RESUMEN
PURPOSE: ß cell mass (BCM) and function are essential to the diagnosis and therapy of diabetes. Diabetic patients serve ß cell loss is, and damage of ß cells leads to severe insulin deficiency. Our understanding of the role of BCM in diabetes progression is extremely limited by lacking efficient methods to evaluate BCM in vivo. In vitro methods of labeling islets, including loading of contrast reagent or integration of exogenous biomarker, require artificial manipulation on islets, of which the clinical application is limited. Imaging methods targeting endogenous biomarkers may solve the above problems. However, traditional reagents targeting GLP-1R and VAMT2 result in a high background of adjacent tissues, complicating the identification of pancreatic signals. Here, we report a non-invasive and quantitative imaging technique by using radiolabeled glycine mimics ([18F]FBG, a boron-trifluoride derivative of glycine) to assay islet function and monitor BCM changes in living animals. METHODS: Glycine derivatives, FBG, FBSa, 2Me-FBG, 3Me-FBG, were successfully synthesized and labeled with 18F. Specificity of glycine derivatives were characterized by in vitro experiment. PET imaging and biodistribution studies were performed in animal models carring GLYT over-expressed cells. In vivo evaluation of BCM with [18F]FBG were performed in STZ (streptozocin) induced T1D (type 1 diabetes) models. RESULTS: GLYT responds to excess blood glycine levels and transports glycine into islet cells to maintain the activity of the glycine receptor (GLYR). Best PET imaging condition was 80 min after given a total of 240 ~ 250 nmol imaging reagent (a mixture of [18F]FBG and natural glycine) intravenously. [18F]FBG can detect both endogenous and exogenous islets clearly in vivo. When applied to STZ induced T1D mouse models, total uptake of [18F]FBG in the pancreas exhibited a linear correlation with survival BCM. CONCLUSION: [18F]FBG targeting the endogenous glycine transporter (GLYT), which is highly expressed on islet cells, avoiding extra modification on islet cells. Meanwhile the highly restricted expression pattern of GLYT excluded the background in adjacent tissues. This [18F]FBG-based imaging technique provides a non-invasive method to quantify BCM in vivo, implying a new evaluation index for diabetic assessment.
Asunto(s)
Glicina , Células Secretoras de Insulina , Animales , Células Secretoras de Insulina/metabolismo , Ratones , Glicina/análogos & derivados , Distribución Tisular , Biomarcadores/metabolismo , Tomografía de Emisión de Positrones/métodos , Masculino , Radioisótopos de Flúor , Humanos , Compuestos de Boro/química , Diabetes Mellitus Experimental/diagnóstico por imagen , Diabetes Mellitus Experimental/metabolismoRESUMEN
Elucidating the mechanism(s) modulating appropriate tissue size is a critical biological issue. Pancreatic ß cells increase during pregnancy via cellular proliferation, but how ß cells promptly decrease to the original amount after parturition remains unclear. Herein, we demonstrate the role and mechanism of macrophage accumulation in this process. In the final stage of pregnancy, HTR1D signaling upregulates murine ß cell CXCL10, thereby promoting macrophage accumulation in pancreatic islets via the CXCL10-CXCR3 axis. Blocking this mechanism by administering an HTR1D antagonist or the CXCR3 antibody and depleting islet macrophages inhibited postpartum ß cell mass reduction. ß cells engulfed by macrophages increased in postpartum islets, but Annexin V administration suppressed this engulfment and the postpartum ß cell mass reduction, indicating the accumulated macrophages to phagocytose ß cells. This mechanism contributes to both maintenance of appropriate ß cell mass and glucose homeostasis promptly adapting to reduced systemic insulin demand after parturition.
