The integrated incretin effect is reduced by both glucose intolerance and obesity in Japanese subjects.

Introduction: Incretin-based drugs are extensively utilized in the treatment of type 2 diabetes (T2D), with remarkable clinical efficacy. These drugs were developed based on findings that the incretin effect is reduced in T2D. The incretin effect in East Asians, whose pancreatic ß-cell function is more vulnerable than that in Caucasians, however, has not been fully examined. In this study, we investigated the effects of incretin in Japanese subjects. Methods: A total of 28 Japanese subjects (14 with normal glucose tolerance [NGT], 6 with impaired glucose tolerance, and 8 with T2D) were enrolled. Isoglycemic oral (75 g glucose tolerance test) and intravenous glucose were administered. The numerical incretin effect and gastrointestinally-mediated glucose disposal (GIGD) were calculated by measuring the plasma glucose and entero-pancreatic hormone concentrations. Results and discussion: The difference in the numerical incretin effect among the groups was relatively small. The numerical incretin effect significantly negatively correlated with the body mass index (BMI). GIGD was significantly lower in participants with T2D than in those with NGT, and significantly negatively correlated with the area under the curve (AUC)-glucose, BMI, and AUC-glucagon. Incretin concentrations did not differ significantly among the groups. We demonstrate that in Japanese subjects, obesity has a greater effect than glucose tolerance on the numerical incretin effect, whereas GIGD is diminished in individuals with both glucose intolerance and obesity. These findings indicate variances as well as commonalities between East Asians and Caucasians in the manifestation of incretin effects on pancreatic ß-cell function and the integrated capacity to handle glucose.

Intestinal Morphology and Glucose Transporter Gene Expression under a Chronic Intake of High Sucrose.

Sucrose is a disaccharide that is degraded into fructose and glucose in the small intestine. High-sucrose and high-fructose diets have been reported, using two-dimensional imaging, to alter the intestinal morphology and the expression of genes associated with sugar transport, such as sodium glucose co-transporter 1 (SGLT1), glucose transporter 2 (GLUT2), and glucose transporter 5 (GLUT5). However, it remains unclear how high-fructose and high-sucrose diets affect the expression of sugar transporters and the intestinal morphology in the whole intestine. We investigate the influence of a chronic high-sucrose diet on the expression of the genes associated with sugar transport as well as its effects on the intestinal morphology using 3D imaging. High sucrose was found to increase GLUT2 and GLUT5 mRNA levels without significant changes in the intestinal morphology using 3D imaging. On the other hand, the delay in sucrose absorption by an α-glucosidase inhibitor significantly improved the intestinal morphology and the expression levels of SGLT1, GLUT2, and GLUT5 mRNA in the distal small intestine to levels similar to those in the proximal small intestine, thereby improving glycemic control after both glucose and sucrose loading. These results reveal the effects of chronic high-sugar exposure on glucose absorption and changes in the intestinal morphology.

Inhibition of GPR120 signaling in intestine ameliorates insulin resistance and fatty liver under high-fat diet feeding.

G protein-coupled receptor (GPR) 120 is expressed in enteroendocrine cells secreting glucagon-like peptide-1 (GLP-1), glucose-dependent insulinotropic polypeptide/gastric inhibitory polypeptide (GIP), and cholecystokinin (CCK). Although GPR120 signaling in adipose tissue and macrophages has been reported to ameliorate obesity and insulin resistance in a high long-chain triglyceride (LCT) diet, intestine-specific roles of GPR120 are unclear. To clarify the metabolic effect of GPR120 in the intestine, we generated intestine-specific GPR120-knockout (GPR120int-/-) mice. In comparison with floxed GPR120 (WT) mice, GPR120int-/- mice exhibited reduced GIP secretion and CCK action without change of insulin, GLP-1, or peptide YY (PYY) secretion after a single administration of LCT. Under a high-LCT diet, GPR120int-/- mice showed a mild reduction of body weight and substantial amelioration of insulin resistance and fatty liver. Moreover, liver and white adipose tissue (WAT) of GPR120int-/-mice exhibited increased Akt phosphorylation and reduced gene expression of suppressor of cytokine signaling (SOCS) 3, which inhibits insulin signaling. In addition, gene expression of inflammatory cytokines in WAT and lipogenic molecules in liver were reduced in GPR120int-/- mice. These findings suggest that inhibition of GPR120 signaling in intestine ameliorates insulin resistance and fatty liver under high-LCT diet feeding.NEW & NOTEWORTHY We generated novel intestine-specific GPR120-knockout (GPR120int-/-) mice and investigated the metabolic effect of GPR120 in the intestine. GPR120int-/- mice exhibited a reduction of GIP secretion and CCK action after a single administration of LCT. Under a high-LCT diet, GPR120int-/- mice showed mild improvement in obesity and marked amelioration of insulin resistance and hepatic steatosis. Our results indicate an important role of intestinal GPR120 on insulin resistance and hepatic steatosis.

