A pilot three-month sitagliptin treatment increases serum adiponectin level in Japanese patients with type 2 diabetes mellitus--a randomized controlled trial START-J study.

BACKGROUND: The dipeptidyl-peptidase-IV (DPP-4) inhibitors, including sitagliptin, are used for the treatment of type 2 diabetes mellitus (T2DM). Adiponectin, an adipocyte-derived circulating protein, has anti-atherosclerotic and anti-diabetic properties and is effectively elevated in bloodstream by thiazolidinediones, an insulin sensitizer. However, the effect of sitagliptin treatment on serum adiponectin level in T2DM has not fully elucidated in Japanese T2DM patients. The aim of the present study was to examine the effect of sitagliptin treatment on serum adiponectin levels in T2DM subjects. METHODS: Twenty-six consecutive Japanese T2DM outpatients were recruited between April 2011 and March 2013, and randomized into the control (conventional treatment, n = 10) group and sitagliptin treatment group (n = 16). Serum adiponectin was measured by enzyme-linked immunosorbent assay. RESULTS: Indices of glycemic control, such as hemoglobin A1c, glycated albumin, and 1.5-anhydro-D-glucitol, were significantly improved after the three-month treatment in both the control and sitagliptin groups. Serum adiponectin level was significantly increased in sitagliptin group from 6.7 ± 0.8 to 7.4 ± 1.0 µg/mL without change of body mass index (p = 0.034), while serum adiponectin level was not altered in the control group (p = 0.601). CONCLUSION: In Japanese T2DM patients, serum adiponectin level was elevated by three-month treatment with sitagliptin without change of body weight. TRIAL REGISTRATION: UMIN000004721.

Vascular complications and changes in body mass index in Japanese type 2 diabetic patients with abdominal obesity.

BACKGROUND: Although many Asian type 2 diabetic patients have been considered to be not obese and have low capacity of insulin secretion, the proportion of obese patients with visceral fat accumulation has increased in recent years. We found previously considerable number of Japanese non-obese subjects (body mass index (BMI) < 25 kg/m²) with visceral fat accumulation and multiple cardiovascular risk factors. The aim of the study was to investigate the difference in clinical features of type 2 diabetic patients with and without visceral fat accumulation, focusing on vascular complications and changes in BMI. METHODS: We enrolled 88 Japanese hospitalized type 2 diabetic patients. Abdominal obesity represented waist circumference (WC) of ≥85 cm for males and ≥90 cm for females (corresponding to visceral fat area of 100 cm²). Subjects were divided into two groups; with or without abdominal obesity. RESULTS: Hypertension, dyslipidemia and cardiovascular diseases were significantly more in the patients with abdominal obesity. The prevalence of cardiovascular disease in the non-obese patients (BMI < 25 kg/m²) with abdominal obesity were similar in obese patients (BMI ≥25 kg/m²). The mean BMI of the patients with abdominal obesity was < 25 kg/m² at 20 years of age, but reached maximum to more than 30 kg/m² in the course. Furthermore, substantial portion of the type 2 diabetic patients (52% in males and 43% in females) were not obese at 20 year-old (BMI < 25 kg/m²), but developed abdominal obesity by the time of admission. CONCLUSION: These results emphasize the need to control multiple risk factors and prevent atherosclerotic disease in patients with abdominal obesity. The significant weight gain after 20 years of age in patients with abdominal obesity stresses the importance of lifestyle modification in younger generation, to prevent potential development of type 2 diabetes and future atherosclerotic cardiovascular disease.

Reciprocal regulation of natriuretic peptide receptors by insulin in adipose cells.

