Nrf2 and Antioxidant Response in Animal Models of Type 2 Diabetes.

This perspective examines the proposition that chronically elevated blood glucose levels caused by type 2 diabetes (T2D) harm body tissues by locally generating reactive oxygen species (ROS). A feed-forward scenario is described in which the initial onset of defective beta cell function T2D becomes sustained and causes chronic elevations in blood glucose, which flood metabolic pathways throughout the body, giving rise to abnormally high local levels of ROS. Most cells can defend themselves via a full complement of antioxidant enzymes that are activated by ROS. However, the beta cell itself does not contain catalase or glutathione peroxidases and thereby runs a greater risk of ROS-induced damage. In this review, previously published experiments are revisited to examine the concept that chronic hyperglycemia can lead to oxidative stress in the beta cell, how this relates to the absence of beta cell glutathione peroxidase (GPx) activity, and whether this deficiency might be ameliorated by genetic enrichment of beta cell GPx and by oral antioxidants, including ebselen, a GPx mimetic.

The demise of islet allotransplantation in the United States: A call for an urgent regulatory update.

Islet allotransplantation in the United States (US) is facing an imminent demise. Despite nearly three decades of progress in the field, an archaic regulatory framework has stymied US clinical practice. Current regulations do not reflect the state-of-the-art in clinical or technical practices. In the US, islets are considered biologic drugs and "more than minimally manipulated" human cell and tissue products (HCT/Ps). In contrast, across the world, human islets are appropriately defined as "minimally manipulated tissue" and not regulated as a drug, which has led to islet allotransplantation (allo-ITx) becoming a standard-of-care procedure for selected patients with type 1 diabetes mellitus. This regulatory distinction impedes patient access to islets for transplantation in the US. As a result only 11 patients underwent allo-ITx in the US between 2016 and 2019, and all as investigational procedures in the settings of a clinical trials. Herein, we describe the current regulations pertaining to islet transplantation in the United States. We explore the progress which has been made in the field and demonstrate why the regulatory framework must be updated to both better reflect our current clinical practice and to deal with upcoming challenges. We propose specific updates to current regulations which are required for the renaissance of ethical, safe, effective, and affordable allo-ITx in the United States.

Factors Predicting Outcomes After a Total Pancreatectomy and Islet Autotransplantation Lessons Learned From Over 500 Cases.

OBJECTIVE: Our objective was to analyze factors predicting outcomes after a total pancreatectomy and islet autotransplantation (TP-IAT). BACKGROUND: Chronic pancreatitis (CP) is increasingly treated by a TP-IAT. Postoperative outcomes are generally favorable, but a minority of patients fare poorly. METHODS: In our single-centered study, we analyzed the records of 581 patients with CP who underwent a TP-IAT. Endpoints included persistent postoperative "pancreatic pain" similar to preoperative levels, narcotic use for any reason, and islet graft failure at 1 year. RESULTS: In our patients, the duration (mean ±â€ŠSD) of CP before their TP-IAT was 7.1 ±â€Š0.3 years and narcotic usage of 3.3 ±â€Š0.2 years. Pediatric patients had better postoperative outcomes. Among adult patients, the odds of narcotic use at 1 year were increased by previous endoscopic retrograde cholangiopancreatography (ERCP) and stent placement, and a high number of previous stents (>3). Independent risk factors for pancreatic pain at 1 year were pancreas divisum, previous body mass index >30, and a high number of previous stents (>3). The strongest independent risk factor for islet graft failure was a low islet yield-in islet equivalents (IEQ)-per kilogram of body weight. We noted a strong dose-response relationship between the lowest-yield category (<2000 IEQ) and the highest (≥5000 IEQ or more). Islet graft failure was 25-fold more likely in the lowest-yield category. CONCLUSIONS: This article represents the largest study of factors predicting outcomes after a TP-IAT. Preoperatively, the patient subgroups we identified warrant further attention.

Arginine is preferred to glucagon for stimulation testing of ß-cell function.