An analysis of intestinal morphology and incretin-producing cells using tissue optical clearing and 3-D imaging.

Tissue optical clearing permits detailed evaluation of organ three-dimensional (3-D) structure as well as that of individual cells by tissue staining and autofluorescence. In this study, we evaluated intestinal morphology, intestinal epithelial cells (IECs), and enteroendocrine cells, such as incretin-producing cells, in reporter mice by intestinal 3-D imaging. 3-D intestinal imaging of reporter mice using optical tissue clearing enabled us to evaluate both detailed intestinal morphologies and cell numbers, villus length and crypt depth in the same samples. In disease mouse model of lipopolysaccharide (LPS)-injected mice, the results of 3-D imaging using tissue optical clearing in this study was consistent with those of 2-D imaging in previous reports and could added the new data of intestinal morphology. In analysis of incretin-producing cells of reporter mice, we could elucidate the number, the percentage, and the localization of incretin-producing cells in intestine and the difference of those between L cells and K cells. Thus, we established a novel method of intestinal analysis using tissue optical clearing and 3-D imaging. 3-D evaluation of intestine enabled us to clarify not only detailed intestinal morphology but also the precise number and localization of IECs and incretin-producing cells in the same samples.

Noninvasive Evaluation of GIP Effects on ß-Cell Mass Under High-Fat Diet.

Pancreatic ß-cell mass (BCM) has an importance in the pathophysiology of diabetes mellitus. Recently, glucagon-like peptide-1 receptor (GLP-1R)-targeted imaging has emerged as a promising tool for BCM evaluation. While glucose-dependent insulinotropic polypeptide/gastric inhibitory polypeptide (GIP) is known to be involved in high-fat diet (HFD)-induced obesity, the effect of GIP on BCM is still controversial. In this study, we investigated indium 111 (111In)-labeled exendin-4 derivative ([Lys12(111In-BnDTPA-Ahx)]exendin-4) single-photon emission computed tomography/computed tomography (SPECT/CT) as a tool for evaluation of longitudinal BCM changes in HFD-induced obese mice, at the same time we also investigated the effects of GIP on BCM in response to HFD using GIP-knockout (GIP-/-) mice. 111In-exendin-4 SPECT/CT was able to distinguish control-fat diet (CFD)-fed mice from HFD-fed mice and the pancreatic uptake values replicated the BCM measured by conventional histological methods. Furthermore, BCM expansions in HFD-fed mice were demonstrated by time-course changes of the pancreatic uptake values. Additionally, 111In-exendin-4 SPECT/CT demonstrated the distinct changes in BCM between HFD-fed GIP-/- (GIP-/-+HFD) and wild-type (WT+HFD) mice; the pancreatic uptake values of GIP-/-+HFD mice became significantly lower than those of WT+HFD mice. The different changes in the pancreatic uptake values between the two groups preceded those in fat accumulation and insulin resistance. Taken together with the finding of increased ß-cell apoptosis in GIP-/-+HFD mice compared with WT+HFD mice, these data indicated that GIP has preferable effects on BCM under HFD. Therefore, 111In-exendin-4 SPECT/CT can be useful for evaluating increasing BCM and the role of GIP in BCM changes under HFD conditions.

Effects of glucagon-like peptide-1 receptor agonists on cardiovascular and renal outcomes: A meta-analysis and meta-regression analysis.

