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Controversy exists regarding the potential regenerative influences of incretin therapy on pancreatic β-cells versus possible adverse pancreatic proliferative effects. Examination of pancreata from age-matched organ donors with type 2 diabetes mellitus (DM) treated by incretin therapy (n = 8) or other therapy (n = 12) and nondiabetic control subjects (n = 14) reveals an ∼40% increased pancreatic mass in DM treated with incretin therapy, with both increased exocrine cell proliferation (P < 0.0001) and dysplasia (increased pancreatic intraepithelial neoplasia, P < 0.01). Pancreata in DM treated with incretin therapy were notable for α-cell hyperplasia and glucagon-expressing microadenomas (3 of 8) and a neuroendocrine tumor. β-Cell mass was reduced by ∼60% in those with DM, yet a sixfold increase was observed in incretin-treated subjects, although DM persisted. Endocrine cells costaining for insulin and glucagon were increased in DM compared with non-DM control subjects (P < 0.05) and markedly further increased by incretin therapy (P < 0.05). In conclusion, incretin therapy in humans resulted in a marked expansion of the exocrine and endocrine pancreatic compartments, the former being accompanied by increased proliferation and dysplasia and the latter by α-cell hyperplasia with the potential for evolution into neuroendocrine tumors.

Pancreatic duct glands (PDGs) have been hypothesized to give rise to pancreatic intraepithelial neoplasia (PanIN). Treatment with the glucagon-like peptide (GLP)-1 analog, exendin-4, for 12 weeks induced the expansion of PDGs with mucinous metaplasia and columnar cell atypia resembling low-grade PanIN in rats. In the pancreata of Pdx1-Cre; LSL-KrasG12D mice, exendin-4 led to acceleration of the disruption of exocrine architecture and chronic pancreatitis with mucinous metaplasia and increased formation of murine PanIN lesions. PDGs and PanIN lesions in rodent and human pancreata express the GLP-1 receptor. Exendin-4 induced proproliferative signaling pathways in human pancreatic duct cells, cAMP–protein kinase A and mitogen-activated protein kinase phosphorylation of cAMP-responsive element-binding protein, and increased cyclin D1 expression. These GLP-1 effects were more pronounced in the presence of an activating mutation of Kras and were inhibited by metformin. These data reveal that GLP-1 mimetic therapy may induce focal proliferation in the exocrine pancreas and, in the context of exocrine dysplasia, may accelerate formation of neoplastic PanIN lesions and exacerbate chronic pancreatitis.

OBJECTIVE Cyclin-dependent kinase 5 (CDK5) regulatory subunit-associated protein 1-like 1 has recently been linked to type 2 diabetes by genome-wide association studies. While CDK5 and its regulatory protein p35 are both expressed and display enzymatic activity in pancreatic β-cells, their precise role in the β-cell remains unknown. Because type 2 diabetes is characterized by a deficit in β-cell mass and increased β-cell apoptosis, we investigated the role of CDK5 in β-cell survival. RESEARCH DESIGN AND METHODS We used INS 832/13 cells, rat islets isolated from wild-type or human islet amyloid polypeptide (h-IAPP) transgenic rats, and pancreatic tissue from rats and humans with and without type 2 diabetes and investigated the effect of CDK5/p35 inhibition (by small interfering RNA or by chemical inhibition) as well as CDK5/p35 overexpression on β-cell vulnerability to apoptosis. RESULTS CDK5 inhibition led to increased β-cell apoptosis. To identify the mechanisms involved, we examined the phosphorylation state of focal adhesion kinase (Fak)Ser732, a known target of CDK5. Following CDK5 inhibition, the phosphorylation of FakSer732 decreased with resulting attenuation of phosphatidylinositol 3-kinase (PI3K)/Akt survival pathway. Conversely, CDK5 overexpression increased FakSer732 phosphorylation and protected β-cells against apoptosis induced by the inhibition of the β-1 integrin signaling pathway. Also, FakSer732 phosphorylation was less abundant in β-cells in both h-IAPP transgenic rats and humans with type 2 diabetes. CONCLUSIONS This study shows that by regulating Fak phosphorylation and subsequently PI3K/Akt survival pathway, CDK5 plays a previously unrecognized role in promoting β-cell survival.

OBJECTIVE The islet in type 2 diabetes is characterized by β-cell apoptosis, β-cell endoplasmic reticulum stress, and islet amyloid deposits derived from islet amyloid polypeptide (IAPP). Toxic oligomers of IAPP form intracellularly in β-cells in humans with type 2 diabetes, suggesting impaired clearance of misfolded proteins. In this study, we investigated whether human-IAPP (h-IAPP) disrupts the endoplasmic reticulum–associated degradation/ubiquitin/proteasome system. RESEARCH DESIGN AND METHODS We used pancreatic tissue from humans with and without type 2 diabetes, isolated islets from h-IAPP transgenic rats, isolated human islets, and INS 832/13 cells transduced with adenoviruses expressing either h-IAPP or a comparable expression of rodent-IAPP. Immunofluorescence and Western blotting were used to detect polyubiquitinated proteins and ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1) protein levels. Proteasome activity was measured in isolated rat and human islets. UCH-L1 was knocked down by small-interfering RNA in INS 832/13 cells and apoptosis was evaluated. RESULTS We report accumulation of polyubiquinated proteins and UCH-L1 deficiency in β-cells of humans with type 2 diabetes. These findings were reproduced by expression of oligomeric h-IAPP but not soluble rat-IAPP. Downregulation of UCH-L1 expression and activity to reproduce that caused by h-IAPP in β-cells induced endoplasmic reticulum stress leading to apoptosis. CONCLUSIONS Our results indicate that defective protein degradation in β-cells in type 2 diabetes can, at least in part, be attributed to misfolded h-IAPP leading to UCH-L1 deficiency, which in turn further compromises β-cell viability.