Atrial- and brain-type natriuretic peptides (ANP and BNP, respectively) have been shown to exert potent lipolytic action in adipocytes. A family of natriuretic peptide receptors (NPRs), NPR-1, NPR-2, and NPR-3, mediates their physiologic effects. NPR-1 and NPR-2 are receptor guanylyl cyclases, while NPR-3 lacks enzymatic activity and functions primarily as a clearance receptor for natriuretic peptides. ANP has a high affinity for NPR-1 and NPR-3 than other natriuretic peptides. There is a possibility that ANP may exhibit its lipolytic effect through the balance of NPR-1 and NPR-3 expressions in adipocytes. However, the regulation of adipose NPRs has not been fully elucidated. We here examined the regulation of mouse adipose NPRs by insulin, an anti-lipolytic hormone. Among the insulin target organs, NPR-1 mRNA levels were higher in white adipose tissue (WAT) than in liver and skeletal muscle. NPR-3 mRNA was expressed most abundantly in WAT. Fasting condition induced NPR-1 mRNA level while suppressed NPR-3 mRNA level in WAT. Administration of streptozotocin resulted in the increase of NPR-1 mRNA level while the decrease of NPR-3 mRNA level in WAT. In ob/ob mice, hyperinsulinemic model, NPR-1 mRNA level was lower whereas NPR-3 mRNA level was higher compared to lean control mice. In 3T3-L1 adipocytes, insulin significantly reduced NPR-1 mRNA level while increased NPR-3 mRNA levels both through phosphatidylinositol 3-kinase (PI3-kinase) pathway. In summary, NPR-1 and NPR-3 were highly expressed in WAT and adipose NPR-1 and NPR-3 were reciprocally regulated by insulin. This study suggests that insulin may efficiently promote lipogenesis partly by reducing the lipolytic action of ANP through the opposite regulation of NPR-1 and NPR-3.

Adiponectin protects against angiotensin II-induced cardiac fibrosis through activation of PPAR-alpha.

OBJECTIVE: Adiponectin is recognized as an antidiabetic, antiatherosclerotic, and anti-inflammatory protein derived from adipocytes. However, the role of adiponectin in cardiac fibrosis remains uncertain. We herein explore the effects of adiponectin on cardiac fibrosis induced by angiotensin II (Ang II). METHODS AND RESULTS: Wild-type (WT), adiponectin knockout (Adipo-KO), and PPAR-alpha knockout (PPAR-alpha-KO) mice were infused with Ang II at 1.2 mg/kg/d. Severe cardiac fibrosis and left ventricular dysfunction were observed in Ang II-infused Adipo-KO mice compared to WT mice. Adenovirus-mediated adiponectin treatment improved the above phenotypes and the dysregulation of reactive oxygen species (ROS)-related mRNAs in Adipo-KO mice, whereas such amelioration was not observed in PPAR-alpha-KO mice despite adiponectin accumulation in heart tissue. In cultured cardiac fibroblasts, adiponectin improved the reduction of AMP-activated protein kinase (AMPK) activity and elevation of extracellular signal-regulated kinase 1/2 (ERK1/2) activity induced by Ang II. Adiponectin significantly enhanced PPAR-alpha activity, whereas the adiponectin-dependent PPAR-alpha activation was diminished by Compound C, an inhibitor of AMPK. CONCLUSIONS: The present study suggests that adiponectin protects against Ang II-induced cardiac fibrosis possibly through AMPK-dependent PPAR-alpha activation.

Role of aquaporin-7 and aquaporin-9 in glycerol metabolism; involvement in obesity.

The discovery of aquaporin (AQP) has had a great impact on life sciences. So far, 13 AQPs have been identified in human. AQP3, 7, 9, and 10 are subcategorized as aquaglyceroporins which permeabilize glycerol as well as water. Many investigators have demonstrated that AQPs play a crucial role in maintaining water homeostasis, but the physiological significance of some AQPs as a glycerol channel has not been fully understood. Adipocyte is considered to be a major source of glycerol which is one of substrates for hepatic gluconeogenesis. This review focuses on recent studies of glycerol metabolism through AQP7 and AQP9, and discusses the importance of glycerol channel in adipose tissues and liver.

URB is abundantly expressed in adipose tissue and dysregulated in obesity.

Dysregulated production of adipocytokines in obesity is involved in the development of metabolic syndrome. URB/DRO1 contains N-terminal signal sequence and is thought to play a role in apoptosis of tumor cells. In the present study, we investigated the expression pattern of URB mRNA in adipose tissue and secretion from cultured adipocytes. In human and mouse, URB mRNA was predominantly expressed in adipose tissue and was downregulated in obese mouse models, such as ob/ob, KKAy, and diet-induced obese mice. In 3T3L1 adipocytes, insulin, TNF-alpha, H(2)O(2) and hypoxia decreased URB mRNA level. This regulation was similar to that for adiponectin and opposite to MCP-1. URB protein was secreted in media of URB cDNA-stably transfected cells and endogenous URB was detected in media of cultured human adipocytes. In conclusion, the expression pattern of URB suggests its role in obesity and the results suggest that URB is secreted, at least in part, from adipocytes.

Effects of peroxisome proliferator-activated receptor ligands, bezafibrate and fenofibrate, on adiponectin level.

