Alpha Cell Ratio Research Articles

Analysis of beta-cell maturity and mitochondrial morphology in juvenile non-human primates exposed to maternal Western-style diet during development.

Using a non-human primate (NHP) model of maternal Western-style diet (mWSD) feeding during pregnancy and lactation, we previously reported altered offspring beta:alpha cell ratio in vivo and insulin hyper-secretion ex vivo. Mitochondria are known to maintain beta-cell function by producing ATP for insulin secretion. In response to nutrient stress, the mitochondrial network within beta cells undergoes morphological changes to maintain respiration and metabolic adaptability. Given that mitochondrial dynamics have also been associated with cellular fate transitions, we assessed whether mWSD exposure was associated with changes in markers of beta-cell maturity and/or mitochondrial morphology that might explain the offspring islet phenotype. We evaluated the expression of beta-cell identity/maturity markers (NKX6.1, MAFB, UCN3) via florescence microscopy in islets of Japanese macaque pre-adolescent (1 year old) and peri-adolescent (3-year-old) offspring born to dams fed either a control diet or WSD during pregnancy and lactation and weaned onto WSD. Mitochondrial morphology in NHP offspring beta cells was analyzed in 2D by transmission electron microscopy and in 3D using super resolution microscopy to deconvolve the beta-cell mitochondrial network. There was no difference in the percent of beta cells expressing key maturity markers in NHP offspring from WSD-fed dams at 1 or 3 years of age; however, beta cells of WSD-exposed 3 year old offspring showed increased levels of NKX6.1 per beta cell at 3 years of age. Regardless of maternal diet, the beta-cell mitochondrial network was found to be primarily short and fragmented at both ages in NHP; overall mitochondrial volume increased with age. In utero and lactational exposure to maternal WSD consumption may increase mitochondrial fragmentation. Despite mWSD consumption having clear developmental effects on offspring beta:alpha cell ratio and insulin secretory response to glucose, this does not appear to be mediated by changes to beta-cell maturity or the beta-cell mitochondrial network. In general, the more fragmented mitochondrial network in NHP beta cells suggests greater ability for metabolic flexibility.

1465-P: Metabolic Remodeling in Pancreatic Beta Cells in Japanese Patients with Type 2 Diabetes

