Protective Effects of Astragaloside IV on Uric Acid-Induced Pancreatic ß-Cell Injury through PI3K/AKT Pathway Activation.

BACKGROUND: Elevated uric acid (UA) has been found to damage pancreatic ß-cell, promote oxidative stress, and cause insulin resistance in type 2 diabetes (T2D). Astragaloside IV (AS-IV), a major active monomer extracted from Astragalus membranaceus (Fisch.) Bunge. which belongs to TRIB. Galegeae (Br.) Torrey et Gray, Papilionaceae, exhibits various activities in a pathophysiological environment and has been widely employed to treat diseases. However, the effects of AS-IV on UA-induced pancreatic ß-cell damage need to be investigated and the associating mechanism needs to be elucidated. This study was designed to determine the protective effects and underlying mechanism of AS-IV on UA-induced pancreatic ß-cell dysfunction in T2D. METHODS: UA-treated Min6 cells were exposed to AS-IV or wortmannin. Thereafter, the 3-(45)-dimethylthiahiazo(-z-y1)-35-di-phenytetrazoliumromide (MTT) assay and flow cytometry were employed to determine the effect of AS-IV on cell proliferation and apoptosis, respectively. Insulin secretion was evaluated using the glucose-stimulated insulin secretion (GSIS) assay. Finally, western blot and quantitative real-time polymerase chain reaction (qRT-PCR) were performed to determine the effect of AS-IV on the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway in UA-treated cells. RESULTS: AS-IV had no cytotoxic effects on Min6 cells. UA significantly suppressed Min6 cell growth, promoted cell apoptosis, and enhanced caspase-3 activity; however, AS-IV abolished these effects in a dose-dependent manner. Further, decreased insulin secretion was found in UA-treated Min6 cells compared to control cells, and the production of insulin was enhanced by AS-IV in a dose-dependent manner. AS-IV significantly increased phosphorylated (p)-AKT expression and the ratio of p-AKT/AKT in Min6 cells exposed to UA. No evident change in AKT mRNA level was found in the different groups. However, the effects of AS-IV on UA-stimulated Min6 cells were reversed by 100 nM wortmannin. CONCLUSION: Collectively, our data suggest that AS-IV protected pancreatic ß-cells from UA-treated dysfunction by activating the PI3K/AKT pathway. Such findings suggest that AS-IV may be an efficient natural agent against T2D.

Impact of inflammatory markers on the relationship between sleep quality and diabetic kidney disease.

PURPOSE: Both poor sleep and diabetic kidney disease are closely associated with inflammation. However, the correlation between poor sleep and diabetic kidney disease has not been well clarified. Thus, the aim of this study was to determine the mediating role that inflammatory markers play in the pathogenic effect of poor sleep on the severity of diabetic kidney disease (DKD). METHODS: A cross-sectional survey was conducted on 336 patients with type 2 diabetes (T2D). DKD was diagnosed according to the guidelines of the National Kidney Foundation-Kidney Disease Outcome Quality Initiative (NKF-K/DOQI). The Pittsburg Sleep Quality Index (PSQI) score was applied to assess patients for the quality of their sleep. Patients with a PSQI score of more than 5 were assigned to the poor sleep group, and the rest of the patients were assigned to the good sleep group. Circulating levels of six inflammatory biomarkers related to poor sleep and DKD were measured. RESULTS: The prevalence of DKD was higher in patients with poor sleep quality than in those with good sleep quality (42% vs. 25%, P = 0.002). After adjustment, poor sleep quality (PSQI score OR 1.075 [95%CI 1.018-1.135], P = 0.009) remained independently associated with DKD. PSQI score was found to be positively related to fibroblast growth factor (FGF23), interleukin 6 (IL-6), P-selectin, and intercellular adhesion molecule-1 (ICAM-1) (P < 0.01), rather than fibrinogen and C-reactive protein (CRP) in linear regression models. As revealed by multiple mediation analysis, FGF23 and IL-6 mediated 26% and 23% of the relationship between PSQI score and urinary microalbumin (UMA), respectively. Similarly, the FGF23 and ICAM-1, instead of IL-6 and P-selectin, mediated 32% and 24% of the association between PSQI and estimated glomerular filtration rate (eGFR), respectively. CONCLUSIONS: Poor sleep quality is independently associated with DKD. These results suggest that inflammatory markers contribute to a pathogenic connection between poor sleep and DKD.