AIM: To evaluate the cardiovascular and renal outcomes of glucagon-like peptide-1 receptor agonists (GLP-1 RAs) and the associations between these outcomes and HbA1c or weight reduction. MATERIALS AND METHODS: We searched PubMed/MEDLINE, EMBASE, and CENTRAL databases for randomized, placebo-controlled trials of GLP-1 RAs reporting major adverse cardiovascular events (MACE; a composite of cardiovascular mortality, stroke, and myocardial infarction) as the primary outcome. We conducted a meta-regression analysis of primary and secondary outcomes with HbA1c or weight reduction following a meta-analysis with a random-effects model for these outcomes. RESULTS: We extracted data of 60 800 individuals from eight eligible studies (ELIXA, LEADER, SUSTAIN-6, EXSCEL, HARMONY, PIONEER 6, REWIND, and AMPLITUDE-O). GLP-1 RAs reduced MACE (hazard ratio [HR] 0.86; 95% CI: 0.80-0.93; P < .001) and secondary outcomes including the composite renal outcome (0.80; 0.73-0.87; P < .001). In meta-regression analysis, every 1% reduction in HbA1c was associated with 26% and 35% decreases in the logarithm of HR of MACE (P = .044; R2  = 0.65) and the composite renal outcome (P = .040; R2  = 0.85), respectively. On the contrary, weight reduction was not associated with any outcome, including MACE (P = .390). CONCLUSIONS: The reduction in HbA1c, but not body weight, is associated with cardiovascular and renal outcomes. The magnitude of HbA1c reduction can be a surrogate for the cardiovascular and renal benefits of treatment with GLP-1 RAs.

Medium-chain triglycerides inhibit long-chain triglyceride-induced GIP secretion through GPR120-dependent inhibition of CCK.

Long-chain triglycerides (LCTs) intake strongly stimulates GIP secretion from enteroendocrine K cells and induces obesity and insulin resistance partly due to GIP hypersecretion. In this study, we found that medium-chain triglycerides (MCTs) inhibit GIP secretion after single LCT ingestion and clarified the mechanism underlying MCT-induced inhibition of GIP secretion. MCTs reduced the CCK effect after single LCT ingestion in wild-type (WT) mice, and a CCK agonist completely reversed MCT-induced inhibition of GIP secretion. In vitro studies showed that medium-chain fatty acids (MCFAs) inhibit long-chain fatty acid (LCFA)-stimulated CCK secretion and increase in intracellular Ca2+ concentrations through inhibition of GPR120 signaling. Long-term administration of MCTs reduced obesity and insulin resistance in high-LCT diet-fed WT mice, but not in high-LCT diet-fed GIP-knockout mice. Thus, MCT-induced inhibition of GIP hypersecretion reduces obesity and insulin resistance under high-LCT diet feeding condition.

Carbonic anhydrase 8 (CAR8) negatively regulates GLP-1 secretion from enteroendocrine cells in response to long-chain fatty acids.

Glucagon-like peptide-1 (GLP-1) is an incretin secreted from enteroendocrine preproglucagon (PPG)-expressing cells (traditionally known as L cells) in response to luminal nutrients that potentiates insulin secretion. Augmentation of endogenous GLP-1 secretion might well represent a novel therapeutic target for diabetes treatment in addition to the incretin-associated drugs currently in use. In this study, we found that PPG cells substantially express carbonic anhydrase 8 (CAR8), which has been reported to inhibit inositol 1,4,5-trisphosphate (IP3) binding to the IP3 receptor and subsequent Ca2+ efflux from the endoplasmic reticulum in neuronal cells. In vitro experiments using STC-1 cells demonstrated that Car8 knockdown increases long-chain fatty acid (LCFA)-stimulated GLP-1 secretion. This effect was reduced in the presence of phospholipase C (PLC) inhibitor; in addition, Car8 knockdown increased the intracellular Ca2+ elevation caused by α-linolenic acid, indicating that CAR8 exerts its effect on GLP-1 secretion via the PLC/IP3/Ca2+ pathway. Car8wdl null mutant mice showed significant increase in GLP-1 response to oral corn oil administration compared with that in wild-type littermates, with no significant change in intestinal GLP-1 content. These results demonstrate that CAR8 negatively regulates GLP-1 secretion from PPG cells in response to LCFAs, suggesting the possibility of augmentation of postprandial GLP-1 secretion by CAR8 inhibition.NEW & NOTEWORTHY This study focused on the physiological significance of carbonic anhydrase 8 (CAR8) in GLP-1 secretion from enteroendocrine preproglucagon (PPG)-expressing cells. We found an inhibitory role of CAR8 in LCFA-induced GLP-1 secretion in vitro and in vivo, suggesting a novel therapeutic approach to diabetes and obesity through augmentation of postprandial GLP-1 secretion by CAR8 inhibition.

Gene expression of nutrient-sensing molecules in I cells of CCK reporter male mice.