OBJECTIVE We sought to establish the extent and mechanisms by which sitagliptin and metformin singly and in combination modify islet disease progression in human islet amyloid polypeptide transgenic (HIP) rats, a model for type 2 diabetes. RESEARCH DESIGN AND METHODS HIP rats were treated with sitagliptin, metformin, sitagliptin plus metformin, or no drug as controls for 12 weeks. Fasting blood glucose, insulin sensitivity, and β-cell mass, function, and turnover were measured in each group. RESULTS Sitagliptin plus metformin had synergistic effects to preserve β-cell mass in HIP rats. Metformin more than sitagliptin inhibited β-cell apoptosis. Metformin enhanced hepatic insulin sensitivity; sitagliptin enhanced extrahepatic insulin sensitivity with a synergistic effect in combination. β-Cell function was partially preserved by sitagliptin plus metformin. However, sitagliptin treatment was associated with increased pancreatic ductal turnover, ductal metaplasia, and, in one rat, pancreatitis. CONCLUSIONS The combination of metformin and sitagliptin had synergistic actions to preserve β-cell mass and function and enhance insulin sensitivity in the HIP rat model of type 2 diabetes. However, adverse actions of sitagliptin treatment on exocrine pancreas raise concerns that require further evaluation.

OBJECTIVE Obesity is a known risk factor for type 2 diabetes. However, most obese individuals do not develop diabetes because they adapt to insulin resistance by increasing β-cell mass and insulin secretion. Islet pathology in type 2 diabetes is characterized by β-cell loss, islet amyloid derived from islet amyloid polypeptide (IAPP), and increased β-cell apoptosis characterized by endoplasmic reticulum (ER) stress. We hypothesized that IAPP-induced ER stress distinguishes successful versus unsuccessful islet adaptation to insulin resistance. RESEARCH DESIGN AND METHODS To address this, we fed wild-type (WT) and human IAPP transgenic (HIP) rats either 10 weeks of regular chow or a high-fat diet and prospectively examined the relations among β-cell mass and turnover, β-cell ER stress, insulin secretion, and insulin sensitivity. RESULTS A high-fat diet led to comparable insulin resistance in WT and HIP rats. WT rats compensated with increased insulin secretion and β-cell mass. In HIP rats, in contrast, neither β-cell function nor mass compensated for the increased insulin demand, leading to diabetes. The failure to increase β-cell mass in HIP rats was the result of ER stress–induced β-cell apoptosis that increased in proportion to diet-induced insulin resistance. CONCLUSIONS IAPP-induced ER stress distinguishes the successful versus unsuccessful islet adaptation to a high-fat diet in rats. These studies are consistent with the hypothesis that IAPP oligomers contribute to increased β-cell apoptosis and β-cell failure in humans with type 2 diabetes.

OBJECTIVE—Endoplasmic reticulum (ER) stress–induced apoptosis may be a common cause of cell attrition in diseases characterized by misfolding and oligomerisation of amyloidogenic proteins. The islet in type 2 diabetes is characterized by islet amyloid derived from islet amyloid polypeptide (IAPP) and increased β-cell apoptosis. We questioned the following: 1) whether IAPP-induced β-cell apoptosis is mediated by ER stress and 2) whether β-cells in type 2 diabetes are characterized by ER stress. RESEARCH DESIGN AND METHODS—The mechanism of IAPP-induced apoptosis was investigated in INS-1 cells and human IAPP (HIP) transgenic rats. ER stress in humans was investigated by β-cell C/EBP homologous protein (CHOP) expression in 7 lean nondiabetic, 12 obese nondiabetic, and 14 obese type 2 diabetic human pancreata obtained at autopsy. To assure specificity for type 2 diabetes, we also examined pancreata from eight cases of type 1 diabetes. RESULTS—IAPP induces β-cell apoptosis by ER stress in INS-1 cells and HIP rats. Perinuclear CHOP was rare in lean nondiabetic (2.6 ± 2.0%) and more frequent in obese nondiabetic (14.6 ± 3.0%) and obese diabetic (18.5 ± 3.6%) pancreata. Nuclear CHOP was not detected in lean nondiabetic and rare in obese nondiabetic (0.08 ± 0.04%) but six times higher (P < 0.01) in obese diabetic (0.49 ± 0.17%) pancreata. In type 1 diabetic pancreata, perinuclear CHOP was rare (2.5 ± 2.3%) and nuclear CHOP not detected. CONCLUSIONS—ER stress is a mechanism by which IAPP induces β-cell apoptosis and is characteristic of β-cells in humans with type 2 diabetes but not type 1 diabetes. These findings are consistent with a role of protein misfolding in β-cell apoptosis in type 2 diabetes.

OBJECTIVE—Type 1 and type 2 diabetes are characterized by an ∼98 and ∼65% loss of pancreatic β-cells, respectively. Efforts to reverse either form of diabetes increasingly focus on the possibility of promoting β-cell replacement and/or regeneration. Islet transplantation has been explored, but it does not provide long-term insulin independence. One possible source of β-cell regeneration is hematopoietic stem cells. In mice, there are conflicting data as to whether hematopoietic stem cells contribute to pancreatic β-cells. We sought to establish whether hematopoietic stem cells (derived from adult donors) transdifferentiate into pancreatic β-cells in adult humans. RESEARCH DESIGN AND METHODS—We addressed this in 31 human pancreata obtained at autopsy from hematopoietic stem cell transplant recipients who had received their transplant from a donor of the opposite sex. RESULTS—Whereas some donor-derived cells were observed in the nonendocrine pancreata, no pancreatic β-cells were identified that were derived from donor hematopoietic stem cells, including two cases with type 2 diabetes. CONCLUSIONS—We conclude that hematopoietic stem cells derived from adult donors contribute minimally to pancreatic β-cells in nondiabetic adult humans. These data do not rule out the possibility that hematopoietic stem cells contribute to pancreatic β-cells in childhood or in individuals with type 1 diabetes.

Insulin secretion from the 2,000–3,000 β-cells in an islet is a highly synchronized activity with discharge of insulin in coordinate secretory bursts at approximately 4-min intervals. Insulin secretion progressively declines in type 2 diabetes and following islet transplantation. Both are characterized by the presence of islet amyloid derived from islet amyloid polypeptide (IAPP). In the present studies, we examined the action of extracellular human IAPP (h-IAPP) on morphology and function of human islets. Because oligomers of h-IAPP are known to cause membrane disruption, we questioned if application of h-IAPP oligomers to human islets would lead to disruption of islet architecture (specifically cell-to-cell adherence) and a decrease in coordinate function (e.g., increased entropy of insulin secretion and diminished coordinate secretory bursts). Both hypotheses are affirmed, leading to a novel hypothesis for impaired insulin secretion in type 2 diabetes and following islet transplantation, specifically disrupted cell-to-cell adherence in islets through the actions of membrane-disrupting IAPP oligomers.