Circulating Serum Myonectin Levels in Obesity and Type 2 Diabetes Mellitus.

OBJECTIVE: Myonectin is one of the myokines and has gained interest as a potential new strategy to combat obesity and its associated disorders, such as type 2 diabetes mellitus (T2DM).The objective of this study was to investigate circulating serum myonectin levels in nondiabetes and T2DM and elucidate possible relationships between serum myonectin levels and metabolic parameters in patients with T2DM. DESIGN: A total of 362 Chinese patients with T2DM and 100 age- and sex-matched healthy controls were recruited in this study. Clinical characteristics, blood biochemistry, and circulating myonectin levels were measured by enzyme-linked immunosorbent assay. RESULTS: Circulating myonectin levels were significantly decreased in T2DM compared with controls. Obese nondiabetic controls had significantly lower serum myonectin levels compared with lean nondiabetic controls. In diabetic patients, serum myonectin concentrations were significantly negatively correlated with body mass index (BMI), total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), C-reactive protein (CRP), hemoglobin A1c (HbA1c), fasting insulin (Fins), the homeostatic model assessment of insulin resistance (HOMA-IR), visceral fat area, and subcutaneous fat area. After adjusting for covariates, multivariate stepwise regression analysis demonstrated that BMI, LDL-C, TG, HOMA-IR, and visceral fat were the main independent predictors of low serum myonectin concentrations. CONCLUSIONS: Circulating myonectin levels were decreased in T2DM patients and in obese subjects. Moreover, serum myonectin levels were correlated with metabolic markers of T2DM. These data suggest that myonectin may be a useful marker in predicting the development of obesity and T2DM.

miRNA­342 suppresses renal interstitial fibrosis in diabetic nephropathy by targeting SOX6.

Diabetic kidney disease (DKD) is one of the major microvascular complications in patients with type 1 and/or type 2 diabetes, the first cause of end­stage renal disease (ESRD) in several countries and regions. However, the pathogenesis of DKD and the mechanisms through which it leads to ESRD remain unknown. Thus, in this study, we aimed to elucidate some of these mechanisms. The expression of microRNA (miRNA or miR)­342­3p and SRY­box 6 (SOX6) in the renal tissues of mice with DKD and mouse renal mesangial cells (MCs) was determined by RT­qPCR and western blot analysis. The diabetic kidney environment was established using high­glucose medium. SOX6 was verified as a target gene of miR­342­3p by dual­luciferase activity assay. In addition, western blot analysis was employed to determine the changes in the levels of several biomarkers of fibrosis [transforming growth factor (TGF)­ß1, fibronectin (FN), collagen IV (referred to as C­IV) and phosphatase and tensin homolog (PTEN)]. Compared with THE control mice, the expression of miR­342­3p in the kidney tissues of mice with DKD was downregulated, whereas that of SOX6 was upregulated. The same phenomenon was observed in the MCs cultured in high­glucose medium. Subsequently, miR­342­3p inhibited SOX6 expression, promoted cell proliferation and inhibited the apoptosis of MCs. Moreover, the overexpression of miR­342­3p suppressed high glucose­induced renal interstitial fibrosis. In addition, it was found that miR­342­3p inhibited SOX6 expression by binding to the 3'­UTR of SOX6. On the whole, the findings of this study demonstrate that miR­342­3p suppresses the progression of DKD by inducing the degradation of SOX6. Thus, the miR­342­3p/SOX6 axis may serve as a novel therapeutic target in the treatment of DKD.

Association between serum total bilirubin levels and the risk of type 2 diabetes mellitus.

AIM: To confirm whether serum bilirubin is an independent risk factor of type 2 diabetes mellitus (T2DM) onset in patients with impaired fasting glycemia (IFG) and impaired glucose tolerance (IGT). METHODS: This was a prospective cohort study carried out at the Diabetic Identification Center of Tianjin Metabolic Diseases Hospital. Serum total bilirubin (TBIL) was measured at baseline and the patients were grouped according to baseline bilirubin quartiles. The outcome was the confirmation of T2DM by oral glucose tolerance test (OGTT) during the 3-year follow-up. Logistic regression was used to determine the risk factors for T2DM development and whether bilirubin levels are independently associated with T2DM development. RESULTS: Finally, 523 patients were analyzed. After 3 years, 310 participants were diagnosed with diabetes based on OGTT. Baseline quartiles of total bilirubin were inversely associated with diabetes risk, even after multivariable adjustment. The adjusted ORs for diabetes were 1.0 (reference), 0.83 (95% CI 0.74-0.96), 0.78 (95% CI 0.68-0.90), 0.74 (95% CI 0.64-0.87) for the 1st, 2nd, 3rd, and 4th quartiles of baseline serum total bilirubin, respectively (P < 0.001). CONCLUSION: In patients with IFG or IGT, low levels of serum total bilirubin were associated with a significantly increased risk of T2DM.