Cholecystokinin (CCK) is secreted from enteroendocrine I cells in response to fat, carbohydrate, and protein ingestion. Gene expression of nutrient-sensing molecules in I cells remains unclear, primarily due to the difficulty in distinguishing I cells from intestinal epithelial cells in vivo. In this study, we generated CCK reporter male mice in which the red fluorescence protein tdTomato (Tomato) is produced by activation of the native murine Cck promoter. Fluorescence microscopy revealed the presence of Tomato-positive cells in upper small intestine (SI), lower SI, and colon. Flow cytometer analysis revealed that Tomato-positive cells among epithelial cells of upper SI, lower SI, and colon occurred at the rate of 0.95, 0.54, and 0.06%, respectively. In upper SI and lower SI, expression levels of Cck mRNA were higher in Tomato-positive cells than those in Tomato-negative cells. The fatty acid receptors Gpr120, Gpr40, and Gpr43 and the oleoylethanolamide receptor Gpr119 were highly expressed in Tomato-positive cells isolated from SI, but were not found in Tomato-positive cells from colon. The glucose and fructose transporters Sglt1, Glut2, and Glut5 were expressed in both Tomato-positive cells and -negative cells, but these expression levels tended to be decreased in Tomato-positive cells from upper SI to colon. The peptide transporter Pept1 and receptor Gpr93 were expressed in both Tomato-positive cells and -negative cells, whereas Casr was expressed only in Tomato-positive cells isolated from SI. Thus, this transgenic mouse reveals that I cell number and gene expression in I cells vary according to region in the gastrointestinal tract.

Absence of GIP secretion alleviates age-related obesity and insulin resistance.

Glucose-dependent insulinotropic polypeptide (GIP) is an incretin secreted from enteroendocine K cells after nutrient ingestion. Fat strongly induces GIP secretion, and GIP hypersecretion is involved in high-fat diet-induced obesity and insulin resistance. Aging also induces GIP hypersecretion, but its effect on body weight gain and insulin sensitivity remains unclear. In the present study, we investigated the effect of GIP on age-related body weight gain and insulin resistance using GIP-knockout homozygous (GIP-/-) and heterozygous (GIP+/-) mice, which have entirely absent and 50% reduced GIP secretion compared to wild-type (WT) mice, respectively. Under 12% fat-containing normal diet feeding condition, body weight was significantly lower in GIP-/- mice compared to that in WT and GIP+/- mice from 38 weeks of age, while there was no significant difference between WT and GIP+/- mice. Visceral and s.c. fat mass were also significantly lower in GIP-/- mice compared to those in WT and GIP+/- mice. During oral glucose tolerance test, blood glucose levels did not differ among the three groups. Insulin levels were significantly lower in GIP-/- mice than those in WT and GIP+/- mice. During insulin tolerance test, GIP-/- mice showed higher insulin sensitivity than that of WT and GIP+/- mice. Adiponectin mRNA levels were increased and leptin mRNA levels tended to be decreased in adipose tissue of GIP-/- mice. These results demonstrate that GIP is involved in age-related obesity and insulin resistance and that inhibition of GIP secretion alleviates age-related fat mass gain and insulin resistance under carbohydrate-based diet feeding condition.

Free fatty acid receptors, G protein-coupled receptor 120 and G protein-coupled receptor 40, are essential for oil-induced gastric inhibitory polypeptide secretion.

AIMS/INTRODUCTION: Incretin hormone glucose-dependent insulinotropic polypeptide/gastric inhibitory polypeptide (GIP) plays a key role in high-fat diet-induced obesity and insulin resistance. GIP is strongly secreted from enteroendocrine K cells by oil ingestion. G protein-coupled receptor (GPR)120 and GPR40 are two major receptors for long chain fatty acids, and are expressed in enteroendocrine K cells. In the present study, we investigated the effect of the two receptors on oil-induced GIP secretion using GPR120- and GPR40-double knockout (DKO) mice. MATERIALS AND METHODS: Global knockout mice of GPR120 and GPR40 were crossbred to generate DKO mice. Oral glucose tolerance test and oral corn oil tolerance test were carried out. For analysis of the number of K cells and gene expression in K cells, DKO mice were crossbred with GIP-green fluorescent protein knock-in mice in which visualization and isolation of K cells can be achieved. RESULTS: Double knockout mice showed normal glucose-induced GIP secretion, but no GIP secretion by oil. We then investigated the number of K cells and gene characteristics in K cells isolated from GIP-green fluorescent protein knock-in mice. Deficiency of both receptors did not affect the number of K cells in the small intestine or expression of GIP messenger ribonucleic acid in K cells. Furthermore, there was no significant difference in the expression of the genes associated with lipid absorption or GIP secretion in K cells between wild-type and DKO mice. CONCLUSIONS: Oil-induced GIP secretion is triggered by the two major fatty acid receptors, GPR120 and GPR40, without changing K-cell number or K-cell characteristics.

