Insulin Receptor Associates with Promoters Genome-wide and Regulates Gene Expression.

Insulin receptor (IR) signaling is central to normal metabolic control and dysregulated in prevalent chronic diseases. IR binds insulin at the cell surface and transduces rapid signaling via cytoplasmic kinases. However, mechanisms mediating long-term effects of insulin remain unclear. Here, we show that IR associates with RNA polymerase II in the nucleus, with striking enrichment at promoters genome-wide. The target genes were highly enriched for insulin-related functions including lipid metabolism and protein synthesis and diseases including diabetes, neurodegeneration, and cancer. IR chromatin binding was increased by insulin and impaired in an insulin-resistant disease model. Promoter binding by IR was mediated by coregulator host cell factor-1 (HCF-1) and transcription factors, revealing an HCF-1-dependent pathway for gene regulation by insulin. These results show that IR interacts with transcriptional machinery at promoters and identify a pathway regulating genes linked to insulin's effects in physiology and disease.

Physiological and pathophysiological actions of insulin in the liver.

The liver plays an important role in the control of glucose homeostasis. When insulin levels are low, such as in the fasting state, gluconeogenesis and glycogenolysis are stimulated to maintain the blood glucose levels. Conversely, in the presence of increased insulin levels, such as after a meal, synthesis of glycogen and lipid occurs to maintain the blood glucose levels within normal range. Insulin receptor signaling regulates glycogenesis, gluconeogenesis and lipogenesis through downstream pathways such as the insulin receptor substrate (IRS)-phosphoinositide 3 (PI3) kinase-Akt pathway. IRS-1 and IRS-2 are abundantly expressed in the liver and are thought to be responsible for transmitting the insulin signal from the insulin receptor to the intracellular effectors involved in the regulation of glucose and lipid homeostasis. Impaired insulin receptor signaling can cause hepatic insulin resistance and lead to type 2 diabetes. In the present study, we focus on a concept called "selective insulin resistance," which has received increasing attention recently: the frequent coexistence of hyperglycemia and hepatic steatosis in people with type 2 diabetes and obesity suggests that it is possible for the insulin signaling regulating gluconeogenesis to be impaired even while that regulating lipogenesis is preserved, suggestive of selective insulin resistance. In this review, we review the progress in research on the insulin actions and insulin signaling in the liver.

Integrin and autocrine IGF2 pathways control fasting insulin secretion in ß-cells.

Elevated levels of fasting insulin release and insufficient glucose-stimulated insulin secretion (GSIS) are hallmarks of diabetes. Studies have established cross-talk between integrin signaling and insulin activity, but more details of how integrin-dependent signaling impacts the pathophysiology of diabetes are needed. Here, we dissected integrin-dependent signaling pathways involved in the regulation of insulin secretion in ß-cells and studied their link to the still debated autocrine regulation of insulin secretion by insulin/insulin-like growth factor (IGF) 2-AKT signaling. We observed for the first time a cooperation between different AKT isoforms and focal adhesion kinase (FAK)-dependent adhesion signaling, which either controlled GSIS or prevented insulin secretion under fasting conditions. Indeed, ß-cells form integrin-containing adhesions, which provide anchorage to the pancreatic extracellular matrix and are the origin of intracellular signaling via FAK and paxillin. Under low-glucose conditions, ß-cells adopt a starved adhesion phenotype consisting of actin stress fibers and large peripheral focal adhesion. In contrast, glucose stimulation induces cell spreading, actin remodeling, and point-like adhesions that contain phospho-FAK and phosphopaxillin, located in small protrusions. Rat primary ß-cells and mouse insulinomas showed an adhesion remodeling during GSIS resulting from autocrine insulin/IGF2 and AKT1 signaling. However, under starving conditions, the maintenance of stress fibers and the large adhesion phenotype required autocrine IGF2-IGF1 receptor signaling mediated by AKT2 and elevated FAK-kinase activity and ROCK-RhoA levels but low levels of paxillin phosphorylation. This starved adhesion phenotype prevented excessive insulin granule release to maintain low insulin secretion during fasting. Thus, deregulation of the IGF2 and adhesion-mediated signaling may explain dysfunctions observed in diabetes.

[Overview on hypoglycemic active constituents of traditional Chinese medicine based on insulin receptor signaling pathway].