Asunto(s)
Células Secretoras de Insulina , Islotes Pancreáticos , Embarazo , Femenino , Ratones , Animales , Células Secretoras de Insulina/fisiología , Parto , Insulina , Macrófagos , FagocitosisRESUMEN
It is crucial to develop practical and noninvasive methods to assess the functional beta-cell mass in a donor pancreas, in which monitoring and precise evaluation is challenging. A patient with type 1 diabetes underwent noninvasive imaging following simultaneous kidney-pancreas transplantation with positron emission tomography/computed tomography (PET/CT) using an exendin-based probe, [18 F]FB(ePEG12)12-exendin-4. Following transplantation, PET imaging with [18 F]FB(ePEG12)12-exendin-4 revealed simultaneous and distinct accumulations in the donor and native pancreases. The pancreases were outlined at a reasonable distance from the surrounding organs using [18 F]FB(ePEG12)12-exendin-4 whole-body maximum intensity projection and axial PET images. At 1 and 2 h after [18 F]FB(ePEG12)12-exendin-4 administration, the mean standardized uptake values were 2.96 and 3.08, respectively, in the donor pancreas and 1.97 and 2.25, respectively, in the native pancreas. [18 F]FB(ePEG12)12-exendin-4 positron emission tomography imaging allowed repeatable and quantitative assessment of beta-cell mass following simultaneous kidney-pancreas transplantation.
Asunto(s)
Trasplante de Riñón , Tomografía Computarizada por Tomografía de Emisión de Positrones , Humanos , Exenatida , Pancrelipasa , Péptidos , Páncreas/diagnóstico por imagenRESUMEN
Imaging with radiolabeled exendin enables detection and characterization of glucagon-like peptide 1 receptors (GLP-1Rs) in vivo with high specificity. The novel radiotracer [68Ga]Ga-NODAGA-exendin-4 forms a stable complex after a simple and fast labeling procedure. Beta-cell mass in the islets of Langerhans can be visualized using [68Ga]Ga-NODAGA-exendin-4, which is promising for research into diabetes mellitus (DM) pathophysiology. Furthermore, this radiotracer enables very sensitive detection of insulinomas, resulting from vast overexpression of GLP-1Rs, and seems promising for the detection of focal lesions in congenital hyperinsulinism (CHI). Here, we describe the procedures involved in [68Ga]Ga-NODAGA-exendin-4 positron emission tomography (PET)/computed tomography (CT) imaging including the radiolabeling of the NODAGA-exendin conjugate with 68Ga, quality controls, and PET/CT.
Asunto(s)
Receptor del Péptido 1 Similar al Glucagón , Neoplasias Pancreáticas , Humanos , Exenatida , Tomografía Computarizada por Tomografía de Emisión de Positrones , Radioisótopos de Galio , Péptidos/química , Neoplasias Pancreáticas/patología , Tomografía de Emisión de Positrones/métodosRESUMEN
Understanding the biology underlying the mechanisms and pathways regulating pancreatic ß cell development is necessary to understand the pathology of diabetes mellitus (DM), which is characterized by the progressive reduction in insulin-producing ß cell mass. Pluripotent stem cells (PSCs) can potentially offer an unlimited supply of functional ß cells for cellular therapy and disease modeling of DM. Homeobox protein NKX6.1 is a transcription factor (TF) that plays a critical role in pancreatic ß cell function and proliferation. In human pancreatic islet, NKX6.1 expression is exclusive to ß cells and is undetectable in other islet cells. Several reports showed that activation of NKX6.1 in PSC-derived pancreatic progenitors (MPCs), expressing PDX1 (PDX1+/NKX6.1+), warrants their future commitment to monohormonal ß cells. However, further differentiation of MPCs lacking NKX6.1 expression (PDX1+/NKX6.1-) results in an undesirable generation of non-functional polyhormonal ß cells. The importance of NKX6.1 as a crucial regulator in MPC specification into functional ß cells directs attentions to further investigating its mechanism and enhancing NKX6.1 expression as a means to increase ß cell function and mass. Here, we shed light on the role of NKX6.1 during pancreatic ß cell development and in directing the MPCs to functional monohormonal lineage. Furthermore, we address the transcriptional mechanisms and targets of NKX6.1 as well as its association with diabetes.