Apigenin prevents metabolic syndrome in high-fructose diet-fed mice by Keap1-Nrf2 pathway.

Chronic dietary high fructose leads to various kinds of undesirable metabolic effects. Apigenin, a naturally occurring plant flavone, is plentiful in fruits and vegetables. The aim of this study was to identify the protective effects of apigenin on metabolic syndrome and elucidate potential underlying mechanisms. The animal model was established by 4-weeks high fructose feeding. Insulin resistance was estimated by oral glucose tolerance test and homeostasis model assessment-insulin resistance index. Liver function was evaluated by serum AST and ALT, hepatic histopathological alternation, and lipid accumulation in the liver. The alterations of lipid profile was evaluated by TG, TC, LDL-C and HDL-C levels in serum. Administration of apigenin exerted beneficial effects through improving insulin resistance, alleviating liver injury, and inhibiting the alterations of lipid profile in high fructose-fed mice. In addition, apigenin potently facilitated the accumulation of Nrf2 nuclear translocation and accompanied by increasing HO-1 and NQO1 protein expressions, which are responsible for attenuating oxidative stress. Molecular docking results demonstrated that potential interaction of apigenin with the Nrf2-binding site in the Keap1 protein. In summary, we demonstrated that apigenin prevented high fructose-induced metabolic syndrome probably by inhibiting binding of Keap1 to Nrf2, and thus Nrf2 nuclear translocation, subsequently resulting in increased the expressions of anti-oxidative genes including HO-1 and NQO1.

Honokiol protects pancreatic ß cell against high glucose and intermittent hypoxia-induced injury by activating Nrf2/ARE pathway in vitro and in vivo.

Obstructive sleep apnea hypopnea syndrome (OSAHS) is associated with glucose intolerance, insulin resistance and type 2 diabetes mellitus (T2DM). Although several studies have revealed that intermittent hypoxia (IH) in OSAHS may further aggravate pancreatic ß cell damage and promote the evolution of type 2 diabetes (T2DM) by increasing oxidative stress, the underlying mechanisms are unclear. Honokiol, a potent radical scavenger, has been demonstrated to ameliorate oxidative stress in many cases. The present study aimed to explore the potential mechanism of IH and diabetes synergistically damage and destruct the pancreatic ß cell, examine the effects of honokiol on ameliorating pancreatic ß cell injury in this context and explore the mechanism of such effects. High glucose (HG) cultured INS-1 cells were exposed to 50 µM of honokiol for 24, 48 and 72 h with or without IH intervention. T2DM rats were treated with honokiol and exposed to 80 s of IH followed by 160 s of normoxia for 8 weeks. The cell proliferation, apoptosis and oxidative stress were measured. Blood glucose, insulin, glucagon and HOMA-IR (Homeostasis model assessment -insulin resistence) were also detected, and the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) were detected by immunofluorescence staining and western blotting. Honokiol can reduce oxidative stress, cytotoxicity and apoptosis in the INS-1 cells of rats receiving HG treatment or both HG and IH treatment. IH can further aggravate pancreas dysfunction, cause a marked elevation in fasting blood glucose, glucagon, HOMA-IR and oxidative stress levels in DM rats. In addition, honokiol can effectively activate the Nrf2/ARE pathway and reverse this pancreatic dysfunction in vivo and in vitro. These findings indicate that honokiol acts as a potent ROS scavenger via Nrf2/ARE pathway and effectively attenuates oxidative stress and improves pancreatic ß cell function of DM rats under IH treatment.

Correlation between Insulin Resistance and Thyroid Nodule in Type 2 Diabetes Mellitus.