A key aspect of research into the prevention and treatment of type 2 diabetes is the availability of reproducible clinical research methodology to assess ß-cell function. One commonly used method employs nonglycemic secretagogues like arginine (arg) or glucagon (glgn). This study was designed to quantify the insulin response to arg and glgn and determine test repeatability and tolerability. Obese overnight-fasted subjects with normal glucose tolerance were studied on 4 separate days: twice using arg (5 g iv) and twice with glgn (1 mg iv). Pre- and postinfusion samples for plasma glucose, insulin, and C-peptide were acquired. Arg and glgn challenges were repeated in the last 10 min of a 60-min glucose (900 mg/min) infusion. Insulin and C-peptide secretory responses were estimated under baseline fasting glucose conditions (AIRarg and AIRglgn) and hyperglycemic (AIRargMAX AIRglgnMAX) states. Relative repeatability was estimated by intraclass correlation coefficient (ICC). Twenty-three (12 men and 11 women) subjects were studied (age: 42.4 ± 8.3 yr; BMI: 31.4 ± 2.8 kg/m²). Geometric means (95% CI) for baseline-adjusted values AIRarg and AIRglgn were 84 (75-95) and 102 (90-115) µU/ml, respectively. After the glucose infusion, AIRargMAX and AIRglgnMAX were 395 (335-466) and 483 (355-658) µU/ml, respectively. ICC values were >0.90 for AIRarg andAIRargMAX. Glucagon ICCs were 0.83, 0.34, and 0.36, respectively, although the exclusion of one outlier increased the latter two values (to 0.84 and 0.86). Both glgn and arg induced mild adverse events that were transient. Glucagon, but not arginine, induced moderate adverse events due to nausea. Taken together, arginine is preferred to glucagon for assessment of ß-cell function.

Total pancreatectomy and islet autotransplantation in chronic pancreatitis: recommendations from PancreasFest.

DESCRIPTION: Total pancreatectomy with islet autotransplantation (TPIAT) is a surgical procedure used to treat severe complications of chronic pancreatitis or very high risk of pancreatic cancer while reducing the risk of severe diabetes mellitus. However, clear guidance on indications, contraindications, evaluation, timing, and follow-up are lacking. METHODS: A working group reviewed the medical, psychological, and surgical options and supporting literature related to TPIAT for a consensus meeting during PancreasFest. RESULTS: Five major areas requiring clinical evaluation and management were addressed: These included: 1) indications for TPIAT; 2) contraindications for TPIAT; 3) optimal timing of the procedure; 4) need for a multi-disciplinary team and the roles of the members; 5) life-long management issues following TPIAP including diabetes monitoring and nutrition evaluation. CONCLUSIONS: TPIAT is an effective method of managing the disabling complications of chronic pancreatitis and risk of pancreatic cancer in very high risk patients. Careful evaluation and long-term management of candidate patients by qualified multidisciplinary teams is required. Multiple recommendations for further research were also identified.

Antioxidants for Early Treatment of Type 2 Diabetes in Rodents and Humans: Lost in Translation?

Reactive oxygen species (ROS) are formed by virtually all tissues. In normal concentrations they facilitate many physiologic activities, but in excess they cause oxidative stress and tissue damage. Local antioxidant enzyme synthesis in cells is regulated by the cytoplasmic KEAP-1/Nrf2 complex, which is stimulated by ROS, to release Nrf2 for entry into the nucleus, where it upregulates antioxidant gene expression. Major antioxidant enzymes include glutathione peroxidase (GPx), catalase (CAT), superoxide dismutases (SOD), hemoxygenases (HO), and peroxiredoxins (Prdx). Notably, the pancreatic islet ß-cell does not express GPx or CAT, which puts it at greater risk for ROS damage caused by postprandial hyperglycemia. Experimentally, overexpression of GPx in ß-cell lines and isolated islets, as well as in vivo studies using genetic models of type 2 diabetes (T2D), has demonstrated enhanced protection against hyperglycemia and oxidative stress. Oral treatment of diabetic rodents with ebselen, a GPx mimetic that is approved for human clinical use, reproduced these findings. Prdx detoxify hydrogen peroxide and reduce lipid peroxides. This suggests that pharmacologic development of more potent, ß-cell-specific antioxidants could be valuable as a treatment for oxidative stress due to postprandial hyperglycemia in early T2D in humans.

Cyclooxygenase-2, not microsomal prostaglandin E synthase-1, is the mechanism for interleukin-1ß-induced prostaglandin E2 production and inhibition of insulin secretion in pancreatic islets.