Insulin resistance,as the main link in the pathogenesis of type 2 diabetes mellitus( T2 DM),runs through the whole process of occurrence and development of T2 DM and is closely related to the insulin receptor signaling pathway. Insulin stimulation causes autophosphorylation of the insulin receptor( IR),which then activates tyrosine phosphorylation of insulin receptor substrate( IRS).Phosphorylation of IRS can induce and activate phosphatidylinositol 3-kinase( PI3 K),subsequently activate downstream 3-phosphoinositide-dependent protease 1( PDK1) and Akt/PKB,and finally promote expression and translocation of glucose transporter 4 to increase glucose uptake of insulin-sensitive tissues and alleviate insulin resistance. Currently,oral hypoglycemic agents for clinical treatment of T2 DM have different side effects on the human body. Traditional Chinese medicine not only has a wide range of sources and abundant types,but also has comprehensive multi-component,multi-link and multi-target effects,showing unique advantages in the treatment of diabetes. In recent years,more and more researchers at home and abroad pay attention to the active ingredients in traditional Chinese medicine for alleviating insulin resistance. In this paper,we would summarize the active hypoglycemic ingredients of traditional Chinese medicine associated with the insulin receptor signaling pathway,which may provide some theoretical guidance for the development of traditional Chinese medicine in the treatment of diabetes.

The Biased G-Protein-Coupled Receptor Agonism Bridges the Gap between the Insulin Receptor and the Metabolic Syndrome.

Insulin signaling, as mediated through the insulin receptor (IR), plays a critical role in metabolism. Aberrations in this signaling cascade lead to several pathologies, the majority of which are classified under the umbrella term "metabolic syndrome". Although many of these pathologies are associated with insulin resistance, the exact mechanisms are not well understood. One area of current interest is the possibility of G-protein-coupled receptors (GPCRs) influencing or regulating IR signaling. This concept is particularly significant, because GPCRs have been shown to participate in cross-talk with the IR. More importantly, GPCR signaling has also been shown to preferentially regulate specific downstream signaling targets through GPCR agonist bias. A novel study recently demonstrated that this GPCR-biased agonism influences the activity of the IR without the presence of insulin. Although GPCR-IR cross-talk has previously been established, the notion that GPCRs can regulate the activation of the IR is particularly significant in relation to metabolic syndrome and other pathologies that develop as a result of alterations in IR signaling. As such, we aim to provide an overview of the physiological and pathophysiological roles of the IR within metabolic syndrome and its related pathologies, including cardiovascular health, gut microflora composition, gastrointestinal tract functioning, polycystic ovarian syndrome, pancreatic cancer, and neurodegenerative disorders. Furthermore, we propose that the GPCR-biased agonism may perhaps mediate some of the downstream signaling effects that further exacerbate these diseases for which the mechanisms are currently not well understood.

The Effects of Peripheral and Central High Insulin on Brain Insulin Signaling and Amyloid-ß in Young and Old APP/PS1 Mice.

Hyperinsulinemia is a risk factor for late-onset Alzheimer's disease (AD). In vitro experiments describe potential connections between insulin, insulin signaling, and amyloid-ß (Aß), but in vivo experiments are needed to validate these relationships under physiological conditions. First, we performed hyperinsulinemic-euglycemic clamps with concurrent hippocampal microdialysis in young, awake, behaving APPswe/PS1dE9 transgenic mice. Both a postprandial and supraphysiological insulin clamp significantly increased interstitial fluid (ISF) and plasma Aß compared with controls. We could detect no increase in brain, ISF, or CSF insulin or brain insulin signaling in response to peripheral hyperinsulinemia, despite detecting increased signaling in the muscle. Next, we delivered insulin directly into the hippocampus of young APP/PS1 mice via reverse microdialysis. Brain tissue insulin and insulin signaling was dose-dependently increased, but ISF Aß was unchanged by central insulin administration. Finally, to determine whether peripheral and central high insulin has differential effects in the presence of significant amyloid pathology, we repeated these experiments in older APP/PS1 mice with significant amyloid plaque burden. Postprandial insulin clamps increased ISF and plasma Aß, whereas direct delivery of insulin to the hippocampus significantly increased tissue insulin and insulin signaling, with no effect on Aß in old mice. These results suggest that the brain is still responsive to insulin in the presence of amyloid pathology but increased insulin signaling does not acutely modulate Aß in vivo before or after the onset of amyloid pathology. Peripheral hyperinsulinemia modestly increases ISF and plasma Aß in young and old mice, independent of neuronal insulin signaling. SIGNIFICANCE STATEMENT: The transportation of insulin from blood to brain is a saturable process relevant to understanding the link between hyperinsulinemia and AD. In vitro experiments have found direct connections between high insulin and extracellular Aß, but these mechanisms presume that peripheral high insulin elevates brain insulin significantly. We found that physiological hyperinsulinemia in awake, behaving mice does not increase CNS insulin to an appreciable level yet modestly increases extracellular Aß. We also found that the brain of aged APP/PS1 mice was not insulin resistant, contrary to the current state of the literature. These results further elucidate the relationship between insulin, the brain, and AD and its conflicting roles as both a risk factor and potential treatment.

