate ERK and Akt, even when IGF1R levels are unaltered. Finally, using a viral vector coding for an IR shRNA, we show that downregulation of Insr expression in CP of adult mice has a profound effect on its structure, as demonstrated by MRI and histological technique. Disclosure C.Mazucanti: None. V.L.Kennedy: None. A.Cho: None. Q.Liu: None. J.F.O’connell: None. S.Camandola: None. J.M.Egan: None.

An SPR-based method for Hill coefficient measurements: the case of insulin-degrading enzyme

Insulin-degrading enzyme (IDE) is a highly conserved zinc metallopeptidase and is capable to catalytically cleave several substrates besides insulin, playing a pivotal role in several different biochemical pathways. Although its mechanism of action has been widely investigated, many conundrums still remain, hindering the possibility to rationally design specific modulators which could have important therapeutical applications in several diseases such as diabetes and Alzheimer's disease. In this scenario, we have developed a novel surface plasmon resonance (SPR) method which allows for directly measuring the enzyme cooperativity for the binding of insulin in the presence of different IDE activity modulators: carnosine, ATP, and EDTA. Results indicate that both positive and negative modulations of the IDE activity can be correlated to an increase and a decrease of the measured Hill coefficient, respectively, giving a new insight into the IDE activity mechanism. The use of the IDE R767A mutant for which oligomerization is hindered confirmed that the positive allosteric modulation of IDE by carnosine is due to a change in the enzyme oligomeric state occurring also for the enzyme immobilized on the gold SPR chip.

Interactions of TUG‐UBL1 and Insulin‐Implications for Glucose Uptake

TUG‐UBL1 (Tether Containing UBX Domain for GLUT‐4 Ubiquitin Like 1) is a glucose transporter 4 (GLUT4) regulating protein which facilitates the release of GLUT4 transport vesicles when insulin stimulation is present. To understand the role of TUG‐UBL1 in glucose metabolism, especially for type two diabetic patients for whom increases in insulin concentration do not correlate with increased glucose uptake, we explored the research question of whether TUG‐UBL1 binds to insulin. Initially, models of full‐length TUG‐UBL1 (amino acids 1‐473), the GLUT4 binding domain of TUG‐UBL1 (amino acids 86‐337), and the insulin heterodimer were built using PHYRE2. All of these structures were further refined with DeepRefiner, and then validated using MolProbity. Including all‐atom contacts, full‐length TUG‐UBL1 scored in the 99th percentile, the GLUT4 binding domain of TUG‐UBL1 scored in the 97th percentile, and the insulin heterodimer scored in the 97th percentile. Insulin binding to both the full‐length TUG‐UBL1 domain and the GLUT4 binding domain were then explored with HawkDock. In both cases, favorable binding energies were observed with predictions of GLUT4 binding region of TUG‐UBL1 binding to insulin at ‐43.31 kcal/mol and full‐length TUG‐UBL1 binding to insulin at ‐32.31 kcal/mol, with predictions favoring insulin binding to the GLUT4 binding domain. To support these computational findings, the GLUT4 binding domain of TUG‐UBL1 is being over‐expressed and purified for use in intrinsic tryptophan fluorescence binding studies with insulin. The finding of TUG‐UBL1 binding to insulin are interesting in light of the potential implications for the care type two diabetic patients as binding characterization could lead to further mechanistic understanding of insulin resistance and possible therapeutic development.