Background and Aims: It has been shown that metabolic remodeling from oxidative phosphorylation to glycolysis via activation of 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) occurs in pancreatic beta cells in non-Asians with diabetes, resulting in impaired beta cell function. We used multiple autopsy pancreatic sections to investigate whether such changes occur in Japanese patients with type 2 diabetes (T2D) , and if so, which clinical and pathological features of islets relate to this remodeling. Methods: Autopsy pancreatic sections from patients with T2D and non-diabetic donors (ND) were stained with antibodies against PFKFB3, insulin, and glucagon. The PFKFB3-positive ratio of pancreatic beta cells and alpha cells between T2D and ND were compared. Next, the relationships between PFKFB3-postive beta cell ratio, patient background factors and pathological features of the islets were explored. Results: In total, 36 cases (18 cases each for T2D and ND) were enrolled and 50-100 islets/case were evaluated. Mean age was 70.6 ± 7.7 vs. 64.3 ± 12.8 years and body mass index (BMI) 22.4 ± 4.2 vs. 23.3 ± 4.5 kg/m2, respectively. T2D duration was 9.8 ± 9.9 years. The PFKFB3-positive beta cell ratio was significantly higher in T2D (29.7 ± 18.6% vs. 11.1 ± 8.7%, p<0.05) , whereas the PFKFB3-positive alpha cell ratio did not differ between groups. A negative correlation between PFKFB3-positive beta cell ratio and BMI was found only in the T2D group (r=-0.65, p<0.05) . No correlation was observed between PFKFB3-positive beta cell ratio and age/disease duration in the T2D group. Thioflavin T staining revealed a positive correlation between amyloid deposition in the islets of T2D and PFKFB3 positivity in beta cells (r=0.67, p<0.05) . Conclusion: Our results indicate that intracellular metabolic remodeling mediated by upregulation of PFKFB3 occurs in pancreatic beta cells of Japanese patients with T2D. Low BMI and amyloid deposition may affect PFKFB3 expression in beta cells from patients with T2D. Disclosure R.Izumihara: None. H.Nomoto: None. K.Chiba: Research Support; Grant for Young Researcher from Japan Association for Diabetes Education and Care, Japan Diabetes Society Junior Scientist Development Grant, Japan Society for the Promotion of Science , MSD Life Science Foundation, Promotion for Young Research Talent and Network, The Akiyama Life Science Foundation, The Uehara Memorial Foundation. H.Kameda: None. K.Cho: None. A.Nakamura: Research Support; Kissei Pharmaceutical Co., Ltd., MSD, Nippon Boehringer Ingelheim, Taisho Pharmaceutical Holdings Co., Ltd. H.Miyoshi: Research Support; Abbott Japan Co., Ltd., Boehringer Ingelheim International GmbH, Daiichi Sankyo, Kowa Company, Ltd., LifeScan, Mitsubishi Tanabe Pharma Corporation, Novo Nordisk, Ono Pharmaceutical Co., Ltd., Sumitomo Dainippon Pharma Co., Ltd., Taisho Pharmaceutical Holdings Co., Ltd., Speaker's Bureau; Astellas Pharma Inc., Boehringer Ingelheim International GmbH, Eli Lilly and Company, Kowa Company, Ltd., Merck & Co., Inc., Mitsubishi Tanabe Pharma Corporation, Novo Nordisk, Ono Pharmaceutical Co., Ltd., Sanofi K.K., Sumitomo Dainippon Pharma Co., Ltd., Taisho Pharmaceutical Holdings Co., Ltd. H.Mizukami: None. T.Atsumi: Consultant; AbbVie Inc., AstraZeneca, MEDICAL & BIOLOGICAL LABORATORIES CO., Nippon Boehringer Ingelheim Co. Ltd., Novartis Pharma K.K., Ono Pharmaceutical Co., Ltd., Pfizer Inc., Research Support; AbbVie Inc., Alexion Pharmaceuticals, Inc., Astellas Pharma Inc., Chugai Pharmaceutical Co., Ltd., Daiichi Sankyo, Mitsubishi Tanabe Pharma Corporation, Nippon Boehringer Ingelheim Co. Ltd., Otsuka Pharmaceutical Co., Ltd., Pfizer Inc., Taiho Pharmaceutical Co. Ltd., Takeda Pharmaceutical Company Limited, Teijin Pharma Limited, Speaker's Bureau; AbbVie Inc., Astellas Pharma Inc., Bristol-Myers Squibb Company, Chugai Pharmaceutical Co., Ltd., Daiichi Sankyo, Eisai Co., Ltd., Eli Lilly and Company, Kyowa Kirin Co., Ltd., Mitsubishi Tanabe Pharma Corporation, Novartis Pharma K.K., Pfizer Inc., Taiho Pharmaceutical Co. Ltd., Takeda Pharmaceutical Company Limited, UCB, Inc. Funding JSPS KAKENHI Grant Number JP20281167.Japan Diabetes Society Junior Scientist Development Grant.MSD Life Science Foundation, Public Interest Incorporated Foundation.

The methionine aminopeptidase 2 inhibitor, TNP-470, enhances the antidiabetic properties of sitagliptin in mice by upregulating xenin