Arachidonic acid is converted to prostaglandin E(2) (PGE(2)) by a sequential enzymatic reaction performed by two isoenzyme groups, cyclooxygenases (COX-1 and COX-2) and terminal prostaglandin E synthases (cPGES, mPGES-1, and mPGES-2). mPGES-1 is widely considered to be the final enzyme regulating COX-2-dependent PGE(2) synthesis. These generalizations have been based in most part on experiments utilizing gene expression analyses of cell lines and tumor tissue. To assess the relevance of these generalizations to a native mammalian tissue, we used isolated human and rodent pancreatic islets to examine interleukin (IL)-1ß-induced PGE(2) production, because PGE(2) has been shown to mediate IL-1ß inhibition of islet function. Rat islets constitutively expressed mRNAs of COX-1, COX-2, cPGES, and mPGES-1. As expected, IL-1ß increased mRNA levels for COX-2 and mPGES-1, but not for COX-1 or cPGES. Basal protein levels of COX-1, cPGES, and mPGES-2 were readily detected in whole cell extracts but were not regulated by IL-1ß. IL-1ß increased protein levels of COX-2, but unexpectedly mPGES-1 protein levels were low and unaffected. In microsomal extracts, mPGES-1 protein was barely detectable in rat islets but clearly present in human islets; however, in neither case did IL-1ß increase mPGES-1 protein levels. To further assess the importance of mPGES-1 to IL-1ß regulation of an islet physiologic response, glucose-stimulated insulin secretion was examined in isolated islets of WT and mPGES-1-deficient mice. IL-1ß inhibited glucose-stimulated insulin secretion equally in both WT and mPGES-1(-/-) islets, indicating that COX-2, not mPGES-1, mediates IL-1ß-induced PGE(2) production and subsequent inhibition of insulin secretion.

Detection, evaluation and treatment of diabetes mellitus in chronic pancreatitis: recommendations from PancreasFest 2012.

DESCRIPTION: Diabetes and glucose intolerance are common complications of chronic pancreatitis, yet clinical guidance on their detection, classification, and management is lacking. METHODS: A working group reviewed the medical problems, diagnostic methods, and treatment options for chronic pancreatitis-associated diabetes for a consensus meeting at PancreasFest 2012. RESULTS: Guidance Statement 1.1: Diabetes mellitus is common in chronic pancreatitis. While any patient with chronic pancreatitis should be monitored for development of diabetes, those with long-standing duration of disease, prior partial pancreatectomy, and early onset of calcific disease may be at higher risk. Those patients developing diabetes mellitus are likely to have co-existing pancreatic exocrine insufficiency. Guidance Statement 1.2: Diabetes occurring secondary to chronic pancreatitis should be recognized as pancreatogenic diabetes (type 3c diabetes). Guidance Statement 2.1: The initial evaluation should include fasting glucose and HbA1c. These tests should be repeated annually. Impairment in either fasting glucose or HbA1c requires further evaluation. Guidance Statement 2.2: Impairment in either fasting glucose or HbA1c should be further evaluated by a standard 75 g oral glucose tolerance test. Guidance Statement 2.3: An absent pancreatic polypeptide response to mixed-nutrient ingestion is a specific indicator of type 3c diabetes. Guidance Statement 2.4: Assessment of pancreatic endocrine reserve, and importantly that of functional beta-cell mass, should be performed as part of the evaluation and follow-up for total pancreatectomy with islet autotransplantation (TPIAT). Guidance Statement 3: Patients with pancreatic diabetes shall be treated with specifically tailored medical nutrition and pharmacologic therapies. CONCLUSIONS: Physicians should evaluate and treat glucose intolerance in patients with pancreatitis.

The B55α-containing PP2A holoenzyme dephosphorylates FOXO1 in islet ß-cells under oxidative stress.