Intrahepatic islet transplantation is an experimental therapy for type 1 diabetes. In the present studies, we sought to address the following questions: 1) In humans, do intrahepatic transplanted islets reestablish coordinated puslatile insulin secretion? and 2) To what extent is insulin secreted by intrahepatic transplanted islets delivered to the hepatic sinusoids (therefore effectively restoring a portal mode of insulin delivery) versus delivered to the hepatic central vein (therefore effectively providing a systemic form of insulin delivery)? To address the first question, we examined insulin concentration profiles in the overnight fasting state and during a hyperglycemic clamp ( approximately 150 mg/dl) in 10 recipients of islet transplants and 10 control subjects. To address the second question, we measured first-pass hepatic insulin clearance in two recipients of islet autografts after pancreatectomy for pancreatitis versus five control subjects by direct catheterization of the hepatic vein. We report that coordinate pulsatile insulin secretion is reestablished in islet transplant recipients and that glucose-mediated stimulation of insulin secretion is accomplished by amplification of insulin pulse mass. Direct hepatic catheterization studies revealed that intrahepatic islets in humans do deliver insulin directly to the hepatic sinusoid because approximately 80% of the insulin is extracted during first pass. In conclusion, intrahepatic islet transplantation effectively restores the liver to pulsatile insulin delivery.

Impaired fasting glucose (IFG) and impaired glucose tolerance (IGT) often coexist and as such represent a potent risk factor for subsequent development of type 2 diabetes. beta-Cell mass is approximately 50% deficient in IFG and approximately 65% deficient in type 2 diabetes. To establish the effect of a approximately 50% deficit in beta-cell mass on carbohydrate metabolism, we performed a approximately 50% partial pancreatectomy versus sham surgery in 14 dogs. Insulin secretion was quantified from insulin concentrations measured in the portal vein at 1-min sampling intervals under basal conditions, after a 30-g oral glucose, and during a hyperglycemic clamp. Insulin sensitivity was measured by a hyperinsulinemic-euglycemic clamp combined with isotope dilution. Partial pancreatectomy resulted in IFG and IGT. After partial pancreatectomy both basal and glucose-stimulated insulin secretion were decreased through the mechanism of a selective approximately 50 and approximately 80% deficit in insulin pulse mass, respectively (P < 0.05). These defects in insulin secretion were partially offset by decreased hepatic insulin clearance (P < 0.05). Partial pancreatectomy also caused a approximately 40% decrease in insulin-stimulated glucose disposal (P < 0.05), insulin sensitivity after partial pancreatectomy being related to insulin pulse amplitude (r = 0.9, P < 0.01). We conclude that a approximately 50% deficit in beta-cell mass can recapitulate the alterations in glucose-mediated insulin secretion and insulin action in humans with IFG and IGT. These data support a mechanistic role of a deficit in beta-cell mass in the evolution of IFG/IGT and subsequently type 2 diabetes.

Type 2 diabetes is characterized by defects in insulin secretion and action and is preceded by impaired fasting glucose (IFG). The islet anatomy in IFG and type 2 diabetes reveals an approximately 50 and 65% deficit in beta-cell mass, with increased beta-cell apoptosis and islet amyloid derived from islet amyloid polypeptide (IAPP). Defects in insulin action include both hepatic and extrahepatic insulin resistance. The relationship between changes in beta-cell mass, beta-cell function, and insulin action leading to type 2 diabetes are unresolved, in part because it is not possible to measure beta-cell mass in vivo, and most available animal models do not recapitulate the islet pathology in type 2 diabetes. We evaluated the HIP rat, a human IAPP transgenic rat model that develops islet pathology comparable to humans with type 2 diabetes, at age 2 months (nondiabetic), 5 months (with IFG), and 10 months (with diabetes) to prospectively examine the relationship between changes in islet morphology versus insulin secretion and action. We report that increased beta-cell apoptosis and impaired first-phase insulin secretion precede the development of IFG, which coincides with an approximately 50% defect in beta-cell mass and onset of hepatic insulin resistance. Diabetes was characterized by approximately 70% deficit in beta-cell mass, progressive hepatic and extrahepatic insulin resistance, and hyperglucagonemia. We conclude that IAPP-induced beta-cell apoptosis causes defects in insulin secretion and beta-cell mass that lead first to hepatic insulin resistance and IFG and then to extrahepatic insulin resistance, hyperglucagonemia, and diabetes. We conclude that a specific beta-cell defect can recapitulate the metabolic phenotype of type 2 diabetes and note that insulin resistance in type 2 diabetes may at least in part be secondary to beta-cell failure.

OBJECTIVE—The purpose of this study was to establish whether hypoglycemia after gastric bypass surgery (GBS) for morbid obesity is due to increased fractional β-cell area or inappropriately increased insulin secretion.RESEARCH DESIGN AND METHODS—We examined pancreata obtained at partial pancreatectomy from 6 patients with post-GBS hypoglycemia and compared these with 31 pancreata from obese subjects and 16 pancreata from lean control subjects obtained at autopsy. We addressed the following questions. In patients with post-GBS hypoglycemia, is β-cell area increased and is β-cell formation increased or β-cell apoptosis decreased?RESULTS—We report that in patients with post-GBS hypoglycemia, β-cell area was not increased compared with that in obese or even lean control subjects. Consistent with this finding, there was no evidence of increased β-cell formation (islet neogenesis and β-cell replication) or decreased β-cell loss in patients with post-GBS hypoglycemia. In control subjects, mean β-cell nuclear diameter correlated with BMI (r2 = 0.79, P < 0.001). In patients with post-GBS hypoglycemia, β-cell nuclear diameter was increased (P < 0.001) compared with that for BMI in matched control subjects but was appropriate for BMI before surgery.CONCLUSIONS—We conclude that post-GBS hypoglycemia is not due to increases in β-cell mass or formation. Rather, postprandial hypoglycemia after GBS is due to a combination of gastric dumping and inappropriately increased insulin secretion, either as a failure to adaptively decrease insulin secretion after GBS or as an acquired phenomenon.