Hepatitis B virus inhibits insulin receptor signaling and impairs liver regeneration via intracellular retention of the insulin receptor.

Hepatitis B virus (HBV) causes severe liver disease but the underlying mechanisms are incompletely understood. During chronic HBV infection, the liver is recurrently injured by immune cells in the quest for viral elimination. To compensate tissue injury, liver regeneration represents a vital process which requires proliferative insulin receptor signaling. This study aims to investigate the impact of HBV on liver regeneration and hepatic insulin receptor signaling. After carbon tetrachloride-induced liver injury, liver regeneration is delayed in HBV transgenic mice. These mice show diminished hepatocyte proliferation and increased expression of fibrosis markers. This is in accordance with a reduced activation of the insulin receptor although HBV induces expression of the insulin receptor via activation of NF-E2-related factor 2. This leads to increased intracellular amounts of insulin receptor in HBV expressing hepatocytes. However, intracellular retention of the receptor simultaneously reduces the amount of functional insulin receptors on the cell surface and thereby attenuates insulin binding in vitro and in vivo. Intracellular retention of the insulin receptor is caused by elevated amounts of α-taxilin, a free syntaxin binding protein, in HBV expressing hepatocytes preventing proper targeting of the insulin receptor to the cell surface. Consequently, functional analyses of insulin responsiveness revealed that HBV expressing hepatocytes are less sensitive to insulin stimulation leading to delayed liver regeneration. This study describes a novel pathomechanism that uncouples HBV expressing hepatocytes from proliferative signals and thereby impedes compensatory liver regeneration after liver injury.

RETRACTED: Quercetin suppresses insulin receptor signaling through inhibition of the insulin ligand-receptor binding and therefore impairs cancer cell proliferation.

Although the flavonoid quercetin is known to inhibit activation of insulin receptor signaling, the inhibitory mechanism is largely unknown. In this study, we demonstrate that quercetin suppresses insulin induced dimerization of the insulin receptor (IR) through interfering with ligand-receptor interactions, which reduces the phosphorylation of IR and Akt. This inhibitory effect further inhibits insulin stimulated glucose uptake due to decreased cell membrane translocation of glucose transporter 4 (GLUT4), resulting in impaired cancer cell proliferation. The effect of quercetin in inhibiting tumor growth was also evident in an in vivo model, indicating a potential future application for quercetin in the treatment of cancers.

4-Quinolone-3-carboxylic acids as cell-permeable inhibitors of protein tyrosine phosphatase 1B.

Protein tyrosine phosphatase 1B is a negative regulator in the insulin and leptin signaling pathways, and has emerged as an attractive target for the treatment of type 2 diabetes and obesity. However, the essential pharmacophore of charged phosphotyrosine or its mimetic confer low selectivity and poor cell permeability. Starting from our previously reported aryl diketoacid-based PTP1B inhibitors, a drug-like scaffold of 4-quinolone-3-carboxylic acid was introduced for the first time as a novel surrogate of phosphotyrosine. An optimal combination of hydrophobic groups installed at C-6, N-1 and C-3 positions of the quinolone motif afforded potent PTP1B inhibitors with low micromolar IC50 values. These 4-quinolone-3-carboxylate based PTP1B inhibitors displayed a 2-10 fold selectivity over a panel of PTP's. Furthermore, the bidentate inhibitors of 4-quinolone-3-carboxylic acids conjugated with aryl diketoacid or salicylic acid were cell permeable and enhanced insulin signaling in CHO/hIR cells. The kinetic studies and molecular modeling suggest that the 4-quinolone-3-carboxylates act as competitive inhibitors by binding to the PTP1B active site in the WPD loop closed conformation. Taken together, our study shows that the 4-quinolone-3-carboxylic acid derivatives exhibit improved pharmacological properties over previously described PTB1B inhibitors and warrant further preclinical studies.