Association of animal and plant protein intakes with biomarkers of insulin and insulin-like growth factor axis

Insulin and insulin-like growth factor (IGF)-1 signaling is a proposed mechanism linking dietary protein and major chronic diseases. However, it is unclear whether animal and plant proteins are associated with biomarkers of insulin and IGF axis. We analyzed a total of 14,709 participants from Nurses' Health Study and Health Professionals Follow-up Study who had provided a blood sample. Detailed dietary information was assessed using validated food frequency questionnaires. We assessed C-peptide, insulin, IGF-1, and IGF binding proteins (BP). Multivariable-adjusted linear regressions were used to examine associations of animal and plant protein intake with biomarkers after adjusting for confounders. The medians (5th-95th percentiles) of animal and plant protein intake (% of total energy) were 13% (8-19%) and 5% (4-7%), respectively. Compared to participants in the lowest quintile, those in the highest quintile of animal protein had 4.8% (95% CI: 1.9, 7.9; P-trend<0.001) higher concentration of IGF-1 and -7.2% (95% CI:-14.8, 1.1; P for trend=0.03) and-11.8% (95% CI:-20.6,-1.9; P-trend<0.001) lower concentration of IGFBP-1 and IGFBP-2, respectively, after adjustment for major lifestyle factors and diet quality. In contrast, no association was observed between animal protein intake and C-peptide, insulin and IGFBP-3. The associations were restricted to participants with at least one unhealthy lifestyle risk factor (i.e., overweight/obese, physical inactivity, smoking, and heavy alcohol intake). Plant protein tended to be strongly associated with numerous biomarkers in age-adjusted analyses but these became largely attenuated or non-significant in multivariable adjustment. Plant protein intake remained positively associated with IGF-1 (P-trend=0.002) and possibly IGFBP-1 (P-trend=0.02) after multivariable adjustment. Substitution of plant protein with animal protein sources was associated with lower IGFBP-1. In additional analysis, IGF-1 and IGFBPs were estimated to mediate approximately 5-20% of the association between animal protein and type 2 diabetes. Higher animal protein intake was associated with higher IGF-1 and lower IGFBP-1 and IGFBP-2, whereas higher plant protein intake was associated with higher IGF-1 and IGFBP-1.

Antidiabetic Action of Mcy Protein: Studies on Gene Expression and Competitive Binding to Insulin Receptors.

Mcy protein, isolated from the fruits of Momordica cymbalaria, was shown to have antihyperglycemic, antihyperlipidemic activities along with renal as well as hepatoprotective activities in streptozotocin-induced diabetic rats. Mcy protein was shown to have insulin-like structure and/or function and/or insulin secretagogue activity. Hence, the present study was conducted to elucidate the molecular mechanism whereby Mcy protein elicits its therapeutic role and also to know whether the Mcy protein has any structural and functional similarity with insulin. Results of our experiments revealed that the Mcy protein is insulin-like protein. Furthermore, we assessed the effect of treatment with Mcy protein on the glucose transport (levels of glucose transporter, GLUT-2) and on the levels of key regulators of glucose and lipid metabolisms like hepatic glucokinase (GK) and sterol regulatory element-binding protein-1c (SREBP-1c). Our findings demonstrated that Mcy protein elevated the expressions of GK, SREBP-1c, and GLUT-2 that were decreased in diabetic animals. Insulin-receptor binding studies using rat erythrocytes demonstrated that mean specific binding of insulin with insulin receptors was significantly increased in Mcy-treated diabetic rats when compared to diabetic control rats. Scatchard analyses of insulin binding studies yielded curvilinear plots, and the number of receptor sites per cell was found to be 180 ± 21.1 in Mcy-treated diabetic animals and found to be significantly superior to those of diabetic control animals. Kinetic analyses also revealed an increase in the average receptor affinity of erythrocytes of Mcy-treated rats compared to diabetic control rats suggesting acute alteration in the number and affinity of insulin receptors on the membranes of erythrocytes.

A Modified Polymeric Nano-formulation to Control Binding and Release of Insulin

The study involves evaluation of the binding affinity of insulin to modified-hydrophobic nanoparticles to control its release. A specific peptide called hormone binding peptide (Hbpep) was designed to encompass most of the hydrophobic residues aiming to provide controlled hydrophobic interactions. Polycaprolactone (PCL) polymer was used here to prepare nanoparticles as carriers for insulin molecules. Hbpep ligand was attached covalently to these nanoparticles after embedding enormous carboxyl groups on the nanoparticles surface using conventional bioconjugation chemistry. The thermodynamic of insulin binding with the ligands was studied using isothermal titration calorimetry (ITC). The study further evaluated the adsorption behavior of insulin with the conjugates under various microenvironmental conditions using equilibrium microdialysis, fluorescence spectroscopy and dynamic light scattering. The adsorption process was found to be affected by changing the pH of the medium. The modified nanoparticles showed a controlled insulin binding especially at neutral pH. The energetics of binding was found to be of high affinity type demonstrating a potential for developing a controlled delivery system for insulin.