Medium-chain triglyceride diet stimulates less GIP secretion and suppresses body weight and fat mass gain compared with long-chain triglyceride diet.

Gastric inhibitory polypeptide (GIP) is an incretin secreted from enteroendocrine K cells and potentiates insulin secretion from pancreatic ß-cells. GIP also enhances long-chain triglyceride (LCT) diet-induced obesity and insulin resistance. Long-term intake of medium-chain triglyceride (MCT) diet is known to induce less body weight and fat mass gain than that of LCT diet. However, the effect of MCT diet feeding on GIP secretion and the effect of GIP on body weight and fat mass under MCT diet-feeding condition are unknown. In this study, we evaluated the effect of single MCT oil administration on GIP secretion and compared the effect of long-term MCT and LCT diet on body weight and fat mass gain in wild-type (WT) and GIP-knockout (GIP KO) mice. Single administration of LCT oil induced GIP secretion but that of MCT oil did not in WT mice. Long-term intake of LCT diet induced GIP hypersecretion and significant body weight and fat mass gain compared with that of control fat (CF) diet in WT mice. In contrast, MCT diet did not induce GIP hypersecretion, and MCT diet-fed mice showed smaller increase in body weight and fat mass gain compared with CF diet-fed mice. In GIP KO mice, body weight and fat mass were markedly attenuated in LCT diet-fed mice but not in MCT diet-fed mice. Our results suggest that long-term intake of MCT diet stimulates less GIP secretion and suppresses body weight and fat mass gain compared with that of LCT diet.

Gastric inhibitory polypeptide/glucose-dependent insulinotropic polypeptide signaling in adipose tissue.

GIPR signaling in adipose tissue plays an important role in HFD-induced insulin resistance and hepatic steatosis in vivo, with no direct effect on fat accumulation, through IL-6 signaling.

Glucose-dependent insulinotropic polypeptide deficiency reduced fat accumulation and insulin resistance, but deteriorated bone loss in ovariectomized mice.

Given the established roles of glucose-dependent insulinotropic polypeptide (GIP) in promoting fat storage and bone formation, we assessed the contribution of GIP to obesity and osteopenia in ovariectomized mice with a gene encoding green fluorescent protein (GFP) inserted into the GIP locus, in which GIP was either reduced (GIPgfp/+ ) or absent (GIPgfp/gfp ). In GIPgfp/gfp mice, weight gain, subcutaneous and visceral fat mass were reduced, and glucose intolerance was improved compared with wild-type mice with the same magnitude of insulin responses. Cancellous bone mineral density and bone cortical thickness were reduced in GIPgfp/gfp mice compared with wild-type mice. In GIPgfp/+ mice, weight gain, glucose intolerance and cancellous bone mineral density were not different from that of wild-type mice. These results indicate that the total elimination of GIP ameliorates weight gain and adiposity in ovariectomized mice, but it enhances osteopenia, particularly in cancellous bone by partly suppressing bone formation.

The Effect of White Rice and White Bread as Staple Foods on Gut Microbiota and Host Metabolism.

The purpose of this study was to examine the influence of two kinds of major Japanese staple foods, white rice and white bread, on gut microbiota against the background in which participants eat common side dishes. Seven healthy subjects completed the dietary intervention with two 1-week test periods with a 1-week wash-out period in cross-over design (UMIN registration UMIN000023142). White bread or white rice and 21 frozen prepared side dishes were consumed during the test periods. At baseline and at the end of each period, fasting blood samples, breath samples, and fecal samples were collected. For fecal samples, 16S rRNA gene sequencing was used to analyze the gut microbiota. After the bread period, the abundance of fecal Bifidobacterium genus (19.2 ± 14.5 vs. 6.2 ± 6.6 (%), p = 0.03), fasting glucagon-like peptide 1 (GLP-1) (13.6 ± 2.0 vs. 10.5 ± 2.9 (pg/mL), p = 0.03), and breath hydrogen (23.4 ± 9.9 vs. 8.2 ± 5.5 (ppm), p = 0.02) were significantly higher than those of after the rice period. Plasma SCFAs also tended to be higher after the bread period. White bread contains more dietary fiber than refined short grain rice. These findings suggest that indigestible carbohydrate intake from short grain rice as a staple food may be smaller than that of white bread.