Type 2 diabetes is characterized by an ∼60% loss of β-cell mass, a marked defect in postprandial insulin secretion, and a failure to suppress postprandial glucagon concentrations. It is possible that postprandial hyperglucagonemia in type 2 diabetes is due to impaired postprandial insulin secretion. To address this, we studied eight adult Goettingen minipigs before and after an ∼60% reduction in β-cell mass induced by alloxan. Pigs were studied fasting and after ingestion of a mixed meal. Insulin and glucagon secretion were determined by deconvolution of blood hormone concentrations measured at 1-min intervals. The relationship between insulin and glucagon release was analyzed using cross-correlation and forward versus reverse cross–approximate entropy. We report that glucagon and insulin were secreted in ∼4-min pulses. Prealloxan, postprandial insulin secretion drove an ∼20% suppression of glucagon concentrations (P < 0.01), through inhibition of glucagon pulse mass. The alloxan-induced ∼60% deficit in β-cell mass lead to an ∼70% deficit in postprandial insulin secretion and loss of the postprandial insulin-driven suppression of glucagon secretion. We conclude that postprandial hyperglucagonemia in type 2 diabetes is likely due to loss of intraislet postprandial suppression of glucagon secretion by insulin.

In health, insulin is secreted in discrete pulses into the portal vein, and the regulation of the rate of insulin secretion is accomplished by modulation of insulin pulse mass. Several lines of evidence suggest that the pattern of insulin delivery by the pancreas determines hepatic insulin clearance. In previous large animal studies, the amplitude of insulin pulses was related to the extent of insulin clearance. In humans (and in large animals), the amplitude of insulin oscillations is approximately 100-fold higher in the portal vein than in the systemic circulation, despite only a fivefold dilution, implying preferential hepatic extraction of insulin pulses. In the present study, by direct hepatic vein sampling in healthy humans, we sought to establish the extent of first-pass hepatic insulin extraction and to determine whether the pattern of insulin secretion (insulin pulse mass and amplitude) dictates the hepatic insulin clearance and thereby delivery of insulin to extrahepatic insulin-responsive tissues. Five nondiabetic subjects (two men and three women, mean age 32 years [range 25-39], BMI 24.9 kg/m(2) [21.2-27.1]) participated. Insulin and C-peptide delivery from the splanchnic bed was measured in basal overnight-fasted state and during a glucose infusion of 2 mg . kg(-1) . min(-1) by simultaneous sampling from the hepatic vein and an arterialized vein along with direct estimation of splanchnic blood flow. Fractional insulin extraction was calculated from the difference between the C-peptide and insulin delivery rates from the liver. The time patterns of insulin concentrations and hepatic insulin clearance were analyzed by deconvolution and Cluster analysis, respectively. Cross-correlation analysis was used to relate C-peptide secretion and insulin clearance. Glucose infusion increased peripheral glucose concentrations from 5.4 +/- 0.1 to 6.4 +/- 0.4 mmol/l (P < 0.05). Likewise, insulin and C-peptide concentrations increased during glucose infusion (P < 0.05). Hepatic insulin clearance increased with glucose infusion (1.06 +/- 0.18 vs. 2.55 +/- 0.38 pmol . kg(-1) . min(-1); P < 0.01), but fractional hepatic insulin clearance was stable (78.2 +/- 4.4 vs. 84 0. +/- 3.9%, respectively; P = 0.18). Insulin secretory-burst mass rose during glucose infusion (P < 0.05), whereas the interburst interval remained unchanged (4.4 +/- 0.2 vs. 4.5 +/- 0.3 min; P = 0.36). Cluster analysis identified an oscillatory pattern in insulin clearance, with peaks occurring approximately every 5 min. Cross-correlation analysis between prehepatic C-peptide secretion and hepatic insulin clearance demonstrated a significant positive association without detectable (

The islet in type 2 diabetes is characterized by a deficit in beta-cell mass, increased beta-cell apoptosis, and impaired insulin secretion. Also, islets in type 2 diabetes often contain deposits of islet amyloid derived from islet amyloid polypeptide (IAPP), a 37-amino acid protein cosecreted with insulin by beta-cells. Several lines of evidence suggest that proteins with a capacity to develop amyloid fibrils may also form small toxic oligomers that can initiate apoptosis. The amino acid sequence of IAPP in rats and mice is identical and differs from that in humans by substitution of proline residues in the amyloidogenic sequence so that the protein no longer forms amyloid fibrils or is cytotoxic. In the present study, we report a novel rat model for type 2 diabetes: rats transgenic for human IAPP (the HIP rat). HIP rats develop diabetes between 5 and 10 months of age, characterized by an approximately 60% deficit in beta-cell mass that is due to an increased frequency of beta-cell apoptosis. HIP rats develop islet amyloid, but the extent of amyloid was not related to the frequency of beta-cell apoptosis (r = 0.10, P = 0.65), whereas the fasting blood glucose was (r = 0.77, P < 0.001). The frequency of beta-cell apoptosis was related to the frequency of beta-cell replication (r = 0.97, P < 0.001) in support of the hypothesis that replicating cells are more vulnerable to apoptosis than nondividing cells. The HIP rat provides additional evidence in support of the potential role of IAPP oligomer formation toward the increased frequency of apoptosis in type 2 diabetes, a process that appears to be compounded by glucose toxicity when hyperglycemia supervenes.

Nondiabetic obese humans adapt to insulin resistance by increasing beta-cell mass. In contrast, obese humans with type 2 diabetes have an approximately 60% deficit in beta-cell mass. Recent studies in rodents reveal that beta-cell mass is regulated, increasing in response to insulin resistance through increased beta-cell supply (islet neogenesis and beta-cell replication) and/or decreased beta-cell loss (beta-cell apoptosis). Prospective studies of islet turnover are not possible in humans. In an attempt to establish the mechanism for the deficit in beta-cell mass in type 2 diabetes, we used an obese versus lean murine transgenic model for human islet amyloid polypeptide (IAPP) that develops islet pathology comparable to that in humans with type 2 diabetes. By 40 weeks of age, obese nontransgenic mice did not develop diabetes and adapted to insulin resistance by a 9-fold increase (P < 0.001) in beta-cell mass accomplished by a 1.7-fold increase in islet neogenesis (P < 0.05) and a 5-fold increase in beta-cell replication per islet (P < 0.001). Obese transgenic mice developed midlife diabetes with islet amyloid and an 80% (P < 0.001) deficit in beta-cell mass that was due to failure to adaptively increase beta-cell mass. The mechanism subserving this failed expansion was a 10-fold increase in beta-cell apoptosis (P < 0.001). There was no relationship between the extent of islet amyloid or the blood glucose concentration and the frequency of beta-cell apoptosis. However, the frequency of beta-cell apoptosis was related to the rate of increase of islet amyloid. These prospective studies suggest that the formation of islet amyloid rather than the islet amyloid per se is related to increased beta-cell apoptosis in this murine model of type 2 diabetes. This finding is consistent with the hypothesis that soluble IAPP oligomers but not islet amyloid are responsible for increased beta-cell apoptosis. The current studies also support the concept that replicating beta-cells are more vulnerable to apoptosis, possibly accounting for the failure of beta-cell mass to expand appropriately in response to obesity in type 2 diabetes.