The therapeutic mechanism of action of methionine aminopeptidase 2 (MetAP2) inhibitors for obesity-diabetes has not yet been fully defined. Xenin, a K-cell derived peptide hormone, possesses an N-terminal Met amino acid residue. Thus, elevated xenin levels could represent a potential pharmacological mechanism of MetAP2 inhibitors, since long-acting xenin analogues have been shown to improve obesity-diabetes. The present study has assessed the ability of the MetAP2 inhibitor, TNP-470, to augment the antidiabetic utility of the incretin-enhancer drug, sitagliptin, in high fat fed (HFF) mice. TNP-470 (1 mg/kg) and sitagliptin (25 mg/kg) were administered once-daily alone, or in combination, to diabetic HFF mice (n = 10) for 18 days. Individual therapy with TNP-470 or sitagliptin resulted in numerous metabolic benefits including reduced blood glucose, increased circulating and pancreatic insulin and improved glucose tolerance, insulin sensitivity, pyruvate tolerance and overall pancreatic islet architecture. Further assessment of metabolic rate revealed that all treatments reduced respiratory exchange ratio and increased locomotor activity. All sitagliptin treated mice also exhibited increased energy expenditure. In addition, treatment with TNP-470 alone, or in combination with sitagliptin, reduced food intake and body weight, as well as elevating plasma and intestinal xenin. Importantly, combined sitagliptin and TNP-470 therapy was associated with further significant benefits beyond that observed by either treatment alone. This included more rapid restoration of normoglycaemia, superior glucose tolerance, increased circulating GIP concentrations and an enhanced pancreatic beta:alpha cell ratio. In conclusion, these data demonstrate that TNP-470 increases plasma and intestinal xenin levels, and augments the antidiabetic advantages of sitagliptin.

MafA is critical for maintenance of the mature beta cell phenotype in mice.

The plasticity of adult somatic cells allows for their dedifferentiation or conversion to different cell types, although the relevance of this to disease remains elusive. Perturbation of beta cell identity leading to dedifferentiation may be implicated in the compromised functions of beta cells in diabetes, which is a current topic of islet research. This study aims to investigate whether or not v-Maf musculoaponeurotic fibrosarcoma oncogene family, protein A (MafA), a mature beta cell marker, is involved in maintaining mature beta cell phenotypes. The fate and gene expression of beta cells were analysed in Mafa knockout (KO) mice and mouse models of diabetes in which the expression of MafA was reduced in the majority of beta cells. Loss of MafA reduced the beta to alpha cell ratio in pancreatic islets without elevating blood glucose to diabetic levels. Lineage tracing analyses showed reduced/lost expression of insulin in most beta cells, with a minority of the former beta cells converted to glucagon-expressing cells in Mafa KO mice. The upregulation of genes that are normally repressed in mature beta cells or transcription factors that are transiently expressed in endocrine progenitors was identified in Mafa KO islets as a hallmark of dedifferentiation. The compromised beta cells in db/db and multiple low-dose streptozotocin mice underwent similar dedifferentiation with expression of Mafb, which is expressed in immature beta cells. The maturation factor MafA is critical for the homeostasis of mature beta cells and regulates cell plasticity. The loss of MafA in beta cells leads to a deeper loss of cell identity, which is implicated in diabetes pathology.

Abnormal infant islet morphology precedes insulin resistance in PCOS-like monkeys.

Polycystic ovary syndrome (PCOS) is prevalent in reproductive-aged women and confounded by metabolic morbidities, including insulin resistance and type 2 diabetes. Although the etiology of PCOS is undefined, contribution of prenatal androgen (PA) exposure has been proposed in a rhesus monkey model as premenopausal PA female adults have PCOS-like phenotypes in addition to insulin resistance and decreased glucose tolerance. PA female infants exhibit relative hyperinsulinemia, suggesting prenatal sequelae of androgen excess on glucose metabolism and an antecedent to future metabolic disease. We assessed consequences of PA exposure on pancreatic islet morphology to identify evidence of programming on islet development. Islet counts and size were quantified and correlated with data from intravenous glucose tolerance tests (ivGTT) obtained from dams and their offspring. Average islet size was decreased in PA female infants along with corresponding increases in islet number, while islet fractional area was preserved. Infants also demonstrated an increase in both the proliferation marker Ki67 within islets and the beta to alpha cell ratio suggestive of enhanced beta cell expansion. PA adult females have reduced proportion of small islets without changes in proliferative or apoptotic markers, or in beta to alpha cell ratios. Together, these data suggest in utero androgen excess combined with mild maternal glucose intolerance alter infant and adult islet morphology, implicating deviant islet development. Marked infant, but subtle adult, morphological differences provide evidence of islet post-natal plasticity in adapting to changing physiologic demands: from insulin sensitivity and relative hypersecretion to insulin resistance and diminished insulin response to glucose in the mature PCOS-like phenotype.

