Xinguo Hou

Metformin reduces intracellular reactive oxygen species levels by upregulating expression of the antioxidant thioredoxin via the AMPK-FOXO3 pathway

BACKGROUND Oxidative stress induced by free fatty acids plays a critical role in the pathogenesis of endothelial dysfunction and atherosclerosis in patients with metabolic syndrome. Reducing oxidative stress in these patients may prevent cardiovascular complications. The antidiabetic agent metformin has been reported to directly protect the cardiovascular system. In this study, we examined the effect of metformin on the intracellular levels of reactive oxygen species (ROS) induced by palmitic acid (PA) in human aortic endothelial cells and studied the molecular mechanisms involved. METHODS AND RESULTS We observed that metformin significantly reduced intracellular ROS levels induced by PA. Additionally, metformin increased the expression of the antioxidant thioredoxin (Trx), which mediated metformins effects on ROS reduction. Metformin increased Trx expression through the AMP-activated protein kinase (AMPK) pathway. Metformin-regulated Trx at the transcriptional level and forkhead transcription factor 3 (FOXO3) was involved in this process. CONCLUSION These results suggest that metformin reduces ROS levels by inducing Trx expression through activation of the AMPK-FOXO3 pathway.

Elevated peripheral frequencies of Th22 cells: a novel potent participant in obesity and type 2 diabetes.

Objective Chronic low-grade inflammation has long been recognized as the central link between obesity and type 2 diabetes (T2D). The novel subset of T helper (Th) cells, Th22, plays an emerging role in chronic inflammation. We investigated the potential association between Th22 and the pathogenesis of obesity and T2D. Methods Ninety T2D inpatients (T2D group), 30 healthy participants with BMI ranged from 19 to 23.9 kg/m2 (CTL group) and 30 metabolically healthy obese controls with BMI ≥ 30 kg/m2 (MHO group) were employed in our study. Peripheral frequencies of Th22 and Th1 and Th17 cells were determined by flow cytometry based on their specific cytokine patterns. Cytokine levels in fresh plasma were quantified by ELISA. Results Compared to that in CTL group (1.18±0.06%, n = 28), peripheral frequency of Th22 cells was significantly increased in MHO group (1.88±0.10%, n = 30) and in T2D group (2.247±0.10%, n = 89). There was a consistent notable increase in plasma interleukin (IL)-22 of T2D patients [47.56 (30.55–76.89) pg/mL] as compared with that of MHO group [36.65 (29.52–55.70) pg/ml; *P<0.0001] and CTLs [36.33 (31.93–40.62) pg/mL; *P<0.0001]. Furthermore, other than Th1/Th17, previously frequently described participants in obesity and T2D, there was a strong correlation between Th22 frequency and the homeostasis model of assessment for insulin resistance index (r = 0.6771, *P<0.0001) and HOMA for β-cell function (r = −0.7264, *P<0.0001). Conclusions There were increased Th22 frequencies and IL-22 levels in obesity and T2D. Elevated Th22 and IL-22 also aided in the differentiation of MHO from T2D patients. The notable correlation implied that Th22 might play a more determinant role in both insulin resistance and β-cell impairment.