An aptamer agonist of the insulin receptor acts as a positive or negative allosteric modulator, depending on its concentration.

Aptamers are widely used as binders that interact with targets with high affinity or as inhibitors of the function of target molecules. However, they have also been used to modulate target protein function, which they achieve by activating the target or stabilizing its conformation. Here, we report a unique aptamer modulator of the insulin receptor (IR), IR-A62. Alone, IR-A62 acts as a biased agonist that preferentially induces Y1150 monophosphorylation of IR. However, when administered alongside insulin, IR-A62 shows variable binding cooperativity depending on the ligand concentration. At low concentrations, IR-A62 acts as a positive allosteric modulator (PAM) agonist that enhances insulin binding, but at high concentrations, it acts as a negative allosteric modulator (NAM) agonist that competes with insulin for IR. Moreover, the concentration of insulin affects the binding of IR-A62 to IR. Finally, the subcutaneous administration of IR-A62 to diabetic mice reduces blood glucose levels with a longer-lasting effect than insulin administration. These findings imply that aptamers can elicit various responses from receptors beyond those of a simple agonist or inhibitor. We expect further studies of IR-A62 to help reveal the mechanism of IR activation and greatly expand the range of therapeutic applications of aptamers.

Synergistic activation of the insulin receptor via two distinct sites.

Insulin receptor (IR) signaling controls multiple facets of animal physiology. Maximally four insulins bind to IR at two distinct sites, termed site-1 and site-2. However, the precise functional roles of each binding event during IR activation remain unresolved. Here, we showed that IR incompletely saturated with insulin predominantly forms an asymmetric conformation and exhibits partial activation. IR with one insulin bound adopts a Γ-shaped conformation. IR with two insulins bound assumes a Ƭ-shaped conformation. One insulin binds at site-1 and another simultaneously contacts both site-1 and site-2 in the Ƭ-shaped IR dimer. We further show that concurrent binding of four insulins to sites-1 and -2 prevents the formation of asymmetric IR and promotes the T-shaped symmetric, fully active state. Collectively, our results demonstrate how the synergistic binding of multiple insulins promotes optimal IR activation.

High-fat diet induced alterations in plasma membrane cholesterol content impairs insulin receptor binding and signalling in mouse liver but is ameliorated by atorvastatin

A high-fat diet (HFD) impairs insulin binding and signalling and may contribute to the development of insulin resistance. In addition, in vitro studies have shown that alterations in plasma membrane cholesterol influence ligand binding and downstream signalling for several receptor-tyrosine kinases (RTKs), including the insulin receptor. Using an ex vivo approach, we explored the effects of a HFD on insulin binding and signalling in mouse liver and relate these to observed changes in plasma membrane cholesterol. Mice fed a HFD demonstrated decreased insulin signalling compared to mice fed a normal chow diet (ND), indicated by a 3-fold decrease in insulin binding (P < 0.001) and a similar decrease in insulin receptor phosphorylation (~2.5-fold; P < 0.0001). Interestingly, we also observed a marked decrease in the cholesterol content of liver plasma membranes in the HFD fed mice (P < 0.0001). These effects of the HFD were found to be ameliorated by atorvastatin treatment (P < 0.0001). However, in ND mice, atorvastatin had no influence on membrane cholesterol content or insulin binding and signalling. The influence of membrane cholesterol on insulin binding and signalling was also corroborated in HepG2 cells. To the best of our knowledge, this is the first demonstration of the effects of a HFD and atorvastatin treatment on changes in plasma membrane cholesterol content and the consequent effects on insulin binding and signalling. Collectively, these findings suggest that changes in membrane cholesterol content could be an important underlying reason for the long-known effects of a HFD on the development of insulin resistance.