OBJECTIVE: The present study explored the association between insulin resistance (IR) and the clinical characteristics of thyroid nodules in patients with type 2 diabetes mellitus (T2DM). METHODS: All the patients were newly diagnosed with T2DM. 201 patients with thyroid nodule disease and 308 patients without the nodular thyroid disease. The participants were evaluated by relevant examination. Correlation analyses and regression analyses were performed to examine the relationships between the two groups. RESULTS: HOMA-IR values, serum FT4 (free thyroxine) levels, and age were higher in the thyroid nodule group than in the control group. The proportion of women in the thyroid nodule group is greater than the proportion of women in the control group. Logistic regression analysis showed that age, sex, FT4, and HOMA-IR were positive factors for thyroid nodule. The volume and size of the thyroid nodule were positively correlated with HOMA-IR, irrespective of gender. The thyroid nodule volume and size and the TSH (thyroid stimulating hormone) were greater in females than in males, whereas FT3 (free triiodothyronine) was lower in females. CONCLUSION: IR might be a risk factor for thyroid nodule. Whether alleviating the IR might slow the growth, or diminish the volume and size of the thyroid nodules, is yet to be elucidated.

The relationship between circulating irisin levels and tissues AGE accumulation in type 2 diabetes patients.

Advanced glycation end-products (AGEs), measured by skin autofluorescence (AF), are a factor in the development or worsening of many degenerative diseases, such as diabetes and atherosclerosis. Irisin levels have been associated with diabetes, endothelial dysfunction and atherosclerosis. The objective of the present study was to investigate whether circulating irisin levels are correlated with skin AF values in type 2 diabetes patients. A total of 362 Chinese type 2 diabetic patients and 100 age- and sex-matched healthy controls were recruited in the present study. Clinical characteristics, blood biochemistry and circulating irisin levels were measured. Skin AF was measured using an AGE reader. Circulating irisin levels were significantly lower, while skin AF values were increased in type 2 diabetes compared with controls (P<0.05 respectively). By dividing the distribution of skin AF values into tertiles, serum irisin levels gradually lowered with increasing skin AF values (P<0.05). After adjusting for covariates, multivariate stepwise regression analysis demonstrated that serum lower irisin levels were independently associated with skin AF (P=0.009). Circulating irisin levels were lower in type 2 diabetes patients compared with healthy controls. Lower levels of irisin are independently associated with elevated skin AF values, indicating that circulating irisin levels could be associated with AGEs accumulation, which is one of the reasons causing vascular complications in diabetic patients.

Altered brain activation and functional connectivity in working memory related networks in patients with type 2 diabetes: An ICA-based analysis.

Type 2 diabetes mellitus (T2DM) can cause multidimensional cognitive deficits, among which working memory (WM) is usually involved at an early stage. However, the neural substrates underlying impaired WM in T2DM patients are still unclear. To clarify this issue, we utilized functional magnetic resonance imaging (fMRI) and independent component analysis to evaluate T2DM patients for alterations in brain activation and functional connectivity (FC) in WM networks and to determine their associations with cognitive and clinical variables. Twenty complication-free T2DM patients and 19 matched healthy controls (HCs) were enrolled, and fMRI data were acquired during a block-designed 1-back WM task. The WM metrics of the T2DM patients showed no differences compared with those of the HCs, except for a slightly lower accuracy rate in the T2DM patients. Compared with the HCs, the T2DM patients demonstrated increased activation within their WM fronto-parietal networks, and activation strength was significantly correlated with WM performance. The T2DM patients also showed decreased FC within and between their WM networks. Our results indicate that the functional integration of WM sub-networks was disrupted in the complication-free T2DM patients and that strengthened regional activity in fronto-parietal networks may compensate for the WM impairment caused by T2DM.

Efficacy and safety of vildagliptin, sitagliptin, and linagliptin as add-on therapy in Chinese patients with T2DM inadequately controlled with dual combination of insulin and traditional oral hypoglycemic agent.