Asunto(s)
Proteínas de Homeodominio , Células Secretoras de Insulina , Diferenciación Celular , Proliferación Celular , Proteínas de Homeodominio/genética , Humanos , Transactivadores/genéticaRESUMEN
BACKGROUND: Ileal transposition (IT) is a surgical procedure to investigate the role of the distal small intestine in metabolic improvements induced by bariatric/metabolic surgery, which has been applied to some human cases. We performed IT in diet-induced obese rats to investigate the effect of IT on glucose metabolism and ß cell senescence. METHODS: Sprague-Dawley rats were fed high-fat diet (60% of total calories from fat) for 12 weeks and randomized into either IT or sham surgery. In the IT group, the distal ileal segment located between 5 and 15 cm proximal to the ileocecal valve was transposed 10 cm distal to the Treitz ligament isoperistaltically. In the sham surgery group, 3 corresponding transections of the intestine were made at the same locations as in IT and reattached in situ. ß cell senescence was examined by the expression of two markers in vivo, p53BP1 and p16. RESULTS: IT did not have a significant effect on body weight and insulin sensitivity, but postprandial insulin secretion was significantly increased. Glucagon-like peptide-1 (GLP-1) and peptide YY secretion were also increased after IT. The histology of the transposed ileum showed distinct hypertrophy with increased GLP-1 positive enteroendocrine cells. Pancreatic ß cell area was significantly increased in the IT group. The percentage of p16 or p53BP1 positive senescent ß cells was significantly lower in the IT group versus the sham group. CONCLUSIONS: IT improved glucose tolerance in diet-induced obese rats mainly through augmented insulin secretion. This improvement was associated with attenuated ß cell senescence.
Asunto(s)
Células Secretoras de Insulina , Obesidad Mórbida , Animales , Glucemia , Senescencia Celular , Prueba de Tolerancia a la Glucosa , Íleon/cirugía , Insulina , Obesidad/cirugía , Obesidad Mórbida/cirugía , Ratas , Ratas Sprague-DawleyRESUMEN
OBJECTIVE: Targeting the glucagon-like peptide-1 receptor with radiolabeled exendin is a very promising method to noninvasively determine the ß cell mass in the pancreas, which is needed to unravel the pathophysiology of type 1 and type 2 diabetes. The present study aimed to explore the effects of both hyperglycemia and insulitis on the uptake of exendin in a spontaneous type 1 diabetes mouse model, nonobese diabetic (NOD) mice. METHODS: NOD mice (n = 75, 7-21 weeks old) were injected intravenously with [111In]In-DTPA-exendin-3, and single-photon emission computed tomography (SPECT) images were acquired 1 h pi. The pancreatic accumulation of [111In]In-DTPA-exendin-3 was quantified in vivo using SPECT and by ex vivo counting and correlated to the ß cell mass (BCM). The influence of insulitis and hyperglycemia on the exendin uptake was assessed. RESULTS: The pancreas could be visualized longitudinally using SPECT. A linear correlation was found between the BCM (%) and pancreatic uptake (%ID/g) as measured by ex vivo counting (Pearson r = 0.64, p < 0.001), which was not affected by either insulitis (Pearson r = 0.66, p = 0.83) or hyperglycemia (Pearson r = 0.57, p = 0.51). Biodistribution and ex vivo autoradiography revealed remaining [111In]In-DTPA-exendin-3 uptake in the pancreas despite total ablation of BCM. CONCLUSIONS: Despite hyperglycemia and severe insulitis, we have found a good correlation between BCM and pancreatic exendin uptake, even in a suboptimal model with relatively high background activity.