BRD7 improves glucose homeostasis independent of IRS proteins.

Bromodomain-containing protein 7 (BRD7) has emerged as a player in the regulation of glucose homeostasis. Hepatic BRD7 levels are decreased in obese mice, and the reinstatement of hepatic BRD7 in obese mice has been shown to establish euglycemia and improve glucose homeostasis. Of note, the upregulation of hepatic BRD7 levels activates the AKT cascade in response to insulin without enhancing the sensitivity of the insulin receptor (InsR)-insulin receptor substrate (IRS) axis. In this report, we provide evidence for the existence of an alternative insulin signaling pathway that operates independently of IRS proteins and demonstrate the involvement of BRD7 in this pathway. To investigate the involvement of BRD7 as a downstream component of InsR, we utilized liver-specific InsR knockout mice. Additionally, we employed liver-specific IRS1/2 knockout mice to examine the requirement of IRS1/2 for the action of BRD7. Our investigation of glucose metabolism parameters and insulin signaling unveiled the significance of InsR activation in mediating BRD7's effect on glucose homeostasis in the liver. Moreover, we identified an interaction between BRD7 and InsR. Notably, our findings indicate that IRS1/2 is not necessary for BRD7's regulation of glucose metabolism, particularly in the context of obesity. The upregulation of hepatic BRD7 significantly reduces blood glucose levels and restores glucose homeostasis in high-fat diet-challenged liver-specific IRS1/2 knockout mice. These findings highlight the presence of an alternative insulin signaling pathway that operates independently of IRS1/2 and offer novel insights into the mechanisms of a previously unknown insulin signaling in obesity.

Effects of alcohol and PARP inhibition on RNA ribosomal engagement in cortical excitatory neurons.

We report on the effects of ethanol (EtOH) and Poly (ADP-ribose) polymerase (PARP) inhibition on RNA ribosomal engagement, as a proxy for protein translation, in prefrontal cortical (PFC) pyramidal neurons. We hypothesized that EtOH induces a shift in RNA ribosomal-engagement (RE) in PFC pyramidal neurons, and that many of these changes can be reversed using a PARP inhibitor. We utilized the translating ribosome affinity purification (TRAP) technique to isolate cell type-specific RNA. Transgenic mice with EGFP-tagged Rpl10a ribosomal protein expressed only in CaMKIIα-expressing pyramidal cells were administered EtOH or normal saline (CTL) i.p. twice a day, for four consecutive days. On the fourth day, a sub-group of mice that received EtOH in the previous three days received a combination of EtOH and the PARP inhibitor ABT-888 (EtOH + ABT-888). PFC tissue was processed to isolate both, CaMKIIα pyramidal cell-type specific ribosomal-engaged RNA (TRAP-RNA), as well as genomically expressed total-RNA from whole tissue, which were submitted for RNA-seq. We observed EtOH effects on RE transcripts in pyramidal cells and furthermore treatment with a PARP inhibitor "reversed" these effects. The PARP inhibitor ABT-888 reversed 82% of the EtOH-induced changes in RE (TRAP-RNA), and similarly 83% in the total-RNA transcripts. We identified Insulin Receptor Signaling as highly enriched in the ethanol-regulated and PARP-reverted RE pool and validated five participating genes from this pathway. To our knowledge, this is the first description of the effects of EtOH on excitatory neuron RE transcripts from total-RNA and provides insights into PARP-mediated regulation of EtOH effects.

Long-Term Hyperglycemia Causes Depressive Behaviors in Mice with Hypoactive Glutamatergic Activity in the Medial Prefrontal Cortex, Which Is Not Reversed by Insulin Treatment.