The FOXO1 (forkhead box O1) transcription factor influences many key cellular processes, including those important in metabolism, proliferation and cell death. Reversible phosphorylation of FOXO1 at Thr(24) and Ser(256) regulates its subcellular localization, with phosphorylation promoting cytoplasmic localization, whereas dephosphorylation triggers nuclear import and transcriptional activation. In the present study, we used biochemical and molecular approaches to isolate and link the serine/threonine PP2A (protein phosphatase 2A) holoenzyme containing the B55α regulatory subunit, with nuclear import of FOXO1 in pancreatic islet ß-cells under oxidative stress, a condition associated with cellular dysfunction in Type 2 diabetes. The mechanism of FOXO1 dephosphorylation and nuclear translocation was investigated in pancreatic islet INS-1 and ßTC-3 cell lines subjected to oxidative stress. A combined chemical cross-linking and MS strategy revealed the association of FOXO1 with a PP2A holoenzyme composed of the catalytic C, structural A and B55α regulatory subunits. Knockdown of B55α in INS-1 cells reduced FOXO1 dephosphorylation, inhibited FOXO1 nuclear translocation and attenuated oxidative stress-induced cell death. Furthermore, both B55α and nuclear FOXO1 levels were increased under hyperglycaemic conditions in db/db mouse islets, an animal model of type 2 diabetes. We conclude that B55α-containing PP2A is a key regulator of FOXO1 activity in vivo.

Brief overview: glucagon history and physiology.

Glucagon is a peptide hormone that is produced primarily by the alpha cells in the islet of Langerhans in the pancreas, but also in intestinal enteroendocrine cells and in some neurons. Approximately 100 years ago, several research groups discovered that pancreatic extracts would cause a brief rise in blood glucose before they observed the decrease in glucose attributed to insulin. An overall description of the regulation of glucagon secretion requires the inclusion of its sibling insulin because they both are made primarily by the islet and they both regulate each other in different ways. For example, glucagon stimulates insulin secretion, whereas insulin suppresses glucagon secretion. The mechanism of action of glucagon on insulin secretion has been identified as a trimeric guanine nucleotide-binding protein (G-protein)-mediated event. The manner in which insulin suppresses glucagon release from the alpha cell is thought to be highly dependent on the peri-portal circulation of the islet through which blood flows downstream from beta cells to alpha cells. In this scenario, it is via the circulation that insulin is thought to suppress the release of glucagon. However, high levels of glucose also have been shown to suppress glucagon secretion. Consequently, the glucose-lowering effect of insulin may be additive to the direct effects of insulin to suppress alpha cell function, so that in vivo both the discontinuation of the insulin signal and the condition of low glucose jointly are responsible for induction of glucagon secretion.

Evolution of Nrf2 Gene Expression in HIT-T15 ß-Cells During Chronic Oxidative Stress and Glucose Toxicity.

Context: Chronic exposure of pancreatic islets to elevated glucose levels causes progressive declines in beta cell Pdx-1 and insulin gene expression, and glucose-induced insulin secretion. This has been shown to be associated with excessive islet reactive oxygen species and consequent damage to beta cell function, a process termed glucose toxicity. In short-term rodent in vivo studies, Nrf2 (Kelch-like ECH-associated protein 1:nuclear factor erythroid-derived-2 related factor complex) has been shown to play a central role in defending beta cells from oxidative damage via activation of antioxidant gene expression. Objective: The current studies were primarily designed to examine the behavior of Nrf2 gene expression during longer term exposure of beta cells to glucose toxicity. Methods and Results: We provide evidence that gene expression of Nrf2 in HIT-T15 cells, an insulin-secreting beta-cell line, undergoes a biphasic response characterized by an initial decrease followed by increased expression during prolonged culturing of these cells in a physiologic (0.8 mM) but not a supraphysiologic (16.0 mM) glucose concentration. This was associated with a slight rise in HO-1 gene expression. Pdx-1 and insulin mRNA levels also decreased but then stabilized in late passages of cells that had been cultured in low glucose concentrations. Conclusion: These complex events support the concept that Nrf2 gene expression plays an important regulatory role in defending beta cells during prolonged exposure to oxidative stress.

Glucolipotoxicity: fuel excess and beta-cell dysfunction.

Glucotoxicity, lipotoxicity, and glucolipotoxicity are secondary phenomena that are proposed to play a role in all forms of type 2 diabetes. The underlying concept is that once the primary pathogenesis of diabetes is established, probably involving both genetic and environmental forces, hyperglycemia and very commonly hyperlipidemia ensue and thereafter exert additional damaging or toxic effects on the beta-cell. In addition to their contribution to the deterioration of beta-cell function after the onset of the disease, elevations of plasma fatty acid levels that often accompany insulin resistance may, as glucose levels begin to rise outside of the normal range, also play a pathogenic role in the early stages of the disease. Because hyperglycemia is a prerequisite for lipotoxicity to occur, the term glucolipotoxicity, rather than lipotoxicity, is more appropriate to describe deleterious effects of lipids on beta-cell function. In vitro and in vivo evidence supporting the concept of glucotoxicity is presented first, as well as a description of the underlying mechanisms with an emphasis on the role of oxidative stress. Second, we discuss the functional manifestations of glucolipotoxicity on insulin secretion, insulin gene expression, and beta-cell death, and the role of glucose in the mechanisms of glucolipotoxicity. Finally, we attempt to define the role of these phenomena in the natural history of beta-cell compensation, decompensation, and failure during the course of type 2 diabetes.