Asunto(s)
Diabetes Mellitus Tipo 1/metabolismo , Hiperglucemia/metabolismo , Células Secretoras de Insulina/metabolismo , Péptidos/metabolismo , Tomografía Computarizada de Emisión de Fotón Único/métodos , Animales , Autorradiografía , Diabetes Mellitus Tipo 1/diagnóstico por imagen , Modelos Animales de Enfermedad , Femenino , Inmunohistoquímica , Radioisótopos de Indio/administración & dosificación , Radioisótopos de Indio/química , Radioisótopos de Indio/metabolismo , Inyecciones Intravenosas , Ratones , Ratones Endogámicos NOD , Ácido Pentético/administración & dosificación , Ácido Pentético/química , Ácido Pentético/metabolismo , Péptidos/administración & dosificación , Péptidos/química , Radiofármacos/metabolismo , Distribución TisularRESUMEN
To assess the potential therapeutic effects of adipose tissue-derived mesenchymal stem cells (ASCs) for the treatment of type 2 diabetes (T2D), we compared the phenotype and functionality of ASCs isolated from high-fat diet and streptozotocin (STZ)-induced T2D and the leptin receptor-deficient (db/db) mice with cells from healthy C57BL/6 mice. ASCs from T2D or db/db mice showed similar expression patterns of cellular markers and abilities to differentiate into adipocytes, osteoblasts, and chondrocytes. However, the rate of proliferation was reduced. ASCs from db/db mice secreted less hepatocyte growth factor (HGF). T2D mice receiving a single intravenous injection of T2D or db/db ASCs showed increased insulin sensitivity, reduced inflammation and fat content in adipose tissue and the liver and increased pancreatic ß cell mass through 5 weeks post-infusion. Our data show that, although ASCs from T2D or db/db mice had inferior proliferative capacity compared to cells from healthy controls, improved insulin sensitivity and less ß cell death was seen in T2D mice receiving mesenchymal stem cell (MSC) therapy. This study offers evidence that ASCs from diabetic donors have the potential to be used for cell therapy in the treatment of insulin resistance and T2D.
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Tejido Adiposo/citología , Diabetes Mellitus Tipo 2/terapia , Células Secretoras de Insulina/citología , Trasplante de Células Madre Mesenquimatosas/métodos , Tejido Adiposo/patología , Animales , Células Cultivadas , Diabetes Mellitus Experimental/etiología , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Tipo 2/etiología , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/patología , Dieta Alta en Grasa/efectos adversos , Modelos Animales de Enfermedad , Resistencia a la Insulina , Masculino , Ratones , Ratones Endogámicos C57BL , Receptores de Leptina/deficiencia , EstreptozocinaRESUMEN
Nutrient sensing receptors are key metabolic mediators of responses to dietary and endogenously derived nutrients. These receptors are largely G-protein-coupled receptors (GPCRs) and many are gaining significant interest as drug targets with a potential therapeutic role in metabolic diseases. A distinct subclass of nutrient sensing GPCRs, two short chain fatty acid (SCFA) receptors (FFA2 and FFA3) are uniquely responsive to gut microbiota derived nutrients (such as acetate, propionate, and butyrate). Pharmacological, molecular, and genetic studies have investigated their role in organismal glucose metabolism and recently in pancreatic ß cell biology. Here, we summarize the present knowledge on the role of these receptors as metabolic sensors in ß cell function and physiology, revealing new therapeutic opportunities for type 2 diabetes.
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Diabetes Mellitus Tipo 2/metabolismo , Células Secretoras de Insulina/metabolismo , Animales , Microbioma Gastrointestinal/fisiología , Humanos , Receptores Acoplados a Proteínas G/metabolismoRESUMEN
PURPOSE: Copper-64 (Cu-64) and Galium-68 (Ga-68) radiolabeled DO3A and NODA conjugates of exendin-4 were used for preclinical imaging of pancreatic ß cells via targeting of glucagon-like peptide-1 receptor (GLP-1R). PROCEDURES: DO3A-VS- and NODA-VS-tagged Cys(40)exendin-4 (DO3A-VS-Cys(40)-exendin-4 and NODA-VS-Cys(40)-exendin-4, respectively) were labeled with Cu-64 and Ga-68 using standard techniques. Biodistribution and dynamic positron emission tomography (PET) were carried out in normal Sprague-Dawley (SD) rats. Ex vivo autoradiography imaging was conducted with freshly frozen pancreatic thin sections. RESULTS: DO3A-VS- and NODA-VS-Cys(40)-exendin-4 analogues were labeled with Cu-64 and Ga-68 to a specific activity of 518.7 ± 3.7 Ci/mmol (19.19 ± 0.14 TBq/mmol) and radiochemical yield above 98 %. Biodistribution data demonstrated pancreatic uptake of 0.11 ± 0.02 %ID/g for [(64)Cu]DO3A-VS-, 0.14 ± 0.02 %ID/g for [(64)Cu]NODA-VS-, 0.11 ± 0.03 for [(68)Ga]DO3A-VS-, and 0.26 ± 0.03 for [(68)Ga]NODA-VS-Cys(40)-exendin-4. Excess exendin-4 and exendin-(9-39)-amide displaced all four Cu-64 and Ga-68 labeled exendin-4 derivatives in blocking studies. CONCLUSIONS: [(64)Cu]/[(68)Ga]DO3A-VS-Cys(40)- and [(64)Cu]/[(68)Ga]NODA-VS-Cys(40)-exendin-4 can be used as PET imaging agents specific for GLP-1R expressed on ß cells. Here, we report the first evidence of pancreatic uptake visualized with exendin-4 derivative in a rat animal model via in vivo dynamic PET imaging.