Type 2 diabetes is characterized by a relative beta-cell deficit as a result of increased beta-cell apoptosis and islet amyloid derived from the beta-cell peptide islet amyloid polypeptide (IAPP). Human IAPP (h-IAPP) but not mouse IAPP (m-IAPP) induces apoptosis when applied to cells in culture, a property that depends on the propensity of h-IAPP to oligomerize. Since beta-cell mass is regulated, the question arises as to why it is not adaptively increased in response to insulin resistance and hyperglycemia in type 2 diabetes. This adaptation might fail if dividing beta-cells preferentially underwent apoptosis. We tested the hypothesis that beta-cells are preferentially vulnerable to h-IAPP-induced apoptosis. We established a microculture environment to perform time-lapse video microscopy (TLVM) and studied beta-cells (RIN) and HeLa cells undergoing replication or apoptosis. Sequential images (every 10 min for 36 h in RIN or 24 h in HeLa cells) of cells in vivo were analyzed, and each mitotic and apoptotic event was documented. Freshly dissolved h-IAPP caused a dose-dependent increased rate of apoptosis (P < 0.0001) in both cell types. At low and medium levels of toxicity, cells that had previously undergone mitosis were more vulnerable to h-IAPP-induced apoptosis than nondividing cells (P < 0.05). In the first 3 h after mitosis (full cell cycle length 26 +/- 0.6 h), beta-cells were particularly susceptible to h-IAPP-induced apoptosis (P < 0.05). Neither m-IAPP nor mature amyloid aggregates of h-IAPP were cytotoxic (P = 0.49). To corroborate these cell culture studies, we examined sections of human pancreatic tissue (five cases of type 2 diabetes) and human islets incubated for 48 h +/- h-IAPP. Both were stained for apoptosis with the transferase-mediated dUTP nick-end labeling method and analyzed for the presence of paired apoptotic cells anticipated in the event of postmitotic apoptosis. In human pancreatic tissue 26 +/- 5% (single plane of examination) and in human islets incubated with h-IAPP 44 +/- 4% of apoptotic islet cells were paired. In conclusion, replicating beta-cells are preferentially vulnerable to h-IAPP-induced apoptosis in cell culture. Postmitotic apoptosis was also documented in humans with type 2 diabetes and in human islet tissue. We postulate that beta-cell deficiency in type 2 diabetes may result in part from failure to adaptively increase beta-cell mass due to increased vulnerability of replicating beta-cells to undergo apoptosis. If this postulate is correct, then inhibition of apoptosis should allow recovery of beta-cell mass in type 2 diabetes.

Most insulin is secreted in discrete pulses at an interval of approximately 6 min. Increased insulin secretion after meal ingestion is achieved through the mechanism of amplification of the burst mass. Conversely, in type 2 diabetes, insulin secretion is impaired as a consequence of decreased insulin pulse mass. beta-cell mass is reported to be deficient in type 2 diabetes. We tested the hypothesis that decreased beta-cell mass leads to decreased insulin pulse mass. Insulin secretion was examined before and after an approximately 60% decrease in beta-cell mass achieved by a single injection of alloxan in a porcine model. Alloxan injection resulted in stable diabetes (fasting plasma glucose 7.4 +/- 1.1 vs. 4.4 +/- 0.1 mmol/l; P0.01) with impaired insulin secretion in the fasting and fed states and during a hyperglycemic clamp (decreased by 54, 80, and 90%, respectively). Deconvolution analysis revealed a selective decrease in insulin pulse mass (by 54, 60, and 90%) with no change in pulse frequency. Rhythm analysis revealed no change in the periodicity of regular oscillations after alloxan administration in the fasting state but was unable to detect stable rhythms reliably after enteric or intravenous glucose stimulation. After alloxan administration, insulin secretion and insulin pulse mass (but not insulin pulse interval) decreased in relation to beta-cell mass. However, the decreased pulse mass (and pulse amplitude delivered to the liver) was associated with a decrease in hepatic insulin clearance, which partially offset the decreased insulin secretion. Despite hyperglycemia, postprandial glucagon concentrations were increased after alloxan administration (103.4 +/- 6.3 vs. 92.2 +/- 2.5 pg/ml; P0.01). We conclude that an alloxan-induced selective decrease in beta-cell mass leads to deficient insulin secretion by attenuating insulin pulse mass, and that the latter is associated with decreased hepatic insulin clearance and relative hyperglucagonemia, thereby emulating the pattern of islet dysfunction observed in type 2 diabetes.

According to the Oxford Dictionary of Proverbs , the oldest written version of the saying “What you don't know can't hurt you” comes from Petit Palace , written in 1576 by G. Pettie: “So long as I know it not, it hurteth mee not.” In this issue of Diabetes Care , Drucker et al. (1) conclude that the safety profile of the newly available glucagon-like peptide 1 (GLP-1) class of drugs is favorable in comparison to their benefits as therapy, and the class of drugs might be considered as next in line after metformin for treatment for type 2 diabetes. The purpose of this counterpoint is to suggest such a conclusion is premature. History has taught us that enthusiasm for new classes of drugs, heavily promoted by the pharmaceutical companies that market them, can obscure the caution that should be exercised when the long-term consequences are unknown. Of perhaps greatest concern in the case of the GLP-1–based drugs, including GLP-1 agonists and dipeptidyl peptidase-4 (DPP-4) inhibitors, is preliminary evidence to suggest the potential risks of asymptomatic chronic pancreatitis and, with time, pancreatic cancer. The GLP-1–related drugs arrived in clinical practice with much fanfare and anticipation. As summarized in the article by Drucker et al., it is a class of drugs that has potential benefits in the treatment of type 2 diabetes. The concept of gut-related factors that enhance glucose-mediated insulin secretion, the incretin effect, has been recognized for many years (2). Once it was demonstrated that an intravenous infusion of GLP-1 could decrease blood glucose concentrations in patients with type 2 diabetes, the race was on to exploit the properties of this action. Many millions of dollars have been invested by the pharmaceutical industry in developing products, the first of which are now in clinical practice. Many millions of dollars …