Islets Transplantation at a Crossroads - Need for Urgent Regulatory Update in the United States: Perspective Presented During the Scientific Sessions 2021 at the American Diabetes Association Congress.

Clinical islet allotransplantation has been successfully regulated as tissue/organ for transplantation in number of countries and is recognized as a safe and efficacious therapy for selected patients with type 1 diabetes mellitus. However, in the United States, the FDA considers pancreatic islets as a biologic drug, and islet transplantation has not yet shifted from the experimental to the clinical arena for last 20 years. In order to transplant islets, the FDA requires a valid Biological License Application (BLA) in place. The BLA process is costly and lengthy. However, despite the application of drug manufacturing technology and regulations, the final islet product sterility and potency cannot be confirmed, even when islets meet all the predetermined release criteria. Therefore, further regulation of islets as drugs is obsolete and will continue to hinder clinical application of islet transplantation in the US. The Organ Procurement and Transplantation Network together with the United Network for Organ Sharing have developed separately from the FDA and BLA regulatory framework for human organs under the Human Resources & Services Administration to assure safety and efficacy of transplantation. Based on similar biologic characteristics of islets and human organs, we propose inclusion of islets into the existing regulatory framework for organs for transplantation, along with continued FDA oversight for islet processing, as it is for other cell/tissue products exempt from BLA. This approach would reassure islet quality, efficacy and access for Americans with diabetes to this effective procedure.

Arguments against the Requirement of a Biological License Application for Human Pancreatic Islets: The Position Statement of the Islets for US Collaborative Presented during the FDA Advisory Committee Meeting.

The Food and Drug Administration (FDA) has been regulating human islets for allotransplantation as a biologic drug in the US. Consequently, the requirement of a biological license application (BLA) approval before clinical use of islet transplantation as a standard of care procedure has stalled the development of the field for the last 20 years. Herein, we provide our commentary to the multiple FDA's position papers and guidance for industry arguing that BLA requirement has been inappropriately applied to allogeneic islets, which was delivered to the FDA Cellular, Tissue and Gene Therapies Advisory Committee on 15 April 2021. We provided evidence that BLA requirement and drug related regulations are inadequate in reassuring islet product quality and potency as well as patient safety and clinical outcomes. As leaders in the field of transplantation and endocrinology under the "Islets for US Collaborative" designation, we examined the current regulatory status of islet transplantation in the US and identified several anticipated negative consequences of the BLA approval. In our commentary we also offer an alternative pathway for islet transplantation under the regulatory framework for organ transplantation, which would address deficiencies of in current system.

International trial of the Edmonton protocol for islet transplantation.

BACKGROUND: Islet transplantation offers the potential to improve glycemic control in a subgroup of patients with type 1 diabetes mellitus who are disabled by refractory hypoglycemia. We conducted an international, multicenter trial to explore the feasibility and reproducibility of islet transplantation with the use of a single common protocol (the Edmonton protocol). METHODS: We enrolled 36 subjects with type 1 diabetes mellitus, who underwent islet transplantation at nine international sites. Islets were prepared from pancreases of deceased donors and were transplanted within 2 hours after purification, without culture. The primary end point was defined as insulin independence with adequate glycemic control 1 year after the final transplantation. RESULTS: Of the 36 subjects, 16 (44%) met the primary end point, 10 (28%) had partial function, and 10 (28%) had complete graft loss 1 year after the final transplantation. A total of 21 subjects (58%) attained insulin independence with good glycemic control at any point throughout the trial. Of these subjects, 16 (76%) required insulin again at 2 years; 5 of the 16 subjects who reached the primary end point (31%) remained insulin-independent at 2 years. CONCLUSIONS: Islet transplantation with the use of the Edmonton protocol can successfully restore long-term endogenous insulin production and glycemic stability in subjects with type 1 diabetes mellitus and unstable control, but insulin independence is usually not sustainable. Persistent islet function even without insulin independence provides both protection from severe hypoglycemia and improved levels of glycated hemoglobin. (ClinicalTrials.gov number, NCT00014911 [ClinicalTrials.gov].).