OBJECTIVE: Adiponectin is adipose-specific secretory protein and acts as anti-diabetic and anti-atherosclerotic molecule. We previously found peroxisome proliferators response element in adiponectin promoter region, suggesting that peroxisome proliferator-activated receptor (PPAR) ligands elevate adiponectin. Fibrates are known to be PPARalpha ligands and were shown to reduce risks of diabetes and cardiovascular disease. Effect of fibrates on adiponectin has not been clarified, whereas thiazolidinediones enhance adiponectin. Thus, we explored the possibility and mechanism that fibrates enhance adiponectin in humans, mice, and cells. METHODS AND RESULTS: Significant increase of serum adiponectin was observed in bezafibrate-treated subjects compared with placebo group in patients enrolled in The Bezafibrate Infarction Prevention study. Higher baseline adiponectin levels were strongly associated with reduced risk of new diabetes. Fibrates, bezafibrate and fenofibrate, significantly elevated adiponectin levels in wild-type mice and 3T3-L1 adipocytes. Such an effect was not observed in PPARalpha-deficient mice and adipocytes. Fibrates activated adiponectin promoter but failed to enhance its activity when the point mutation occurred in peroxisome proliferators response element site and the endogenous PPARalpha was knocked down by PPARalpha-RNAi. CONCLUSIONS: Our results suggest that fibrates enhance adiponectin partly through adipose PPARalpha and measurement of adiponectin might be a useful tool for searching subjects at high risk for diabetes.

Aquaporins and glycerol metabolism.

The discovery of aquaporin (AQP) has made a great impact on life sciences. AQPs are a family of homologous water channels widely distributed in plants, unicellular organisms, invertebrates, and vertebrates. So far, 13 AQPs have been identified in human. AQP3, 7, 9, and 10 are subcategorized as aquaglyceroporins which permeabilize glycerol as well as water. Many investigators have demonstrated that AQPs play a crucial role in maintaining water homeostasis, but the physiological significance of some AQPs as a glycerol channel is not fully understood. Adipose tissue is a major source of glycerol and glycerol is one of substrates for gluconeogenesis. This review focuses on recent studies of glycerol metabolism through aquaglyceroporins, and briefly discusses the importance of glycerol channel in adipose tissues and liver.

The heart requires glycerol as an energy substrate through aquaporin 7, a glycerol facilitator.

AIMS: Cardiomyocytes require fatty acids and glucose for energy production. However, other nutrients and substrates that may serve as possible candidates for a cardiac energy source have not been fully studied. Several reports showed that a moderate expression of aquaporin 7 (AQP7), a member of the aquaglyceroporin family that is permeated by glycerol and water, is observed in heart tissue. However, the functional role of cardiac AQP7 is not clear. The aim of this study was to investigate the significance of glycerol as a cardiac energy substrate and to clarify the role of cardiac AQP7. METHODS AND RESULTS: Heart function and morphology were examined in AQP7-knockout (KO) mice under basal conditions and during pressure overload [isoproterenol infusion and transverse aortic constriction (TAC)]. Glycerol uptake and glycerol-dependent ATP production were measured in AQP7-knockdown cardiac cells. Cardiac glycerol consumption was analysed in ex vivo beating hearts. Cardiac morphology and function in KO mice were similar to those of wild-type (WT) mice under basal conditions, although low glycerol and ATP content were noted in hearts of KO mice. In H9c2 cardiomyotubes, knockdown of AQP7 was associated with a significant reduction of glycerol uptake. The ex vivo heart study demonstrated that cardiac glycerol consumption levels in KO mice were significantly lower than those of WT mice. Furthermore, isoproterenol challenge induced severe left ventricular hypertrophy in KO mice, and TAC resulted in a higher mortality rate in KO mice than in WT mice. CONCLUSION: The results indicate that AQP7 acts as a glycerol facilitator in cardiomyocytes and that glycerol is a substrate for cardiac energy production.

Blockade of mineralocorticoid receptor reverses adipocyte dysfunction and insulin resistance in obese mice.