We have previously shown that hemizygous transgenic mice expressing human islet amyloid polypeptide (hIAPP) in pancreatic beta-cells have no diabetic phenotype, whereas in the homozygous state, they developed severe, early-onset hyperglycemia associated with impaired insulin secretion and beta-cell death. We investigated the possibility that when the hemizygous mice are crossed onto an obese, insulin-resistant strain such as agouti viable yellow (A(vy)/a), they would exhibit a phenotype more akin to human type 2 diabetes. The hIAPP-expressing A(vy) males (TG-Y) displayed fasting hyperglycemia at 90 days of age and by 1 year progressed to severe hyperglycemia relative to their nontransgenic counterparts. Plasma insulin concentrations and pancreatic insulin content dropped 10- to 20-fold, suggesting severe impairment of beta-cell function. Histopathological findings revealed beta-cell degeneration and loss consistent with the drop in the plasma insulin concentration. In addition, large deposits of IAPP amyloid were present in TG-Y islets. We conclude that in transgenic mice expressing hIAPP, insulin resistance can induce overt, slow-onset diabetes associated with islet amyloid and decreased beta-cell mass.

Although sulfonylureas enhance insulin secretion, it is unknown whether these hypoglycemic chemicals stimulate insulin secretion through the augmentation of the pulsatile or basal modes of insulin release. Enhanced pulsatile insulin could occur in turn through amplification of the burst mass or an increase in burst frequency. To address the mechanism of sulfonylurea action, we employed a recently validated canine model with a portal vein sampling catheter and flow probe to measure pulsatile insulin secretion in vivo directly in response to tolbutamide infusion or ingestion. After a 16-h fast, seven dogs were studied in the postabsorptive basal state and during a tolbutamide (0.2 mg/min) infusion when their plasma glucose concentrations were clamped at euglycemia. Insulin concentrations in the carotid artery (basal vs. tolbutamide, 85 ± 12 vs. 325 ± 66 pmol/l; P < 0.01) and portal vein (basal vs. tolbutamide, 345 ± 55 vs. 1,288 ± 230 pmol/l; P < 0.01) increased during tolbutamide infusion, but the portal vein plasma flow did not change. Increased plasma insulin concentrations were achieved by a fourfold increase in the total insulin secretion rate (2.3 ± 0.2 to 9.4 ± 1.9 pmol · kg−1 · min−1; basal vs. tolbutamide, P < 0.01). The augmented total insulin secretion was achieved mechanistically via a marked and selective increase in the insulin secretory burst mass (basal vs. tolbutamide, 266 ± 64 vs. 817 ± 144 pmol/pulse; P < 0.01), with no change in portal-vein insulin pulse frequency (basal vs. tolbutamide, 10.1 ± 0.6 vs. 11.1 ± 0.8 pulses/h; P = 0.3). Oral (250 mg) tolbutamide also magnified the endogenous insulin secretion rate by the preferential amplification of the secretory pulse mass (basal vs. tolbutamide, 167 ± 37 vs. 362 ± 50 pmol/pulse; P < 0.01). Neither the infusion nor the ingestion of tolbutamide changed the calculated clearance rates of endogenously secreted insulin. We conclude that sulfonylurea (tolbutamide) induced insulin secretion in vivo is achieved by the highly selective amplification of insulin secretory burst mass with no change in basal insulin release or the frequency of the β-cell-network pacemaker.

In the present studies, we used a recently validated canine model to determine 1) if glucose ingestion stimulates insulin secretion by amplifying the pulsatile component of insulin release, and if so, 2) whether this effect is achieved preferentially through burst mass or frequency modulation, and 3) if the mechanism of incretin effect of insulin secretion is mediated via the pulsatile mode of secretion. We report that 30 g of glucose ingestion stimulates an approximately 550% increase in the overall rate of insulin secretion (1.8 +/- 0.2 to 11.6 +/- 1.5 pmol.kg-1.min-1), which is achieved via an approximately 400% increase in the mass of insulin secreted per burst (202 +/- 38 to 1,003 +/- 147 pmol/pulse, P0.001) and a approximately 40% increase in burst frequency (8.7 +/- 0.5 to 12.3 +/- 0.6 pulse/h, P0.001). Of the insulin secreted after glucose ingestion, 68% (+/-4) was released in discrete secretory bursts. Further analyses showed that the incretin effect of ingested (GPO) versus infused glucose (GIV) is achieved through regulation of pulsatile insulin secretion. Glucose ingestion led to an approximately 70% greater rate of insulin secretion than intravenous glucose delivery (10.0 +/- 1.6 vs. 5.9 +/- 0.9 pmol.kg-1.min-1, P0.005, GPO vs. GIV). This incretin effect was achieved by the specific mechanism of an approximately 70% greater pulse mass (930 +/- 196 vs. 558 +/- 97 pmol/pulse, P0.02, GPO vs. GIV) but with a comparable pulse frequency (13.1 +/- 0.9 vs. 12.0 +/- 0.5 pulses/h, P = 0.14, n = 9 dogs, GPO vs. GIV). We conclude that in vivo glucose regulates overall insulin secretion almost exclusively by amplification of the pulsatile mode of insulin secretion, and that the incretin effect is achieved by preferential enhancement of insulin secretory burst mass.