Pancreatic Islet Transplantation in Humans: Recent Progress and Future Directions.

Pancreatic islet transplantation has become an established approach to ß-cell replacement therapy for the treatment of insulin-deficient diabetes. Recent progress in techniques for islet isolation, islet culture, and peritransplant management of the islet transplant recipient has resulted in substantial improvements in metabolic and safety outcomes for patients. For patients requiring total or subtotal pancreatectomy for benign disease of the pancreas, isolation of islets from the diseased pancreas with intrahepatic transplantation of autologous islets can prevent or ameliorate postsurgical diabetes, and for patients previously experiencing painful recurrent acute or chronic pancreatitis, quality of life is substantially improved. For patients with type 1 diabetes or insulin-deficient forms of pancreatogenic (type 3c) diabetes, isolation of islets from a deceased donor pancreas with intrahepatic transplantation of allogeneic islets can ameliorate problematic hypoglycemia, stabilize glycemic lability, and maintain on-target glycemic control, consequently with improved quality of life, and often without the requirement for insulin therapy. Because the metabolic benefits are dependent on the numbers of islets transplanted that survive engraftment, recipients of autoislets are limited to receive the number of islets isolated from their own pancreas, whereas recipients of alloislets may receive islets isolated from more than one donor pancreas. The development of alternative sources of islet cells for transplantation, whether from autologous, allogeneic, or xenogeneic tissues, is an active area of investigation that promises to expand access and indications for islet transplantation in the future treatment of diabetes.

Thyroid Dysfunction and Diabetes Mellitus: Two Closely Associated Disorders.

Thyroid dysfunction and diabetes mellitus are closely linked. Several studies have documented the increased prevalence of thyroid disorders in patients with diabetes mellitus and vice versa. This review critically discusses the different underlying mechanisms linking type 1 and 2 diabetes and thyroid dysfunction to demonstrate that the association of these two common disorders is unlikely a simple coincidence. We assess the current state of knowledge on the central and peripheral control of thyroid hormone on food intake and glucose and lipid metabolism in target tissues (such as liver, white and brown adipose tissue, pancreatic ß cells, and skeletal muscle) to explain the mechanism linking overt and subclinical hypothyroidism to type 2 diabetes and metabolic syndrome. We also elucidate the common susceptibility genes and the pathogenetic mechanisms contributing to the autoimmune mechanism involved in the onset of type 1 diabetes mellitus and autoimmune thyroid disorders. An untreated thyroid dysfunction can impair the metabolic control of diabetic patients, and this association can have important repercussions on the outcome of both of these disorders. Therefore, we offer recommendations for the diagnosis, management, and screening of thyroid disorders in patients with diabetes mellitus, including the treatment of diabetic patients planning a pregnancy. We also discuss the major causes of failure to achieve an optimal management of thyroid dysfunction in diabetic patients and provide recommendations for assessing and treating these disorders during therapy with antidiabetic drugs. An algorithm for a correct approach of these disorders when linked is also provided.

Zinc, not insulin, regulates the rat alpha-cell response to hypoglycemia in vivo.

The intra-islet insulin hypothesis proposes that the decrement in beta-cell insulin secretion during hypoglycemia provides an activation signal for alpha-cells to release glucagon. A more recent hypothesis proposes that zinc atoms suppress glucagon secretion via their ability to open alpha-cell ATP-sensitive K(+) channels. Since insulin binds zinc, and zinc is co-secreted with insulin, we tested whether decreased zinc delivery to the alpha-cell activates glucagon secretion. In streptozotocin-induced diabetic Wistar rats, we observed that switching off intrapancreatic artery insulin infusions in vivo during hypoglycemia greatly improved glucagon secretion (area under the curve [AUC]: control group 240 +/- 261 and experimental group 4,346 +/- 1,259 pg x ml(-1) x 90 min(-1); n = 5, P < 0.02). Switching off pancreatic artery infusions of zinc chloride during hypoglycemia also improved the glucagon response (AUC: control group 817 +/- 107 and experimental group 3,445 +/- 573 pg x ml(-1) x 90 min(-1); n = 6, P < 0.01). However, switching off zinc-free insulin infusions had no effect. Studies of glucose uptake in muscle and liver cell lines verified that the zinc-free insulin was biologically active. We conclude that zinc atoms, not the insulin molecule itself, provide the switch-off signal from the beta-cell to the alpha-cell to initiate glucagon secretion during hypoglycemia.