The etiology of hyperglycemic-induced depressive behaviors is unclear. We hypothesized that long-term hyperglycemia may induce long-lasting disturbances in glutamatergic signaling and neural damages, causing depressive behaviors. To prove our hypothesis, a C57BL/6N mouse model of hyperglycemia was maintained for 4 weeks (equivalent to approximately 3 years in humans), after which insulin treatment was administered for an additional 4 weeks to normalize hyperglycemia-induced changes. Hyperglycemic mice showed depressive-like behaviors. Glutamatergic neurons and glial cells in the medial prefrontal cortex (mPFC) were affected by hyperglycemia. Insulin treatment improved blood glucose, water intake, and food intake to normoglycemic levels, but did not improve depressive-like behaviors. Glutamatergic signaling decreased with long-term hyperglycemia and did not normalize with insulin-induced normoglycemia. Importantly, hyperglycemia-induced changes in the mPFC were almost not reversed by the 4-week insulin treatment. In particular, levels of insulin receptor beta subunit (IRß), IRS-1, vesicular glutamate transporter 1, glutamine transporter SNAT2, phosphate-activated glutaminase, and GLUT-3 were not changed by insulin. Nitration and the dephosphorylation of IRß in the PFC also did not improve with insulin treatment. Therefore, our results suggest that hypoactive glutamatergic activity in the mPFC is involved in diabetic-associated depressive behaviors, and it is difficult to cure with glycemic regulation alone.

Brain insulin resistance and the therapeutic value of insulin and insulin-sensitizing drugs in Alzheimer's disease neuropathology.

The incidence of Alzheimer's disease (AD) is significantly higher in people with diabetes. Insulin and insulin receptor (IR) signaling intermediates are expressed in the brain. Insulin exerts multiple function in the brain. The role of compromised IR signaling in AD pathogenesis and the therapeutic value of insulin attract broad attention. This review summarizes the collective insulin action in the brain related to key factors of AD pathogenesis, updates the key features of insulin resistance in the AD brain and assesses the therapeutic potential of insulin and insulin-sensitizing drugs. Insulin stimulates neural growth and survival, suppresses amyloidogenic processing of the amyloid precursor protein (AßPP) and inhibits the Tau phosphorylation kinase, glycogen synthase kinase 3ß. Central nervous IR signaling regulates systemic metabolism and increases glucose availability to neurons. The expression of IR and its downstream effectors is reduced in AD brain tissues. Insulin and insulin-sensitizing drugs can improve cognitive function in AD patients and AD animal models. Systemic insulin delivery is less effective than intranasal insulin treatment. The penetrance of insulin-sensitizing drugs to the blood brain barrier is problematic and new brain-prone drugs need be developed. Insulin resistance manifested by the degradation and the altered phosphorylation of IR intermediates precedes overt AD syndrome. Type 3 diabetes as a pure form of brain insulin resistance without systemic insulin resistance is proposed as a causal factor in AD. Further research is needed for the identification of critical factors leading to impaired IR signaling and the development of new molecules to stimulate brain IR signaling.

Comparative proteomic analysis of insulin receptor isoform A and B signaling.

The insulin receptor (IR) gene undergoes differential splicing generating two IR isoforms, IR-A and IR-B. The roles of IR-A in cancer and of IR-B in metabolic regulation are well known but the molecular mechanisms responsible for their different biological effects are poorly understood. We aimed to identify different or similar protein substrates and signaling linked to each IR isoforms. We employed mouse fibroblasts lacking IGF1R gene and expressing exclusively either IR-A or IR-B. By proteomic analysis a total of 2530 proteins were identified and quantified. Proteins and pathways mostly associated with insulin-activated IR-A were involved in cancer, stemness and interferon signaling. Instead, proteins and pathways associated with insulin-stimulated IR-B-expressing cells were mostly involved in metabolic or tumor suppressive functions. These results show that IR-A and IR-B recruit partially different multiprotein complexes in response to insulin, suggesting partially different functions of IR isoforms in physiology and in disease.

A Neonatal Mild Defect in Brain Insulin Signaling Predisposes a Subclinical Model of Sporadic Alzheimer's to Develop the Disease.

Brain insulin system dysfunction has been proposed as a key player in the pathogenesis of sporadic Alzheimer's disease (sAD). Given this fact, an adult rat model for sAD has been developed by intracerebroventricular injection of a subdiabetogenic streptozotocin dosage (icv-STZ). A low dose of icv-STZ in adult rats leads to a subclinical model of Alzheimer's disease. According to the brain developmental origin for sAD occurrence, the present study evaluated the effect of neonatal injection of icv-STZ on the development and progression of Alzheimer's disease later in the adult animals treated with a low dose of icv-STZ. Although no alteration was observed in the rats receiving an adult low dose of icv-STZ, these animals displayed cognitive deficits if they were also treated neonatally with icv-STZ. These impairments were associated with altered gene expression of insulin receptor, tau and choline acetyltransferase, along with increased astrocyte and dark neuron densities in the hippocampus. This study highlights neonatal brain insulin system dysfunction in the programming of brain insulin signaling sensitivity and provides more evidence for the developmental origin of sAD.