Amyloid deposits that characteristically form in the pancreatic islets of patients with non-insulin-dependent diabetes mellitus (NIDDM) and in insulinomas are both derived from islet amyloid polypeptide (IAPP). Evidence from previous studies has suggested that deposition of IAPP-derived amyloid is related to inherent amyloidogenic sequences present within normal human IAPP, together with an increased production and local concentration of IAPP. However, whether the aggregation of IAPP to form amyloid fibrils is primarily an intra- or extracellular event is not clear. To address this question, we studied 20 human insulinomas by light and electron microscopy. By light microscopy, amyloid deposits were demonstrated in 13 of 20 (65%) human insulinomas. Furthermore, evaluation of Congo red-stained tumor sections showed small, globular or irregular, congophilic amyloid deposits within the cytoplasm of many tumor cells in 10 of 13 (77%) amyloid-containing insulinomas. Dense, punctate areas of IAPP immunoreactivity within tumor cells corresponded with the congophilic intracellular deposits. Ubiquitin immunoreactivity also was observed as punctate intracellular labeling and within large extracellular amyloid deposits. Among the 10 insulinomas available for electron microscopic evaluation, pathological IAPP-immunoreactive (immunogold) deposits were found in 3 of 5 insulinomas in which amyloid was demonstrated by light microscopy and in none of 5 tumors found negative for amyloid by light microscopy. Morphology of IAPP-immunoreactive deposits varied from those with the classical distinct 7- to 10-nm diameter nonbranching fibrils to those with distinct but faint fibrillarity to those without discernable fibrils. In each of the 3 tumors with ultrastructurally demonstrable amyloid deposits, intracellular aggregates of IAPP-immunoreactive amyloid fibrils were observed either free in the cytoplasmic matrix or in membrane-bound structures. Intracellular IAPP-immunoreactive deposits were up to 10 urn in the greatest dimension. Free intracellular aggregates of amyloid fibrils sometimes contained degenerating secretory vesicles and fragments of membranous organelles. These observations suggest that IAPP-derived amyloid deposits are initially formed intracellularly and are subsequently released to the extracellular space by exocytosis of the membrane-bound structures and/or after necrosis of the tumor cells. Our observations raise the possibility that IAPP-derived islet amyloid deposits (characteristically present in NIODM patients) also may be initially formed within the cytoplasm of β-cells, thus leading to β-cell necrosis and a reduction of β-cell mass as seen in NIDDM.

Excessive amounts of glucose enter the systemic circulation when patients with non-insulin-dependent diabetes mellitus (NIDDM) eat a carbohydrate-containing meal. To determine the contribution of hepatic glucose cycling (defined as the net effect of glucose/glucose-6-phosphate cycling and uptake and release of glucose from hepatic glycogen) to postprandial hyperglycemia, diabetic, glucose-intolerant, and nondiabetic subjects were fed mixed meals. The meal contained both [2-3H]glucose (an isotope that is extensively detritiated during hepatic glucose cycling) and [6-3H]glucose (an isotope that is not detritiated during hepatic glucose cycling). Of the 50 g of carbohydrate contained in the meal, ∼4–8 g underwent hepatic glucose cycling. Although total cycling of ingested glucose did not differ between diabetic, glucose-intolerant, and nondiabetic subjects (361 ± 67 vs. 494 ± 106 vs. 322 ± 44 μmol · kg−1 · 5 h−1, respectively), the data suggested that hepatic cycling was increased in the diabetic and glucose-intolerant individuals but not in the nondiabetic subjects during the first 2 h after eating. Hepatic cycling during the first 2 h after eating was correlated with the prevailing glucagon concentration (r = 0.6, P < 0.01) and increased (P < 0.05) as hepatic glucose release increased. Hepatic glucose cycling had a marked effect on the measurement of so-called initial splanchnic glucose uptake. Nevertheless, however measured, initial splanchnic glucose uptake was not decreased and, if anything, was increased in diabetic and glucose-intolerant patients. Integrated postprandial hepatic glucose release increased (r < 0.01) with the severity of fasting hyperglycemia. These results indicate that postprandial hyperglycemia in diabetic and glucose-intolerant patients is not due to enhanced hepatic glucose cycling or a lack of uptake of ingested glucose by the splanchnic bed but rather to excessive release of glucose by the liver coupled with a failure of extrahepatic tissues to appropriately increase glucose disposal.

Insulinlike growth factor binding protein 1 (IGFBP-1) has been shown to modulate the metabolic and mitogenic actions of the growth hormone (GH)- dependent peptide insulinlike growth factor I. Previous studies showed that levels of IGFBP-1 are regulated by insulin. The relative role of GH in the regulation of IGFBP-1 levels is less well defined and was examined in our study with a contiguous two-part protocol. Overnight (part A) and pre- and post-morning meal (part B) blood samples were obtained from eight healthy adults during a constant infusion of saline (SAL) or 4 μg · kg−1 · min−1 GH. Five of eight subjects were restudied with glucose (GLUC) infused during part B (SAL + GLUC) to match glucose and insulin to levels observed during GH infusion. During SAL infusion, IGFBP-1 levels measured by specific radioimmunoassay showed a marked immediate decline after the evening meal in part A, with a subsequent nocturnal rise of 2.4- to 17.3-fold. GH infusion resulted in a similar meal-induced fall in IGFBP-1 levels but led to a delayed nocturnal rise in IGFBP-1, which was associated with elevated postprandial insulin concentrations. During part B, changes in plasma IGFBP-1 levels showed a similar pattern, with a delayed postprandial increase observed during both GH and SAL + GLUC infusions. The half-life of IGFBP-1 disappearance was calculated at ∼2 h for all three infusion groups. Comparison of venous and arterialized blood samples showed no consistent pattern of difference, arguing against peripheral tissue clearance or compartmentalization as the mechanism for the rapid rise and fall in IGFBP-1 levels. Our studies, the first to measure GH, insulin, and glucose in concurrent sampling and under controlled physiological conditions, support previous investigations suggesting that insulin (not GH or glucose) is the primary regulator of plasma IGFBP-1 levels. Furthermore, we postulate that the observed fluctuations in plasma levels are caused by a direct insulin effect on hepatic IGFBP-1 production.

The importance of portal insulin delivery in the regulation of postprandial carbohydrate metabolism is uncertain. To address this question, three groups of dogs were studied: one group in which pancreatic venous drainage was transected and reanastomosed (portal insulin delivery), one in which the pancreatic drainage was transected and anastomosed to the inferior vena cava (peripheral insulin delivery), and one that received only a sham operation. Plasma insulin was greater (P < 0.05) during peripheral insulin delivery than in either the portal or sham groups, respectively, before and after meal ingestion. On the other hand, C-peptide concentrations did not differ between groups, resulting in a higher (P < 0.001) insulin to C-peptide ratio in the peripheral group. This indicated that the hyperinsulinemia in the peripheral group was due to decreased insulin clearance rather than increased insulin secretion. Isotopically determined splanchnic uptake of ingested glucose, postprandial suppression of hepatic glucose release, incorporation of CO2 into glucose (a qualitative measure of gluconeogenesis), and total-body glucose uptake were virtually identical in all groups. Similarly, plasma lipid, β-hydroxybutyrate, and lactate concentrations did not differ between groups. Our data indicate that, despite differences in systemic insulin concentration, portal and peripheral insulin delivery comparably regulate hepatic and extrahepatic carbohydrate metabolism after meal ingestion.