Glucagon-like peptide 1 regulates adipogenesis in 3T3-L1 preadipocytes

Glucagon-like peptide 1 (GLP-1), a gut-derived peptide, has been reported to have profound effects on metabolism and to reduce insulin resistance. Adipocyte hyperplasia stimulated by preadipocyte differentiation has a positive effect on adipose tissue insulin sensitivity. However, it remains less clear whether GLP-1 plays a role in adipogenesis. In this study, we examined the effect of GLP-1 on preadipocyte differentiation and investigated the mechanisms that may be involved in this effect. In our 3T3-L1 cell study, we tested the levels of adipocyte-specific markers and signaling pathways during preadipocyte differentiation. In addition, Oil Red O staining was used to examine lipid accumulation. Image Pro Plus 5.02 was used to analyze the size and number of lipid droplets. We found that GLP-1 elevated the protein expression levels of free fatty acid-binding protein 4 (aP2) and the transcription factor peroxisome proliferator-activated receptor-γ (PPAR‑γ) in a dose-dependent manner during 3T3‑L1 preadipocyte differentiation. Furthermore, RT‑PCR results showed that GLP-1 promoted CCAAT/enhancer-binding protein α (C/EBPα) and lipoprotein lipase (LPL) expression at the transcriptional level. These data suggest that GLP-1 promotes preadipocyte differentiation. Our study also found that treatment of the cells with 100 nM GLP-1 enhanced the phosphorylation of Akt signaling during the first 24 h of differentiation. Although Oil Red O staining showed that GLP‑1 had no significant effect on lipid accumulation, there were increased numbers of small adipocytes in the cells treated with 100 nM GLP‑1. Taken together, these results indicate that GLP-1 regulates 3T3‑L1 adipogenesis and the Akt signaling pathway may be involved in this process. The differentiated small adipocytes may have a positive effect against insulin resistance and obesity.

Glucagon-like peptide 1 protects microvascular endothelial cells by inactivating the PARP-1/iNOS/NO pathway.

Increasing studies suggest that the activity of GLP-1 might be of significant importance in the development of type 2 diabetes beyond its serum glucose-lowering effects. However, to date, the anti-apoptosis mechanism by which GLP-1 acts on MILE SVEN 1 (MS-1) cells has not been fully explored with regard to the intracellular signaling pathway. Increasing evidence shows that apoptosis of islet microvascular endothelial cells (IMECs) participates in the pathogenesis of diabetes. We wondered whether GLP-1 exerts its anti-apoptosis effects by inactivating the PARP-1/iNOS/NO pathway in oxidized low-density-lipoprotein (oxLDL)-induced apoptosis in mouse IMECs (MS-1 cells), which may linked to GLP-1R/cAMP levels. MTT assay revealed that 2-h pre-incubation with GLP-1 markedly restored the oxLDL-induced loss of MS-1 viability in a concentration-dependent manner. This effect was accompanied by a significant decrease in intracellular nitric oxide (NO) activity. Moreover, GLP-1 suppressed lipid peroxidation, restored the activities of endogenous antioxidants, and decreased the level of NO. Pre-incubating MS-1 cells with GLP-1 reduced cell apoptosis. Finally, GLP-1 could efficiently prevent the upregulation of poly(ADP-ribose) polymerase-1/nitrotyrosine and inducible NO synthase protein. Simultaneously, the expression of GLP-1 receptor and the level of cAMP was consistent with the administration of GLP-1. Our findings suggest that GLP-1 can effectively protect MS-1 cells against oxLDL-induced apoptosis, which may be important in preventing the pathogenesis of diabetes mellitus.

Hyperglycemia induces apoptosis of pancreatic islet endothelial cells via reactive nitrogen species-mediated Jun N-terminal kinase activation

Hyperglycemia significantly stimulates pancreatic islet endothelial cell apoptosis; however, the precise mechanisms are not fully understood. In the present study, treating pancreatic islet endothelial (MS-1) cells with high glucose (30mmol/l) but not mannitol significantly increased the number of apoptotic cells as compared with a physiological glucose concentration (5.5mmol/l). Hyperglycemia significantly stimulated the expression of inducible nitric oxide synthase (iNOS) and production of NO and peroxynitrite (ONOO(-)), relevant to MS-1 cell apoptosis. Moreover, induced reactive nitrogen species (RNS) significantly increased the expression of bax, cleaved caspase-3 and poly adenosine diphosphate (ADP)-ribose polymerase (PARP) via JNK activation, but the expression of bcl-2 was not altered. Furthermore, SP600125 (a specific inhibitor of JNK) and 1400W (a specific inhibitor of iNOS) significantly attenuated cell apoptosis induced by high glucose. Therefore, hyperglycemia triggers MS-1 cell apoptosis by activating an intrinsic-dependent apoptotic pathway via RNS-mediated JNK activation.