Renal gluconeogenesis in insulin resistance: A culprit for hyperglycemia in diabetes.

Renal gluconeogenesis is one of the major pathways for endogenous glucose production. Impairment in this process may contribute to hyperglycemia in cases with insulin resistance and diabetes. We reviewed pertinent studies to elucidate the role of renal gluconeogenesis regulation in insulin resistance and diabetes. A consensus on the suppressive effect of insulin on kidney gluconeogenesis has started to build up. Insulin-resistant models exhibit reduced insulin receptor (IR) expression and/or post-receptor signaling in their kidney tissue. Reduced IR expression or post-receptor signaling can cause impairment in insulin's action on kidneys, which may increase renal gluconeogenesis in the state of insulin resistance. It is now established that the kidney contributes up to 20% of all glucose production via gluconeogenesis in the post-absorptive phase. However, the rate of renal glucose release excessively increases in diabetes. The rise in renal glucose release in diabetes may contribute to fasting hyperglycemia and increased postprandial glucose levels. Enhanced glucose release by the kidneys and renal expression of the gluconeogenic-enzyme in diabetic rodents and humans further point towards the significance of renal gluconeogenesis. Overall, the available literature suggests that impairment in renal gluconeogenesis in an insulin-resistant state may contribute to hyperglycemia in type 2 diabetes.

S-Propargyl-Cysteine Attenuates Diabetic Cardiomyopathy in db/db Mice Through Activation of Cardiac Insulin Receptor Signaling.

Background: Endogenous hydrogen sulfide (H2S) is emerging as a key signal molecule in the development of diabetic cardiomyopathy. The aim of this study was to explore the effect and underlying mechanism of S-propargyl-cysteine (SPRC), a novel modulator of endogenous H2S, on diabetic cardiomyopathy in db/db diabetic mice. Methods and Results: Vehicle or SPRC were orally administered to 8-month-old male db/db mice and their wild type littermate for 12 weeks. SPRC treatment ameliorated myocardial hypertrophy, fibrosis, and cardiac systolic dysfunction assessed by histopathological examinations and echocardiography. The functional improvement by SPRC was accompanied by a reduction in myocardial lipid accumulation and ameliorated plasma lipid profiles. SPRC treatment improved glucose tolerance in db/db mice, with fasting blood glucose and peripheral insulin resistance remaining unchanged. Furthermore, insulin receptor signaling involving the phosphorylation of protein kinase B (Akt/PKB) and glycogen synthase kinase 3ß (GSK3ß) were elevated and activated by SPRC treatment. Primary neonatal mice cardiomyocytes were cultured to explore the mechanisms of SPRC on diabetic cardiomyopathy in vitro. Consistent with the results in vivo, SPRC not only up-regulated insulin receptor signaling pathway in cardiomyocytes in dose-dependent manner in the basal state, but also relieved the suppression of insulin receptor signaling induced by high concentrations of glucose and insulin. Furthermore, SPRC also enhanced the expression of glucose transporter 4 (GLUT4) and 3H glucose uptake in cardiomyocytes. Conclusions: In this study, we found a novel beneficial effect of SPRC on diabetic cardiomyopathy, which was associated with activation of insulin receptor signaling. SPRC may be a promising medication for diabetic cardiomyopathy in type 2 diabetes mellitus patients.

Yunpi Heluo decoction attenuates insulin resistance by regulating liver miR-29a-3p in Zucker diabetic fatty rats.