Transcriptional factor Pdx1 is involved in age-related GIP hypersecretion in mice.

Fat accumulation with aging is a serious problem; glucose-dependent insulinotropic polypeptide/gastric inhibitory polypeptide (GIP) is an incretin that plays an important role in fat accumulation. GIP receptor knockout mice show reduced fat mass and improved insulin sensitivity associated with aging. Therefore, GIP is involved in fat accumulation and insulin resistance with aging. However, age-related changes of GIP secretion remain unclear. The present study aimed to elucidate age-related changes of GIP secretion and enteroendocrine K cells using GIP reporter [GIP-green fluorescent protein (GFP) knock-in heterozygous (GIPgfp/+)] mice. Aged 1-yr-old GIPgfp/+ mice exhibited a phenotype of fat accumulation, insulin resistance, and GIP hypersecretion compared with young (3-4 mo old) GIPgfp/+ mice. In aged mice, K-cell number in the small intestine and the mRNA expression levels of GIP and transcriptional factor pancreatic and duodenal homeobox-1 (Pdx1) in K cells were increased. K-cell number, GIP mRNA expression and content in small intestine, and GIP secretion were decreased after posteriori suppression of Pdx1 using intestine-specific gene transfer. Thus, Pdx1 positively regulates GIP mRNA and K-cell number in small intestine. Increased Pdx1 expression might be involved in GIP hypersecretion with aging. NEW & NOTEWORTHY Age-related changes of glucose-dependent insulinotropic polypeptide/gastric inhibitory polypeptide (GIP) secretion and K cells were investigated. We found that K-cell number and GIP and pancreatic and duodenal homeobox-1 (Pdx1) expression in K cells were increased in aged mice, which showed greater GIP secretion compared with young mice. In addition, we have succeeded in posteriori suppression of Pdx1 in small intestine using the method of intestine-specific gene transfer, and showed that K-cell number, GIP expression, and GIP secretion were decreased in the Pdx1-knockdown intestine.

ß-cell-specific overexpression of adiponectin receptor 1 does not improve diabetes mellitus in Akita mice.

Adiponectin, a metabolically-active cytokine secreted from adipose tissue, is reported to have anti-apoptotic effects on ß-cells as well as anti-hyperglycemic effects through adiponectin receptor signaling. However, the anti-apoptotic effects of adiponectin on ß-cells have not been confirmed in established diabetic models, and the anti-hyperglycemic effects and their associated signal cascades remain controversial. To investigate the effects of adiponectin on ß-cell protection and its down-stream signaling events, we have generated ß-cell-specific rat insulin promoter (RIP)-AdipoR1 transgenic mice (AdipoR1 mice), in which the adiponectin receptor, AdipoR1, is overexpressed in ß-cells in a manner synchronous with insulin demand. AdipoR1 mice were then mated with Akita mice, a diabetes model in which ß-cell apoptosis results from endoplasmic reticulum (ER) stress. AdipoR1 protein expression and localization in islets from AdipoR1 mice as well as in an AdipoR1-transfected mouse insulinoma cell line were confirmed, as was the activation of both AMPK and Akt in AdipoR1 mice by adiponectin. Nevertheless, there were no significant differences in Ad lib feed and fasting blood glucose levels, or in glucose tolerance tests, between Akita mice [Ins2Akita (C96Y) +/- mouse model] and AdipoR1/Akita and from 4 weeks to 10 weeks of age. Similarly, pancreatic insulin contents of AdipoR1/Akita mice were not significantly different from those in Akita mice from 15 to 20 weeks of age, but they were significantly lower than in wild-type mice. Immunostaining for insulin and subsequent electron microscopy showed that ß-cell destruction in AdipoR1/Akita mice was not markedly improved in comparison with that in Akita mice. Serum adiponectin concentrations were confirmed to be extremely high (> 30 µg / ml) compared with the Kd value (0.06 µg / ml) in all mouse groups at 15 to 20 weeks of age. Therefore, although the physiological levels of adiponectin are sufficient to activate AMPK and Akt when AdipoR1 is overexpressed in ß-cells, yet adiponectin cannot protect ß-cells in Akita mice from ER stress-induced destruction.