Circulating Levels of Betatrophin and Irisin Are Not Associated with Pancreatic β-Cell Function in Previously Diagnosed Type 2 Diabetes Mellitus Patients

Betatrophin and irisin are two recently identified hormones which may participate in regulating pancreatic β-cell function. However, the associations of these two hormones with β-cell function remain unclear. The present study aims to demonstrate the associations of circulating betatrophin and irisin levels with β-cell function, assessed by the area under the curve (AUC) of C-peptide, and the possible correlation between these two hormones in previously diagnosed type 2 diabetes mellitus (T2DM) patients. In total, 20 age-, sex-, and body mass index- (BMI-) matched normal glucose tolerance (NGT) subjects and 120 previously diagnosed T2DM patients were included in this study. Partial correlation analysis was used to evaluate the relationships between these two hormones and indexes of β-cell function and insulin resistance. Our results showed that betatrophin levels were significantly elevated, while irisin levels were significantly decreased, in patients with T2DM compared with NGT subjects. However, partial correlation analysis showed that betatrophin levels did not correlate with β-cell function-related variables or insulin resistance-related variables before or after controlling multiple covariates, while irisin correlated positively with insulin sensitivity but is not associated with β-cell function-related variables. Besides, no correlation was observed between betatrophin and irisin levels. Hence we concluded that betatrophin and irisin were not associated with β-cell function in previously diagnosed T2DM patients.

Irisin has no effect on lipolysis in 3T3-L1 adipocytes or fatty acid metabolism in HepG2 hepatocytes

Irisin, a newly identified myokine responsible for browning of white or beige adipocytes, has been reported to be present at reduced levels in diabetic patients and associated with obesity, serum triglyceride (TG) levels, and intrahepatic TG levels. We wondered whether irisin could directly affect fatty acid and TG metabolism in adipocytes and hepatocytes. We examined the effects of various concentrations of irisin on lipolysis (according to Oil Red O staining, free fatty acid release, and glycerol release), protein expression of HSL and ATGL, and mRNA expression of other lipid-related genes (UCP-1, PPARγ, FABP-4, HSL, ATGL, PPARα, and CPT-1) in mature 3T3-L1 adipocytes, as well as mRNA levels of genes involved in the synthesis (SREBP-1C and FAS) and β-oxidation (PPARα and CPT-1) of fatty acids in HepG2 hepatocytes under physiological or hyperglycemic conditions. Our results revealed that although irisin significantly increased the mRNA levels of UCP-1 and PPARα, it failed to show detectable effects on lipolysis, HSL or ATGL protein levels, or the mRNA expression of other lipid-related genes in mature 3T3-L1 adipocytes. In HepG2 hepatocytes, high glucose induced the upregulation of SREBP-1C and FAS and the downregulation of PPARα; however, no significant effect of irisin on gene expression was observed under either physiological or hyperglycemic conditions. We therefore conclude that irisin has no significant direct effect on lipolysis in 3T3-L1 adipocytes or on fatty acid metabolism in HepG2 hepatocytes.

C-Peptide Is Independently Associated with an Increased Risk of Coronary Artery Disease in T2DM Subjects: A Cross-Sectional Study

Objective C-peptide has been reported to be a marker of subclinical atherosclerosis in type 2 diabetes mellitus (T2DM) patients, whereas its role in coronary artery disease (CAD) has not been clarified, especially in diabetics with differing body mass indices (BMIs). Design and Methods This cross-sectional study included 501 patients with T2DM. First, all subjects were divided into the following two groups: CAD and non-CAD. Then, binary logistic regression was used to determine the risk factors for CAD for all patients. To clarify the role of obesity, we re-divided all subjects into two additional groups (obese and non-obese) based on BMI. Finally, binary logistic regression was used to determine the risk factors for CAD for each weight group. Results The patients with CAD showed a higher BMI and fasting C-peptide level in addition to an increased prevalence of traditional risk factors for CAD, such as hypertension, insulin resistance, higher cholesterol, cysteine-C (Cys-C) and lower estimated glomerular filtration rate (eGFR). Logistic regression analysis showed that fasting C-peptide (OR=1.513, p=0.005), insulin treatment (OR=1.832, p=0.027) hypertension (OR=1.987, p=0.016) and hyperlipidemia (OR=4.159, p<0.001) significantly increased the risk of clinical CAD in the T2DM patients independent of age, gender, diabetes duration, smoking and alcohol statuses, fasting insulin and glucose, hypoglycemic episodes, UA and eGFR. Additionally, in both of the obese (OR=1.488, p=0.049) and non-obese (OR=1.686, p=0.037) DM groups, C-peptide was associated with an increased risk of CAD after multiple adjustments. Conclusions C-peptide is associated with an increased CAD risk in T2DM patients, no matter whether they are obese or not.