Insulin blood-brain barrier transport and interactions are greater following exercise in mice.

Exercise has multiple beneficial effects including improving peripheral insulin sensitivity, improving central function such as memory, and restoring a dysregulated blood-brain barrier (BBB). Central nervous system (CNS) insulin resistance is a common feature of cognitive impairment, including Alzheimer's disease. Delivery of insulin to the brain can improve memory. Endogenous insulin must cross the BBB to directly act within the CNS and this transport system can be affected by various physiological states and serum factors. Therefore, the current study sought to investigate whether exercise could enhance insulin BBB transport as a mechanism for the underlying benefits of exercise on cognition. We investigated radioactive insulin BBB pharmacokinetics following an acute bout of exercise in young, male and female CD-1 mice. In addition, we investigated changes in serum levels of substrates that are known to affect insulin BBB transport. Finally, we measured the basal level of a downstream protein involved in insulin receptor signaling in various brain regions as well as muscle. We found insulin BBB transport in males was greater following exercise, and in males and females to both enhance the level of insulin vascular binding and alter CNS insulin receptor signaling, independent of changes in serum factors known to alter insulin BBB transport.NEW & NOTEWORTHY Central nervous system (CNS) insulin and exercise are beneficial for cognition. CNS insulin resistance is present in Alzheimer's disease. CNS insulin levels are regulated by transport across the blood-brain barrier (BBB). We show that exercise can enhance insulin BBB transport and binding of insulin to the brain's vasculature in mice. There were no changes in serum factors known to alter insulin BBB pharmacokinetics. We conclude exercise could impact cognition through regulation of insulin BBB transport.

Structural Investigations of Full-Length Insulin Receptor Dynamics and Signalling

Insulin regulates glucose homeostasis via binding and activation of the insulin receptor dimer at two distinct pairs of binding sites 1 and 2. Here, we present cryo-EM studies of full-length human insulin receptor (hIR) in an active state obtained at non-saturating, physiologically relevant insulin conditions. Insulin binds asymmetrically to the receptor under these conditions, occupying up to three of the four possible binding sites. Deletion analysis of the receptor together with site specific peptides and insulin analogs used in binding studies show that both sites 1 and 2 are required for high insulin affinity. We identify a homotypic interaction of the fibronectin type III domain (FnIII-3) of IR resulting in tight interaction of membrane proximal domains of the active, asymmetric receptor dimer. Our results show how insulin binding at two distinct types of sites disrupts the autoinhibited apo-IR dimer and stabilizes the active dimer. We propose an insulin binding and activation mechanism, which is sequential, exhibits negative cooperativity, and is based on asymmetry at physiological insulin concentrations with one to three insulin molecules activating IR.

Study of serum magnesium levels and its correlation with glycemic status in type II diabetes patients

Magnesium is most important and vital element of body. It needs to be supplemented adequately. It plays a vital role in insulin secretion, insulin binding and homeostasis. When Serum Magnesium is adequate, the glycemic control is better and HbA1c values will fall, thus proving that serum magnesium plays a major role in glycemic control. It is now established that diabetes can by itself induce hypomagnesemia and hypomagnesemia can in turn induce onset or worsen diabetes mellitus.: A cross-sectional study was conducted in 48 diagnosed cases of type II diabetes mellitus. This study was planned to study if any correlation exists between the level of Serum Magnesium and HbA1C in diagnosed Type II diabetics.: The correlation between the two parameters was not found to be statistically significant.: Owing to COVID-19 restrictions history regarding the duration of disease, the dietary history of the participants could not be obtained: Serum magnesium does not bear a constant relationship with the diabetic control according to the findings of the current study and detailed studies including multi-parametric analysis along with duration of diabetes is required.