AIMS: In obesity, chronic low-grade inflammation and overproduction of reactive oxygen species (ROS) in fat contribute to the development of metabolic syndrome. Suppression of inflammation and ROS production in fat may attenuate the metabolic syndrome. Activation of mineralocorticoid receptor (MR) promotes inflammation in heart, kidney, and vasculature via ROS generation. However, the significance of MR in fat remains elusive. Here we investigated whether MR blockade attenuates obesity-related insulin resistance and improves adipocyte dysfunction. METHODS AND RESULTS: Obese ob/ob and db/db mice were treated with eplerenone, a MR antagonist, for 3 weeks. 3T3-L1 adipocytes were treated with aldosterone or H2O2, with and without eplerenone or MR-siRNA. High levels of MR mRNA were detected in adipose tissue of obese ob/ob and db/db mice. Eplerenone treatment significantly reduced insulin resistance, suppressed macrophage infiltration and ROS production in adipose tissues, and corrected the mRNA levels of obesity-related genes in obese mice. In 3T3-L1 adipocytes, aldosterone and H2O2 increased intracellular ROS levels and MR blockade inhibited such increases. H2O2 and aldosterone resulted in dysregulation of mRNAs of various genes related to ROS and cytokines, whereas MR blockade corrected such changes. CONCLUSION: MR blockade attenuates obesity-related insulin resistance partly through reduction of fat ROS production, inflammatory process, and induction of cytokines.

Adiponectin replenishment ameliorates obesity-related hypertension.

Patients with obesity are susceptible to hypertension. We have reported that the plasma adiponectin levels are decreased in obesity and that adiponectin has many defensive properties against obesity-related diseases, such as type 2 diabetes and coronary artery disease. The aim of this study was to determine the relationship between adiponectin and hypertension in mice. We measured blood pressure and heart rate directly by a catheter in the carotid artery and indirectly by automatic sphygmomanometer at the tail artery. Obese KKAy mice had significantly lower plasma adiponectin levels and higher systolic blood pressure than control C57BL/6J mice at 21 weeks of age. Adenovirus-delivered adiponectin significantly decreased blood pressure in KKAy mice. The direct role of adiponectin on blood pressure regulation under insulin resistance-free state was investigated in adiponectin-knockout (KO) mice. Adiponectin KO mice developed hypertension when maintained on a high-salt diet (8% NaCl) without insulin resistance. The hypertension of salt-fed adiponectin KO mice was associated with reduced mRNA levels of endothelial NO synthase (eNOS) and prostaglandin I(2) synthase in aorta and low metabolite levels of endothelial NO synthase and prostaglandin I(2) synthase in plasma. Adiponectin therapy lowered the elevated blood pressure and corrected the above mRNA levels to those of the wild type. Our results suggest that hypoadiponectinemia contributes to the development of obesity-related hypertension, at least in part, directly, in addition to its effect via insulin resistance, and that adiponectin therapy can be potentially useful for hypertension in patients with the metabolic syndrome.

Aquaporin 7 deficiency is associated with development of obesity through activation of adipose glycerol kinase.

In adipocytes, hydrolysis of triglycerides results in the release of free fatty acids and glycerol. Aquaporin 7 (AQP7), a member of aquaglyceroporins, is known to permeabilize glycerol and water. We recently generated Aqp7-knockout (KO) mice and demonstrated that such mice have low plasma glycerol levels and impaired glycerol release in response to beta3-adrenergic agonist, suggesting that AQP7 acts as a glycerol gateway molecule in adipocytes for the efficient release of glycerol in vivo. Although there was no difference in body weight between WT and KO mice until 10 weeks of age, here we found that KO mice developed adult-onset obesity. The body weight and fat mass increased significantly in KO mice compared with WT mice after 12 weeks of age. Adipocytes of KO mice were large and exhibited accumulation of triglycerides compared with WT mice. The KO mice developed obesity and insulin resistance even at a young age after consumption of high-fat/high-sucrose diet. We demonstrated the enhanced glycerol kinase enzymatic activity in Aqp7-KO and -knockdown adipocytes. A series of our results indicate that AQP7 disruption elevates adipose glycerol kinase activity, accelerates triglycerides synthesis in adipocytes, and, finally, develops obesity.

Adaptation to fasting by glycerol transport through aquaporin 7 in adipose tissue.