OBJECTIVE: We aimed to evaluate the efficacy and safety of the three dipeptidyl peptidase 4 (DPP-4) inhibitors (vildagliptin, sitagliptin, and linagliptin) as add-on therapy in Chinese patients with type 2 diabetes mellitus (T2DM)inadequately controlled on dual combination of insulin and metformin or acarbose. METHODS: A total of 535 T2DM patients who failed to achieve glycemic control with insulin and a traditional oral hypoglycemic agent were randomized to receive vildagliptin, sitagliptin, or linagliptin. Body mass index, glycosylated hemoglobin (HbA1c), fasting and postprandial plasma glucose (FPG and PPG), insulin dose, and adverse events were evaluated during the study. RESULTS: The baseline HbA1c was 9.59 ± 1.84 % (vildagliptin group), 9.22 ± 1.60 % (sitagliptin group), and 9.58 ± 1.80 % (linagliptin group). At week 12 it was 8.16 ± 1.29 % (vildagliptin), 8.56 ± 1.96 % (linagliptin), and 8.26 ± 1.10 % (sitagliptin). The changes in HbA1c from baseline were -1.33 ± 0.11 % (vildagliptin), -0.84 ± 0.08 % (sitagliptin) and -0.81 ± 0.08 % (linagliptin), the vildagliptin group had the greatest reduction in HbA1c (P < 0.05). The proportions of patients that reached target HbA1c were 66.27 % (vildagliptin), 52.73 % (sitagliptin), and 55.49 % (linagliptin), the vildagliptin group had the highest one (P < 0.05). The baseline FPG and PPG values in the three groups were at the same level. At week 12, mean FPG levels in the vildagliptin (7.31 ± 1.50 mmol/L) and linagliptin (6.90 ± 1.55 mmol/L) groups were significantly lower than in the sitagliptin group (8.02 ± 4.48 mmol/L; P < 0.05); the linagliptin group had the lowest mean PPG followed by the vildagliptin group which was also significant lower (P = 0.000) than the sitagliptin group. Additionally, the required insulin dosage in the vildagliptin group was the lowest among the groups at weeks 6 and 12. Only mild AEs were reported during the study. CONCLUSION: The three DPP-4 inhibitors appear to be effective and safe as add-on therapy for T2DM patients on dual combination of insulin and a traditional OHA. Vildagliptin was more effective in decreasing insulin requirement and achieving glycemic control when compared to the other two.

Impact of inflammatory markers on the relationship between sleep quality and incident cardiovascular events in type 2 diabetes.

AIMS: To analyze relevance of sleep quality with CVD in T2D patients and determine whether inflammation prompted by poor sleep has impact on the CVD. METHODS: 332 T2D patients were recruited and their sleep qualities were evaluated by PSQI. The patients with PSQI score >7 were in the poor sleep group, and the others were in the good sleep group. Plasma samples of the patients were obtained to measure inflammatory markers. Correlation analyses and regression analyses were performed to examine the cross-sectional relationships among the poor sleep, inflammatory markers and CVD. RESULTS: The morbidity of CVD was significantly higher in the poor sleep patients compared to the good sleep patients (P=0.000). PSQI score ORs were both >1 for CVD in model 1 and model 2 (P<0.05). PSQI score were positively related to IL-6 and ICAM-1(P<0.05), negatively to FBI (P<0.05), but not related to CRP in linear regression models. Multiple logistic regression analysis showed IL-6 and ICAM-1, but not FBI and CRP, were related to CVD (P<0.05). CONCLUSIONS: Poor sleep is regarded as a plausible risk factor for CVD in T2D patients, and may be mediated by certain inflammatory markers.

The intestinal fatty acid binding protein-2 Ala54Thr polymorphism is associated with diabetic retinopathy in Chinese population.

OBJECTIVE: Endothelial dysfunction which is induced by serum saturated fatty acids increasing is one of pathogenesis of diabetic retinopathy (DR). The intestinal fatty acid binding protein-2 (FABP2) Ala54Thr polymorphism results in serum saturated fatty acids elevating. In the present study, we assessed the association of FABP2 gene polymorphism (Ala54Thr) with DR in Chinese population. MATERIALS/METHODS: In this case-control association study, 810 T2DM patients were recruited. 420 patients with retinal neovascularization, microneurysms and hemorrhages were considered as cases (DR) and 390 patients with T2DM and no clinical signs of retinopathy (DNR), were recruited as controls. Genotypes for FABP2(Ala54Thr) polymorphisms were assessed with the PCR-RFLP method. RESULTS: A significant difference in genotype distribution and allele frequency was observed between cases and controls. Patients with DR had significantly higher frequency of the Ala/Thr + Thr/Thr genotypes compared to DNR group [62.6% vs. 46.2%; OR (95% CI), 1.95 (1.48-2.59); p < 0.001]. The DR group showed a significantly higher frequency of the the Thr allele compared to the DNR group [39.5% vs. 29.4%; OR (95% CI), 1.56 (1.16-2.09); p = 0.003]. Binary logistic analyses showed FFA levels (p = 0.014) and Ala54Thr (p = 0.011) were independent correlates of the presence of DR. CONCLUSIONS: We examined that FABP2 polymophism on the Ala54Thr is significant and independent associated with DR.