Glycogen synthase kinase 3β inhibition and insulin-receptor binding enhancement of compounds isolated from wild leafy vegetable Acalypha alnifolia

BackgroundThe traditional information of Acalypha alnifolia is captivate towards its therapeutic efficacy against diabetes along with the scientific confirmation were leads to an instinct to extract phytocompounds and to evaluate antidiabetic-molecular mechanism. MethodsColumn chromatographic compound isolation were adopted to extract the pure compounds from bioactive leaf extract and interactions of the compounds with diabetic metabolic proteins were accomplished through in-silico docking studies. By the direction, the anti-diabetic property was valued by the expressions of Glycogen Synthase Kinase 3β(GSK 3β) and Insulin on insulin-Receptor on Hep G2 cells. ResultsThe flavone compound - 5,7-dihydroxy-2-(4-hydroxyphenyl)-3-methoxy-4H-chromen-4-one and pyridine alkaloid - (2,6-dihydroxypyridin-4-yl)(3,5-dihydroxytetrahydro-2H-pyran-4-yl) acetic acid were isolated. Both the compounds reduced the GSK 3β expression and the flavone contributed also to enhance insulin binding ability expediently. ConclusionsThe results concluded that the compounds from A. alnifolia having potent to maintain glucose homeostasis in liver insulin-metabolism. By exploring pharmaceutical importance of the isolated compounds, this study will be an important breakthrough in developing new drug for Diabetes.

Insulin resistance; mechanisms and related health hazards: a review article

Insulin resistance (IR) is related to metabolic syndrome, which is manifested by biochemical and physical disturbances that could be related to increased weight, atherogenic dyslipidemia, pro-inflammatory conditions, pro-thrombotic conditions, and hypertension. The process is complex, and the operating mechanisms are not well understood. Thus, this article aimed to determine the mechanisms of IR and its related serious health problems. It was found that subjects with impaired glucose-6-phosphate action and muscle glycogen production carry a 40% chance of developing type-2 diabetes mellitus. Further, the pro-inflammatory mediator interleukin-6 (IL-6) increases the insulin-induced glucose take-up by myocytes and decreases IR or increases insulin sensitivity in the entire body. Many reports documented an increase in IL-6 in the skeletal muscles of obese people. Cultured myocytes from obese diabetic individuals produce more monocyte chemo-attractant protein 1 and tumor necrotic factor alpha (TNF-α). TNF-α could affect mitochondrial function in myocytes and stimulate IR. The binding of insulin to the insulin receptors on endothelial cells activates the mitogen-activated protein kinase and the phosphatidylinositol 3-kinase pathways. In contrast to the first pathway, the second pathway is pro-atherogenic and is activated by IR. It was concluded that IR is associated with increased release of pro-inflammatory mediators, oxidative stress molecules, and modulation of insulin receptors in fatty tissues and endothelial cells.

Wearables, Respiratory Rehabilitation, Acute Myocardial Infarction and Frailty, and High-Intensity Respiratory Muscle Training

Wearables, Respiratory Rehabilitation, Acute Myocardial Infarction and Frailty, and High-Intensity Respiratory Muscle Training

Alterations of neurovascular insulin receptor in Alzheimer's disease.

The hypothesis that central insulin resistance plays a role in Alzheimer's disease (AD) is gaining interest. However, insulin is secreted by the pancreas and must first interact with the blood-brain barrier (BBB) before having an impact on brain function. In addition, β-amyloid peptides (Aβ) can compete with insulin binding to its insulin receptor (INSR). The aim of the present study was thus to investigate the association between INSR and Aβ in brain microvessels isolated from frozen human brain samples. We used microvessel-enriched brain samples from the Religious Order Study from individuals classified as either Controls, mild cognitive impaired (MCI), or AD. First, the INSR was found enriched in human brain microvessels, compared to parenchymal samples. We next observed lower levels of the INSR precursor (proINSR) and INSRα-B in the parietal cortex of subject diagnosed with AD, while INSRβ remained unchanged between groups. A shift toward a higher INSRα-A: INSRα-B ratio was present in AD brain, consistent with insulin resistance. Western blot analyses showed that INSRα-B levels positively correlated with cognitive scores. Moreover, proINSR, and mature INSR (INSRβ and INSRα-B) were inversely correlated with Aβ plaques in brain cortex and β-site APP cleaving enzyme 1 (BACE1) of microvessels. In addition, positive associations between Insulin-degrading enzyme (IDE) and neprilysin or ATP Binding Cassette Subfamily B Member 1 (ABCB1) were established. Overall, our data support the hypothesis of brain insulin resistance in AD, implicating INSR localized in microvessels. Alterations of vascular INSR were associated with ante mortem cognitive impairment and with Aβ or proteins involved in its production or clearance.