Deficient Glucagon Response to Hypoglycemia During a Mixed Meal in Total Pancreatectomy/Islet Autotransplantation Recipients.

Context: Total pancreatectomy and intrahepatic islet autotransplantation (TP/IAT) is performed to alleviate severe abdominal pain, avoid narcotic use, maintain islet function, and avoid diabetes in patients with chronic pancreatitis. However, many TP/IAT recipients complain of postprandial hypoglycemia. Objective: This study was designed to discover the mechanisms of this problem. Design: Participants consumed a triple-isotope mixed meal. Setting: This study was performed in a hospital research unit. Participants: We studied 10 TP/IAT recipients and 10 age- and body mass index-matched control subjects. Seven of 10 recipients had a history of postprandial hypoglycemia. Interventions: Participants were given a [1-13C]-labeled mixed meal and two tracer infusions ([6,6-2H2]- and [6-3H]-glucose). Main Outcome Measures: Glucose kinetics and concentrations of regulatory hormones were determined. Results: Immediately after the meal, peak glucose was elevated in recipients compared with control subjects [266 ± 20 mg/dL (14.8 ± 1.1 mmol/L) vs 185 ± 13 mg/dL (10.3 ± 0.7 mmol/L); P = 0.01]. However, mean Δ glucose for TP/IAT recipients between minutes 240 and 360 postprandially was significantly lower than for control subjects (P < 0.05); six of the seven recipients with a history of hypoglycemia experienced abnormally low postprandial Δ glucose. Δ Glucagon remained unchanged (minutes 240 to 360; P = 0.58) in TP/IAT recipients despite abnormal decreases in postprandial glucose. Radioisotopic studies revealed that meal appearance, glucose disappearance, and endogenous glucose production in TP/IAT recipients were not different from control subjects. Conclusion: Initially high glucose levels followed by hypoglycemia with an absent glucagon response is a mechanistic sequence that contributes to postprandial hypoglycemia after TP/IAT.

Mixed Meal and Intravenous L-Arginine Tests Both Stimulate Incretin Release Across Glucose Tolerance in Man: Lack of Correlation with ß Cell Function.

BACKGROUND: The aims of this study were to 1. define the responses of glucagon-like peptide-1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP), glucagon, and peptide YY (PYY) to an oral meal and to intravenous L-arginine; and 2. examine correlation of enteroendocrine hormones with insulin secretion. We hypothesized a relationship between circulating incretin concentrations and insulin secretion. METHODS: Subjects with normal glucose tolerance (NGT, n = 23), prediabetes (PDM, n = 17), or with type 2 diabetes (T2DM, n = 22) were studied twice, following a mixed test meal (470 kCal) (mixed meal tolerance test [MMTT]) or intravenous L-arginine (arginine maximal stimulation test [AST], 5 g). GLP-1 (total and active), PYY, GIP, glucagon, and ß cell function were measured before and following each stimulus. RESULTS: Baseline enteroendocrine hormones differed across the glucose tolerance (GT) spectrum, T2DM generally >NGT and PDM. In response to MMTT, total and active GLP-1, GIP, glucagon, and PYY increased in all populations. The incremental area-under-the-curve (0-120 min) of analytes like total GLP-1 were often higher in T2DM compared with NGT and PDM (35-51%; P < 0.05). At baseline glucose, L-arginine increased total and active GLP-1 and glucagon concentrations in all GT populations (all P < 0.05). As expected, the MMTT and AST provoked differential glucose, insulin, and C-peptide responses across GT populations. Baseline or stimulated enteroendocrine hormone concentrations did not consistently correlate with either measure of ß cell function. CONCLUSIONS/INTERPRETATION: Both MMTT and AST resulted in insulin and enteroendocrine hormone responses across GT populations without consistent correlation between release of incretins and insulin, which is in line with other published research. If a defect is in the enteroendocrine/ß cell axis, it is probably reduced response to rather than diminished secretion of enteroendocrine hormones.