Interaction of sleep quality and sleep duration on glycemic control in patients with type 2 diabetes mellitus.

BACKGROUND: Copious evidence from epidemiological and laboratory studies has revealed that sleep status is associated with glucose intolerance, insulin resistance, thus increasing the risk of developing type 2 diabetes. The aim of this study was to reveal the interaction of sleep quality and sleep quantity on glycemic control in patients with type 2 diabetes mellitus. METHODS: From May 2013 to May 2014, a total of 551 type 2 diabetes patients in Tianjin Metabolic Diseases Hospital were enrolled. Blood samples were taken to measure glycosylated hemoglobin (HbA1c), and all the patients completed the Chinese version of the Pittsburgh Sleep Quality Index (PSQI) questionnaire to evaluate their sleep status. "Good sleep quality" was defined as PQSI <5, "average sleep quality" was defined as PQSI 6-8, and "poor sleep quality" was defined as PQSI >8. Poor glycemic control was defined as HbA1c ≥7%. Sleep quantity was categorized as <6, 6-8, and >8 hours/night. Short sleep time was defined as sleep duration <6 hours/night. RESULTS: In the poor glycemic control group, the rate of patients who had insufficient sleep was much higher than that in the other group (χ(2) = 11.16, P = 0.037). The rate of poor sleep quality in poor glycemic control group was much greater than that in the average control group (χ(2) = 9.79, P = 0.007). After adjusted by gender, age, body mass index, and disease duration, the adjusted PSQI score's OR was 1.048 (95% CI 1.007-1.092, P = 0.023) for HbA1c level. The sleep duration's OR was 0.464 (95% CI 0.236-0.912, P = 0.026) for HbA1c level. One-way analysis of variance showed that the poor sleep quality group had the highest homeostasis model assessment-insulin resistance (P < 0.01). CONCLUSIONS: Inadequate sleep, in both quality and quantity, should be regarded as a plausible risk factor for glycemic control in type 2 diabetes. Poor sleep might bring much more serious insulin resistance and could be the reason for bad glycemic control. A good night's sleep should be seen as a critical health component tool in the prevention and treatment of type 2 diabetes. It is important for clinicians to target the root causes of short sleep duration and/or poor sleep quality.

Liraglutide enhances glucose transporter 4 translocation via regulation of AMP-activated protein kinase signaling pathways in mouse skeletal muscle cells.

OBJECTIVE: Liraglutide is an anti-diabetic drug and human glucagon-like peptide-1 (GLP-1) analog that primarily functions in the pancreas. However, its extra-pancreatic functions are not clear. Skeletal muscle tissue is an important determinant of blood glucose and cells take in approximately 80% of dietary glucose via glucose transporter 4 (GLUT4) on the plasma membrane. Insulin and muscle contraction are two physiological stimuli of GLUT4 translocation to the cell membrane from intracellular storage compartments, but the signaling mechanisms that mediate these processes are different. AMP-activated protein kinase (AMPK) and Akt are the key signal molecules mediating the effects of muscle contraction and insulin, respectively, on GLUT4 translocation. Here, we investigate the effect of liraglutide on GLUT4 translocation and the roles of AMPK and Akt in this mechanism in skeletal muscle cells by stably expressing GLUT4myc with an exofacial myc-epitope C(2)C(12)-GLUT4myc. MATERIALS/METHODS: The cell surface GLUT4myc levels were determined by an antibody-coupled colorimetric assay. The phosphorylation levels of AMPK, Akt, AS160, TBC1D1, and GLUT4 were determined by western blotting. The cAMP levels were measured by an ELISA kit. siRNA was transfected with Lipofectamine 2000. Analysis of variance (ANOVA) was used for data analysis. RESULTS: Liraglutide stimulated GLUT4 translocation in C(2)C(12)-GLUT4myc myotubes. Liraglutide increased the intracellular cAMP levels and the phosphorylation of AMPK, AS160, and TBC1D1. Akt phosphorylation and GLUT4 expression were not affected. Inhibition of AMPK by siRNA or Compound C reduced liraglutide-induced GLUT4 translocation. CONCLUSION: Our results suggest that liraglutide may induce GLUT4 translocation by activation of AMPK in muscle cells.