Collegiate male athletes exhibit conditions of the Male Athlete Triad.

The primary purpose of this study was to determine prevalence of the Male Athlete Triad (MAT) conditions: low energy availability (EA), low bone mineral density (BMD), and low testosterone in male collegiate athletes from different sports. Participants included 44 collegiate male athletes (age, 20.4 ± 0.2 years; body mass index, 25.3 ± 1.3kg/m2) from 7 sports (cross country, soccer, basketball, wrestling, track, golf, and baseball). Resting metabolic rate, 3-day food intake, 7-day exercise energy expenditure, body composition, and reproductive and metabolic hormones were assessed. Of the total participants, 15% had low EA, 0% had low BMD, 28% had low total testosterone (TT), and 80% had low calculated free testosterone (cFT). There were no significant correlations between EA, BMD, TT, and cFT. Insulin and sex hormone binding globulin (SHBG) were below and on the upper end of the reference range for healthy male adults, respectively. Insulin was negatively correlated with total (r = -0.330, p= 0.043) and lumbar spine BMD z-scores (r = -0.413, p= 0.010). Low TT and low cFT were the most prevalent MAT conditions among all athletes. Further research should investigate the relationship between insulin and SHBG and the role of these hormones in the MAT. Novelty: Assessment of energy availability alone is not sufficient to identify physiological disturbances in collegiate male athletes. Low total and/or free testosterone may be present in some collegiate male athletes, regardless of BMD status. Low insulin and high SHBG concentration may portray the presence of conditions of the MAT in male collegiate athletes.

ROLE OF GSK-3 IN Wnt/β-CATENIN SIGNALING PATHWAY IN OBESITY

The complexity of the adipogenesis mechanism results from the impact of multiple cues, among which an important place is held by the components of the Wnt signaling pathway. The search for potential markers of the development of diseases related to obesity aroused an interest in the study of GSK-3 (glycogen synthase kinase), β-catenin. GSK-3β is an intracellular serine / threonine kinase found in the cytoplasm, nucleus, mitochondria, synthesized in all body tissues and involved in regulating metabolic processes, cell proliferation, apoptosis etc. The first of the discovered functions of GSK-3β was the regulation of glycogen synthesis. Active GSK-3β phosphorylates and thereby inhibits glycogen synthase. As a result of the insulin binding to the cell receptor via inositol-3-phosphate, protein kinase B (Akt1) is activated, which, in turn, phosphorylates and inhibits GSK-3β. In addition, GSK-3β is involved in the regulating glucose metabolism. The most important function of GSK-3β is the inhibition of the β-catenin protein. In a resting cell, GSK-3β in complex with the APC and Axin proteins binds and phosphorylates the β-catenin transcription factor, which leads to its ubiquitination and degradation. When Wnt proteins act on the cell, the Dvl protein is activated, which, by binding to GSK-3β, releases β-catenin, preventing its degradation, however, the role of GSK3α/β in the adipocyte inflammatory response has not yet been fully investigated, therefore it seems promising to study the role of GSK-3 in the Wnt/β-catenin signaling pathway in obesityThe aim of the study was to assess the activity of the components of the Wnt signaling pathway in obese patients by measuring the serum level of GSK-3 and β-catenin. There were enrolled 32 patients with progressive forms of I-III degree obesity in the absence of diabetes mellitus. The concentration of serum GSK-3α, GSK-3β, and β-catenin was measured by enzyme-linked immunoassay. Data are presented as absolute and relative (%) number of patients; arithmetic mean; medians, 1 and 3 quartiles – Ме (Q0.25-Q0.75). Obese patients contained a 7.5-fold increased serum level of GSK-3α (785 (371-1317.5) pg/ml) compared to healthy individuals 105 (102.5-110) pg/ml, (p &lt; 0.001), paralleled with increased amount of GSK-3β, which level in obese patients was 295 (190-695) pg/ml, which is by 18.3% higher than those in healthy individuals 241 (218.75-287.5) pg/ml, p = 0.111. Amount of GSK-3 depending on the degree of obesity tended to increase, most often coupled to decreased β-catenin level which is consistent with the literature data and can be considered as a prognostic criterion for the course of pathological processes in obesity.