The role of caspase-8 in amyloid-induced beta cell death in human and mouse islets

Reduced beta cell mass due to increased beta cell apoptosis is a key defect in type 2 diabetes. Islet amyloid, formed by the aggregation of human islet amyloid polypeptide (hIAPP), contributes to beta cell death in type 2 diabetes and in islet grafts in patients with type 1 diabetes. In this study, we used human islets and hIAPP-expressing mouse islets with beta cell Casp8 deletion to (1) investigate the role of caspase-8 in amyloid-induced beta cell apoptosis and (2) test whether caspase-8 inhibition protects beta cells from amyloid toxicity. Human islet cells were cultured with hIAPP alone, or with caspase-8, Fas or amyloid inhibitors. Human islets and wild-type or hIAPP-expressing mouse islets with or without caspase-8 expression (generated using a Cre/loxP system) were cultured to form amyloid. Caspase-8 and -3 activation, Fas and FLICE inhibitory protein (FLIP) expression, islet beta cell and amyloid area, IL-1β levels, and the beta:alpha cell ratio were assessed. hIAPP treatment induced activation of caspase-8 and -3 in islet beta cells (via Fas upregulation), resulting in apoptosis, which was markedly reduced by blocking caspase-8, Fas or amyloid. Amyloid formation in cultured human and hIAPP-expressing mouse islets induced caspase-8 activation, which was associated with Fas upregulation and elevated islet IL-1β levels. hIAPP-expressing mouse islets with Casp8 deletion had comparable amyloid, IL-1β and Fas levels with those expressing hIAPP and Casp8, but markedly lower beta cell apoptosis, higher beta:alpha cell ratio, greater beta cell area, and enhanced beta cell function. Beta cell Fas upregulation by endogenously produced and exogenously applied hIAPP aggregates promotes caspase-8 activation, resulting in beta cell apoptosis. The prevention of amyloid-induced caspase-8 activation enhances beta cell survival and function in islets.

Effects of Combining Linagliptin Treatment with BI-38335, A Novel SGLT2 Inhibitor, on Pancreatic Islet Function and Inflammation in db/db Mice

Dipeptidyl peptidase-4 (DPP-4) inhibitors enhance incretin actions and beta-cell function. Concurrently, sodium-glucose co-transporter 2 (SGLT2) inhibitors block renal glucose reabsorption promoting excretion. In this study, we investigated the effects of linagliptin (a DPP-4 inhibitor) and BI-38335 (an SGLT2 inhibitor), individually and in combination, on glucose homeostasis, islet function, and pancreatic islet morphology in db/db mice. Diabetic and non-diabetic mice received linagliptin (3 mg/kg), BI-38335 (1 mg/kg), the two drugs in combination or control once daily for 8 weeks. Blood glucose homeostasis and insulin sensitivity were assessed. Pancreatic islet function and morphology as well as inflammatory factors and toll like receptor 2 (TLR2) pathways involved in islet inflammation were investigated. Active treatments markedly reduced blood glucose and glycated hemoglobin A1c (HbA(1c)) levels, with the combined treatment showing the greater effects. Insulin resistance was improved in the BI-38335 and combination groups with the enhancement of insulin sensitivity and significant increase of serum adiponectin levels. The combined treatment exhibited greater effects on enhanced islet glucose-stimulated insulin secretion and improved glucose tolerance. Moreover, the combination restored the islet beta-/alpha-cell ratio, reduced beta-cell apoptosis, decreased expression of islet immune cell markers, and suppressed factors related to the TLR2 pathway. In addition, all active treatments reduced serum lipid profiles, though the combination produced more robust effects. Collectively, our data show that combined treatment with BI-38335 and linagliptin work, at least in part, synergistically to benefit islet cell function/architecture and insulin resistance, thus improving glycemic control.