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Radioisótopos de Cobre/metabolismo , Radioisótopos de Galio/metabolismo , Receptor del Péptido 1 Similar al Glucagón/agonistas , Células Secretoras de Insulina/diagnóstico por imagen , Imagen Molecular/métodos , Tomografía de Emisión de Positrones/métodos , Radiofármacos/metabolismo , Animales , Autorradiografía , Exenatida , Receptor del Péptido 1 Similar al Glucagón/metabolismo , Compuestos Heterocíclicos con 1 Anillo/química , Masculino , Péptidos/química , Ratas Sprague-Dawley , Procesamiento de Señales Asistido por Computador , Distribución Tisular , Tomografía Computarizada por Rayos X , Ponzoñas/química , Compuestos de Vinilo/químicaRESUMEN
Protein kinase Bα (PKBα)/AKT1 and PKBß/AKT2 are required for normal peripheral insulin action but their role in pancreatic ß cells remains enigmatic as indicated by the relatively mild islet phenotype of mice with deficiency for either one of these two isoforms. In this study we have analysed proliferation, apoptosis, ß cell size and glucose-stimulated insulin secretion in human islets overexpressing either PKBα or PKBß. Our results reveal redundant and specific functions. Overexpression of either isoform resulted in increased ß cell size, but insulin production and secretion remained unchanged. Proliferation and apoptosis of ß cells were only significantly stimulated and inhibited, respectively, by PKBα/AKT1. Importantly, overexpression of PKBα/AKT1 in dissociated islets increased the ratio of ß cells to non-ß cells. These results confirm our previous findings obtained with rodent islets and strongly indicate that PKBα/AKT1 can regulate ß cell mass also in human islets.
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Células Secretoras de Insulina/enzimología , Proteínas Proto-Oncogénicas c-akt/fisiología , Apoptosis , Proliferación Celular , Tamaño de la Célula , Células Cultivadas , Humanos , Insulina/metabolismo , Secreción de Insulina , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/fisiologíaRESUMEN
Gi-GPCRs, G protein-coupled receptors that signal via Gα proteins of the i/o class (Gαi/o), acutely regulate cellular behaviors widely in mammalian tissues, but their impact on the development and growth of these tissues is less clear. For example, Gi-GPCRs acutely regulate insulin release from pancreatic ß cells, and variants in genes encoding several Gi-GPCRs--including the α-2a adrenergic receptor, ADRA2A--increase the risk of type 2 diabetes mellitus. However, type 2 diabetes also is associated with reduced total ß-cell mass, and the role of Gi-GPCRs in establishing ß-cell mass is unknown. Therefore, we asked whether Gi-GPCR signaling regulates ß-cell mass. Here we show that Gi-GPCRs limit the proliferation of the insulin-producing pancreatic ß cells and especially their expansion during the critical perinatal period. Increased Gi-GPCR activity in perinatal ß cells decreased ß-cell proliferation, reduced adult ß-cell mass, and impaired glucose homeostasis. In contrast, Gi-GPCR inhibition enhanced perinatal ß-cell proliferation, increased adult ß-cell mass, and improved glucose homeostasis. Transcriptome analysis detected the expression of multiple Gi-GPCRs in developing and adult ß cells, and gene-deletion experiments identified ADRA2A as a key Gi-GPCR regulator of ß-cell replication. These studies link Gi-GPCR signaling to ß-cell mass and diabetes risk and identify it as a potential target for therapies to protect and increase ß-cell mass in patients with diabetes.