Effects of tauroursodeoxycholic acid on glucose homeostasis: Potential binding of this bile acid with the insulin receptor

AimsThe bile acid (BA), tauroursodeoxycholic acid (TUDCA) regulates glucose homeostasis; however, it is not clear whether its effects on insulin signaling are due to its direct interaction with the insulin receptor (IR) or through activation of the G-coupled BA receptor, TGR5. We, herein, investigated whether the actions of TUDCA on glucose homeostasis occur via IR or TGR5 activation. Main methodsGlucose homeostasis was evaluated in high-fat diet (HFD)-obese or control (CTL) mice, after 30 days or one intraperitoneal (ip) injection of 300 mg/kg TUDCA, respectively. Molecular docking was performed to investigate the potential binding of TUDCA on the IR and TGR5. Key findingsAfter 30 days of TUDCA treatment, HFD mice exhibited improvements in glucose tolerance and insulin sensitivity, which were abolished when these rodents received the IR antagonist, S961. Molecular docking experiments showed that TUDCA demonstrates high binding affinity for TGR5 and IR and strongly interacts with the insulin binding sites 1 and 2 of the IR. Consistent with this potential agonist activity of TUDCA on IR, CTL mice displayed increased hepatic phosphorylation of AKT after an ip injection of TUDCA. This effect was not associated with altered glycemia in CTL mice and was dependent on IR activation, as S961 prevented hepatic AKT activation by TUDCA. Furthermore, TUDCA activated the hepatic protein kinase A (PKA) and cAMP response element-binding protein (CREB) pathway in CTL mice, even after the administration of S961. SignificanceWe provide novel evidence that TUDCA may be an agonist of the IR, in turn activating AKT and contributing, at least in part, to its beneficial effects upon glucose homeostasis.

Insights From Single Cell RNA Sequencing Into the Immunology of Type 1 Diabetes- Cell Phenotypes and Antigen Specificity.

In the past few years, huge advances have been made in techniques to analyse cells at an individual level using RNA sequencing, and many of these have precipitated exciting discoveries in the immunology of type 1 diabetes (T1D). This review will cover the first papers to use scRNAseq to characterise human lymphocyte phenotypes in T1D in the peripheral blood, pancreatic lymph nodes and islets. These have revealed specific genes such as IL-32 that are differentially expressed in islet –specific T cells in T1D. scRNAseq has also revealed wider gene expression patterns that are involved in T1D and can predict its development even predating autoantibody production. Single cell sequencing of TCRs has revealed V genes and CDR3 motifs that are commonly used to target islet autoantigens, although truly public TCRs remain elusive. Little is known about BCR repertoires in T1D, but scRNAseq approaches have revealed that insulin binding BCRs commonly use specific J genes, share motifs between donors and frequently demonstrate poly-reactivity. This review will also summarise new developments in scRNAseq technology, the insights they have given into other diseases and how they could be leveraged to advance research in the type 1 diabetes field to identify novel biomarkers and targets for immunotherapy.