Deletion of Fas protects islet beta cells from cytotoxic effects of human islet amyloid polypeptide.

AIMS/HYPOTHESIS: Islet amyloid, which is mainly composed of human islet amyloid polypeptide (hIAPP), is a pathological characteristic of type 2 diabetes and also forms in cultured and transplanted islets. We used islet beta cells as well as two ex vivo models of islet amyloid formation, cultured human islets and hIAPP-expressing transgenic mouse islets with or without beta cell Fas deletion, to test whether: (1) the aggregation of endogenous hIAPP induces Fas upregulation in beta cells; and (2) deletion or blocking of Fas protects beta cells from amyloid toxicity. METHODS: INS-1, mouse or human islet cells were cultured with hIAPP alone, or with amyloid inhibitor or Fas antagonist. Non-transduced islets, and human islets or hIAPP-expressing mouse islets transduced with an adenovirus that delivers a human proIAPP-specific small interfering RNA (siRNA) (Ad-ProhIAPP-siRNA) were cultured to form amyloid. Mouse islets expressing hIAPP with or without Fas were similarly cultured. Beta cell Fas upregulation, caspase-3 activation, apoptosis and function, and islet IL-1β levels were assessed. RESULTS: hIAPP treatment induced Fas upregulation, caspase-3 activation and apoptosis in INS-1 and islet cells. The amyloid inhibitor or Fas antagonist reduced apoptosis in hIAPP-treated beta cells. Islet cells with Fas deletion had lower hIAPP-induced beta cell apoptosis than those expressing Fas. Ad-ProhIAPP-siRNA-mediated amyloid inhibition reduced Fas upregulation and IL-1β immunoreactivity in human and hIAPP-expressing mouse islets. Cultured hIAPP-expressing mouse islets with Fas deletion had similar amyloid levels, but lower caspase-3 activation and beta cell apoptosis, and a higher islet beta:alpha cell ratio and insulin response to glucose, compared with islets expressing Fas and hIAPP. CONCLUSIONS/INTERPRETATION: The aggregation of biosynthetic hIAPP produced in islets induces beta cell apoptosis, at least partially, via Fas upregulation and the Fas-mediated apoptotic pathway. Deletion of Fas protects islet beta cells from the cytotoxic effects of endogenously secreted (and exogenously applied) hIAPP.

The protective effect of ursodeoxycholic acid in alloxan-induced diabetes.

The purpose of this work was to study the effect of ursodeoxycholic acid (UDCA) on the morphological and functional alterations in pancreatic islet beta-cells in rats with diabetes induced by alloxan (150 mg kg(-1), i.p.). UDCA (40 mg kg(-1), i.g.) was administered daily from the fifth to the 35th day after the alloxan treatment. The treatment of diabetic rats with UDCA improved the pancreatic morphology disturbed by the alloxan treatment: UDCA increased the number of pancreatic islets and beta-cells, the beta-/alpha-cell ratio and decreased the number of alpha-cells. As the morphometric data suggest, the treatment of diabetic animals with UDCA significantly increased the area of beta-cell cytoplasmatic granules stained by paraldehyde-fuchsin. The concentration of blood glucose in diabetic rats was gradually decreased after the UDCA treatment, and at the end of the experiment reached the control value. The treatment with UDCA raised the serum insulin level in diabetic rats about 2.5-fold, but this concentration was significantly lower as compared to the control group. The content of lipid peroxidation end-products, hydroxyalkenals and malondialdehyde, was significantly elevated in the alloxan-treated rats, whereas the treatment with UDCA normalized these parameters. The present data indicate that UDCA acts as an effective antidiabetic agent in alloxan-induced diabetes and its beneficial effects in diabetic rats can be related to the antioxidant properties of UDCA.