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Proliferación Celular , Diabetes Mellitus Tipo 2/metabolismo , Subunidades alfa de la Proteína de Unión al GTP Gi-Go/metabolismo , Células Secretoras de Insulina/metabolismo , Receptores Adrenérgicos alfa 2/metabolismo , Transducción de Señal , Animales , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/patología , Subunidades alfa de la Proteína de Unión al GTP Gi-Go/genética , Glucosa/genética , Glucosa/metabolismo , Células Secretoras de Insulina/patología , Ratones , Ratones Transgénicos , Receptores Adrenérgicos alfa 2/genéticaRESUMEN
Induction of ß-cell mass regeneration is a potentially curative treatment for diabetes. We have recently found that long-term gastrin treatment results in improved metabolic control and ß-cell mass expansion in 95% pancreatectomised (Px) rats. In this study, we investigated the underlying mechanisms of gastrin-induced ß-cell mass expansion after Px. After 90%-Px, rats were treated with gastrin (Px+G) or vehicle (Px+V), pancreatic remnants were harvested on days 1, 3, 5, 7, and 14 and used for gene expression, protein immunolocalisation and morphometric analyses. Gastrin- and vehicle-treated Px rats showed similar blood glucose levels throughout the study. Initially, after Px, focal areas of regeneration, showing mesenchymal cells surrounding ductal structures that expressed the cholecystokinin B receptor, were identified. These focal areas of regeneration were similar in size and cell composition in the Px+G and Px+V groups. However, in the Px+G group, the ductal structures showed lower levels of keratin 20 and ß-catenin (indicative of duct dedifferentiation) and higher levels of expression of neurogenin 3 and NKX6-1 (indicative of endocrine progenitor phenotype), as compared with Px+V rats. In Px+G rats, ß-cell mass and the number of scattered ß-cells were significantly increased compared with Px+V rats, whereas ß-cell replication and apoptosis were similar in the two groups. These results indicate that gastrin treatment-enhanced dedifferentiation and reprogramming of regenerative ductal cells in Px rats, increased ß-cell neogenesis and fostered ß-cell mass expansion.
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Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Gastrinas/farmacología , Células Secretoras de Insulina/citología , Pancreatectomía/métodos , Conductos Pancreáticos/citología , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Expresión Génica/efectos de los fármacos , Proteínas de Homeodominio/genética , Hormonas/farmacología , Inmunohistoquímica , Células Secretoras de Insulina/metabolismo , Masculino , Microscopía Confocal , Proteínas del Tejido Nervioso/genética , Páncreas/citología , Páncreas/metabolismo , Páncreas/fisiología , Conductos Pancreáticos/metabolismo , Ratas , Ratas Sprague-Dawley , Receptor de Colecistoquinina B/metabolismo , Regeneración/efectos de los fármacos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de TiempoRESUMEN
In a previous study, we observed that multiple low doses of streptozotocin (STZ) can induce spontaneous restoration of normoglycemia in adult mouse diabetes models. In the present study, we sought to identify when spontaneous recovery from diabetes occurs and to disclose the changes in the diabetic data of diabetic mice induced by multiple low doses of streptozotocin (MLD-STZ mice). After inducing type 1 diabetes mellitus, radioimmunoassay, indirect immunofluorescence and hematoxylin-eosin staining were used to determine the diabetic data of MLD-STZ mice. In MLD-STZ mice, the diabetic indicators, including food intake, water consumption, body weight, blood glucose level and serum insulin concentration, developed gradually until week 20. Thereafter, the symptoms of diabetes gradually improved. By the week 36, although body weight and ß cell masses remained significantly different between the MLD-STZ mice and the age-matched control animals, food intake, water consumption, blood glucose levels and serum insulin concentrations had all returned to normal levels, and no lymphocytic infiltrations were observed in pancreatic islets. This data demonstrates that MLD-STZ can induce spontaneous recovery from diabetes mellitus in adult mice, suggesting further research into the processes by which normoglycemia is recovered.