Triglyceride levels are closely associated with mild declines in estimated glomerular filtration rates in middle-aged and elderly Chinese with normal serum lipid levels.

Objective To investigate the relationship between lipid profiles [including total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C)] and a mild decline in the estimated glomerular filtration rate (eGFR) in subjects with normal serum lipid levels. Design and Methods In this study, we included 2647 participants who were ≥40 years old and had normal serum lipid levels. The Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equation was used to estimate the GFR. A mildly reduced eGFR was defined as 60–90 mL/min/1.73 m2. First, multiple linear regression analysis was used to estimate the association of lipid profiles with the eGFR. Then, the levels of each lipid component were divided into four groups, using the 25th, 50th and 75th percentiles as cut-off points. Finally, multiple logistic regression analysis was used to investigate the association of different lipid components with the risk of mildly reduced eGFR. Results In the group with a mildly reduced eGFR, TG and LDL-C levels were significantly increased, but HDL-C levels were significantly decreased. After adjusting for age, gender, body mass index (BMI), systolic blood pressure (SBP), glycated hemoglobin (HbA1c), smoking and drinking, only TC and TG were independently related to the eGFR. Additionally, only TG showed a linear relationship with an increased risk of a mildly reduced eGFR, with the highest quartile group (TG: 108–150 mg/dl [1.22–1.70 mmol/L]) having a significantly increased risk after adjusting for the above factors. Conclusions Triglyceride levels are closely associated with a mildly reduced eGFR in subjects with normal serum lipid levels. Dyslipidemia with lower TG levels could be used as new diagnostic criteria for subjects with mildly reduced renal function.

Association of the Number of Years Since Menopause with Metabolic Syndrome and Insulin Resistance in Chinese Urban Women

BACKGROUND This study aimed to assess the prevalence of metabolic syndrome (MetS) and the association of years since menopause with MetS and Insulin Resistance (IR) in Chinese women. METHOD A total of 4436 Chinese subjects aged 40-80 years participated in the study; 790 were premenopausal women, and 3646 were postmenopausal women. IR was arbitrarily defined as a homeostasis model assessment-IR index (HOMA-IR) value above the 75th percentile of normal glucose tolerance (NGT). MetS was defined according to the International Diabetes Federation consensus definition. To test whether there was an association between the number of years since menopause and MetS, multivariate logistic analysis was conducted. Premenopausal women were used as a comparison group in regression analyses. RESULTS After adjustment for age, body mass index (BMI), and γ-glutamyltransferase (GGT), more years since menopause was highly associated with an increased risk of MetS (p for trend <0.05) ; the number of years since menopause was not correlated with fasting insulin and HOMA-IR. Postmenopausal women with 10 to 14 years since menopause had the highest risk (odds ratio [OR], 2.10; 95% confidence interval [CI], 1.52-2.89, p < .05) of MetS, high triglycerides (TG; OR, 1.80; 95% CI, 1.34-2.42, p < .05) and high glucose (OR, 1.52; 95% CI, 1.14-2.05, p < .05) and low high-density lipoprotein cholesterol (HDL-C; OR, 1.38; 95% CI, 1.18-2.32, p < .05). Postmenopausal women with more than 15 years since menopause had the highest risk of abdominal obesity (OR, 1.69; 95% CI, 1.05-2.71, p < .05). CONCLUSION In China, more years since menopause was highly associated with an increased risk of MetS. Menopausal history may help identify women with increased risk of developing MetS.