BACKGROUND AND OBJECTIVE: Yunpiheluo (YPHL) decoction is a Chinese herbal formula with unique advantages for the treatment of type 2 diabetes mellitus (T2DM). The aim of the present study was to investigate changes in miRNA expression and downstream gene expression in Zucker diabetic fatty (ZDF) rats treated with YPHL to determine whether YPHL could be used as an adjuvant treatment of T2DM. METHODS: Serum and liver total cholesterol (TC) and triglycerides (TG) levels, insulin resistance index (IR) and differentially expressed miRNAs were detected in a T2DM ZDF rat model. miRNA target prediction was based on bioinformatic algorithms and dual luciferase reporter assay. Protein expression of genes in the insulin receptor signaling pathway was detected by Western blot. The IR cell model was established and the effects of lyophilized YPHL powder on the protein expressions were observed by transfecting specific miRNA mimics and inhibitors. RESULTS: The miR-29a-3p expression level was significantly increased in the liver of ZDF rats. Insulin receptor substrate 1 (IRS1) was the target gene of miR-29a-3p. IRS1 mRNA and protein expressions of IRS1, IRS1 (phospho S307), protein kinase B (Akt), Akt (phosphor ser473) and pyruvate dehydrogenase lipoamide kinase isozyme 1 (PDK1) were decreased significantly. miR-29a-3p over-expression decrease IRS1 and the others protein expressions in the HepG2 IR cell model while anti-miR-29a-3p showed the opposite result. The miR-29a-3p level was decreased, and the expressions of IRS1 mRNA and the above proteins were all increased after YPHL treatment. CONCLUSION: miR-29a-3p played a functional role in insulin receptor signaling in the liver of ZDF rats. YPHL decoction attenuated IR in T2DM probably by down-regulating or maintaining the miR-29a-3p level, increasing the expression of IRS1 mRNA and its phosphorylated proteins, and regulating the expression of insulin receptor signaling-related proteins. YPHL may prove to be an alternative treatment for T2DM.

Mechanisms of insulin resistance by simvastatin in C2C12 myotubes and in mouse skeletal muscle.

Statins inhibit cholesterol biosynthesis and lower serum LDL-cholesterol levels. They are generally well tolerated, but can cause insulin resistance in patients. Therefore, we investigated the mechanisms underlying the statin-induced insulin resistance. We used mice and C2C12 myotubes (murine cell line): mice (n = 10) were treated with oral simvastatin (5 mg/kg/day) or water (control) for 21 days and C2C12 cells were exposed to 10 µM simvastatin for 24 h. After intraperitoneal glucose application (2 g/kg), simvastatin-treated mice had higher glucose but equal insulin plasma concentrations than controls and lower glucose transport into skeletal muscle. Similarly, glucose uptake by C2C12 myotubes exposed to 10 µM simvastatin for 24 h was impaired compared to control cells. In simvastatin-treated C2C12 myotubes, mRNA and protein expression of the insulin receptor (IR) ß-chain was increased, but the phosphorylation (Tyr1361) was impaired. Simvastatin decreased numerically Akt/PKB Thr308 phosphorylation (via insulin signaling pathway) and significantly Akt/PKB Ser473 phosphorylation (via mTORC2), which was explained by impaired phosphorylation of mTOR Ser2448. Reduced phosphorylation of Akt/PKB impaired downstream phosphorylation of GSK3ß, leading to impaired translocation of GLUT4 into plasma membranes of C2C12 myotubes. In contrast, reduced phosphorylation of AS160 could be excluded as a reason for impaired GLUT4 translocation. In conclusion, simvastatin caused insulin resistance in mice and impaired glucose uptake in C2C12 myotubes. The findings in myotubes can be explained by diminished activation of Akt/PKB by mTORC2 and downstream effects on GSK3ß, impairing the translocation of GLUT4 and the uptake of glucose.

Non-Nutritive Sweeteners and Their Implications on the Development of Metabolic Syndrome.

Individuals widely use non-nutritive sweeteners (NNS) in attempts to lower their overall daily caloric intake, lose weight, and sustain a healthy diet. There are insufficient scientific data that support the safety of consuming NNS. However, recent studies have suggested that NNS consumption can induce gut microbiota dysbiosis and promote glucose intolerance in healthy individuals that may result in the development of type 2 diabetes mellitus (T2DM). This sequence of events may result in changes in the gut microbiota composition through microRNA (miRNA)-mediated changes. The mechanism(s) by which miRNAs alter gene expression of different bacterial species provides a link between the consumption of NNS and the development of metabolic changes. Another potential mechanism that connects NNS to metabolic changes is the molecular crosstalk between the insulin receptor (IR) and G protein-coupled receptors (GPCRs). Here, we aim to highlight the role of NNS in obesity and discuss IR-GPCR crosstalk and miRNA-mediated changes, in the manipulation of the gut microbiota composition and T2DM pathogenesis.