Adipocytes hydrolyze triglycerides and secrete free fatty acids and glycerol into the circulation. The molecular mechanism involved in glycerol transport from adipocytes has not been elucidated. Here, we investigated glycerol and glucose metabolism in mice lacking aquaporin 7 (Aqp7), a member of the aquaglyceroporins expressed in adipose tissue, and demonstrated that Aqp7 functions as a glycerol gateway molecule in vivo. Aqp7-knockout (KO) mice had lower plasma glycerol levels compared with WT mice but had normal plasma free fatty acid levels. The increase in plasma glycerol level in response to beta(3)-adrenergic agonist was severely impaired in KO mice. Epinephrine-stimulated glycerol secretion was also impaired in Aqp7 knockdown adipocytes. During prolonged fasting, plasma glycerol was elevated and the plasma glucose level was maintained in WT mice. In contrast, KO mice showed a disrupted increase of plasma glycerol and rapid reduction of plasma glucose during prolonged fasting. Our findings indicate that the lack of effective glycerol transport from adipocytes by glycerol gateway molecule causes defective adaptation to prolonged fasting.

Measurement of fasting serum apoB-48 levels in normolipidemic and hyperlipidemic subjects by ELISA.

The present study was designed to evaluate the metabolism of chylomicron and chylomicron remnants by measuring serum apolipoprotein B-48 (apoB-48) levels in 335 normolipidemic and 253 hyperlipidemic subjects using a novel ELISA system. The distribution of fasting serum apoB-48 levels in normolipidemic subjects varied widely, ranging from <1 to >24 microgram/ml (mean, 5.2 +/- 3.8 microgram/ml; median, 3.9 microgram/ml). Serum apoB-48 levels correlated with serum triglyceride (TG) concentrations (r = 0.45, P < 0.001), but not with total cholesterol levels. Serum apoB-48 levels were 7 to 18 times higher in patients with Type I, Type V, and Type III hyperlipidemia, and only slightly higher in patients with Type IIa, Type IIb, and Type IV hyperlipidemia, compared with normolipidemic subjects. The calculated apoB-48/TG ratio was elevated only in patients with dysbetalipoproteinemia (apoE2/2 phenotype). In normolipidemic subjects, oral fat loading resulted in about 2-fold increase in serum apoB-48 levels, with a peak level recorded at 3-4 h postloading, and then returned to the baseline level within 6 h. On the other hand, in patients with dysbetalipoproteinemia, serum apoB-48 levels did not change considerably. Our results indicate that serum apoB-48 is a very useful parameter for evaluating lipoprotein metabolism in exogenous pathways.

Disturbed secretion of mutant adiponectin associated with the metabolic syndrome.

Adiponectin, an adipocyte-derived protein, consists of collagen-like fibrous and complement C1q-like globular domains, and circulates in human plasma in a multimeric form. The protein exhibits anti-diabetic and anti-atherogenic activities. However, adiponectin plasma concentrations are low in obese subjects, and hypoadiponectinemia is associated with the metabolic syndrome, which is a cluster of insulin resistance, type 2 diabetes mellitus, hypertension, and dyslipidemia. We have recently reported a missense mutation in the adiponectin gene, in which isoleucine at position 164 in the globular domain is substituted with threonine (I164T). Subjects with this mutation showed markedly low level of plasma adiponectin and clinical features of the metabolic syndrome. Here, we examined the molecular characteristics of the mutant protein associated with a genetic cause of hypoadiponectinemia. The current study revealed (1) the mutant protein showed an oligomerization state similar to the wild-type as determined by gel filtration chromatography and, (2) the mutant protein exhibited normal insulin-sensitizing activity, but (3) pulse-chase study showed abnormal secretion of the mutant protein from adipose tissues. Our results suggest that I164T mutation is associated with hypoadiponectinemia through disturbed secretion into plasma, which may contribute to the development of the metabolic syndrome.

Divergent effects of soy protein diet on the expression of adipocytokines.

Adipose tissue secretes various bioactive molecules called adipocytokines. Dysregulation of adipocytokines plays an important role in the development of atherosclerotic vascular diseases. Consumption of vegetable protein reduces the risks of atherosclerotic vascular diseases. Here, we investigated the effects of 10-day dietary soy protein isolate (SPI) on the regulation of adipocytokines in Wistar rats. No significant difference in body weight was observed between SPI and Casein (animal protein) group. Expression of adipose PAI-1 was lower and expression and plasma concentration of adiponectin were higher in SPI than Casein group. Triglyceride content was lower and fatty acid synthase mRNA level in adipose tissue was lower in SPI than Casein group. Although SREBP-1 mRNA expression was decreased in the liver of SPI group, adipose SREBP and PPARgamma mRNA levels remained unchanged. Our data suggest that dietary SPI alters the gene expressions in adipose tissue and has beneficial effects on the expression of adipocytokines.