It is well established that patients with non-insulindependent diabetes mellitus (NIDDM) are resistant to insulin. However, the contribution of hepatic and extrahepatic tissues to insulin resistance remains controversial. The uncertainty may be at least in part due to errors introduced by the unknowing use in previous studies of impure isotopes to measure glucose turnover. To determine hepatic and extrahepatic responses to insulin in the absence of these errors, steady-state glucose turnover was measured simultaneously with [6-3H]- and [6-14C]glucose during sequential 5- and 4-h infusions of insulin at rates of 0.4 and 10 mil · kg−1 · min−1 in diabetic and nondiabetic subjects. At low insulin concentrations, [6-3H]- and [6-14C]glucose gave similar estimates of glucose turnover. Hepatic glucose release was equal to but not below zero in the nondiabetic subjects, but persistent glucose release (P < 0.001) and decreased glucose uptake (P < 0.001) was observed in the diabetic patients. At high insulin concentrations, both isotopes underestimated glucose turnover during the 1st h after initiation of the highdose insulin infusion. More time (P < 0.05) was required to reachieve steady state in NIDDM than nondiabetic subjects. At steady state, [6-3H] but not [6-14C]glucose systematically underestimated (P < 0.05) glucose turnover in both groups due to the presence of a tritiated nonglucose contaminant. The percentage of radioactivity in plasma due to tritiated contaminants was linearly related to turnover. When plasma [6-3H]glucose specific activity was corrected for the presence of the contaminant in each subject, both [6-3H]- and [6-14C]glucose indicated that diabetic patients had hepatic and extrahepatic insulin resistance. We conclude that in these experiments, although the presence of a low percentage of tritiated nonglucose contaminant in the tracer may alter estimates of the severity of hepatic and extrahepatic insulin resistance in patients with NIDDM, it does not obscure their presence

Progress in biomedical research is documented in journals after peer review, a process with flaws but the best we have. Once an article is published, in general it faces three fates. Most commonly, after some measure of initial interest, it quietly slips into obscurity, only occasionally to resurface if cited by a vigilant successor who recognizes the prior contribution to a once again advancing field. A quick glance through a copy of any major medical journal published ≥50 years ago is a humbling experience for any active scientist. Many of the confidently stated conclusions, with hindsight, were quite simply wrong. How many of our conclusions today will seem mildly amusing to a reader 50 years from now? Occasionally, an article makes a tremendously important contribution that advances a field and is widely cited, becoming a so-called citation classic with little controversy. Reading such a paper that is ≥50 years old can also be a humbling experience. How did the authors have such insight from what we now consider an incomplete dataset obtained by insensitive and nonspecific methods compared with those available to us now? It was clear to the editorial team …

This month marks the “official” transition for the Editorial Board of the journal Diabetes . For the last 5 years, Diabetes has flourished under the editorial guidance of the Editor in Chief, Dr. Franz Matschinsky, and his Associate Editors. Indeed, the number of manuscripts submitted to Diabetes has increased substantially during this time period (Fig. 1), and the journal’s citation impact has continued to rise. In addition to the excellent leadership provided by Dr. Matschinsky, the journal has benefited from a highly efficient editorial office in Philadelphia, managed by Vesselina Panteva. On behalf of the diabetes research community, we thank Dr. Matschinsky and his Associate Editorial team, as well as Vesselina Panteva and Rachel Edwards, for their service to Diabetes journal. Editorial transition offers an opportunity for change, although the first priority of the incoming team is not to disturb the formula of success that has been so effective under the leadership of Dr. Matschinsky. However, one key change involves the geographic distribution of the Associate Editors. With the benefits of improved communication technologies and the global impact of …

This December 2011 edition of Diabetes marks the completion of the 5-year term of the outgoing editorial team. It has been my honor and pleasure to serve as editor with such a dedicated and accomplished team of associate editors. I thank each of them for their gift of time, their patience, their scholarship, their integrity, and their good humor. We would also like to collectively thank Edwin Gale, the former editor in chief of Diabetologia and Mayer Davidson, former editor in chief of Diabetes Care , who both gave us generous and useful advice as we took on the task of editing Diabetes . We would also like to thank the editorial staff at the University of California, Los Angeles, and at the American Diabetes Association office in Indiana, for assisting us in striving to give submitting authors a timely decision, even if not always one they wanted. As Diabetes now moves forward under …

We read the recent paper by Tushuizen et al. (1) with interest. In that study, the authors measured pancreatic fat content by use of proton magnetic resonance spectroscopy in 12 type 2 diabetic and 24 nondiabetic men and examined the relationship between pancreatic fat content and β-cell function. They report a negative correlation between pancreatic fat and a measure of β-cell function in nondiabetic controlsubjects (but not individuals with type 2 diabetes) and conclude that “pancreatic lipid content may contribute to β-cell dysfunction and possibly to the subsequent development of type 2 diabetes in susceptible humans.” We respectfully suggest that the report contains insufficient evidence …

OBJECTIVE To compare incidence rates of childhood-onset IDDM among black African-heritage populations age 0–19 years in the Caribbean region. RESEARCH DESIGN AND METHODS Population-based registries for IDDM were established on the eastern Caribbean islands of Antigua, Barbados, Dominica, St. Croix, St. Kitts, St. Thomas, and Tortola using standardized criteria from the World Health Organization (WHO) Multinational Project for Childhood Diabetes (DiaMond). Average annual incidence rates (IR) with 95% CI for 0–19 years olds were computed using the DiaMond Registry program for the 5-year period from 1989 to 1993. Poisson regression analysis was used to determine differences in incidence rates. RESULTS The highest incidence rate for 0–19 year olds was for the black African-heritage population of St. Croix (IR 10.09 per 100,000; 95% CI 4.35–19.89), one of the U.S. Caribbean islands. A significant (P < 0.05) 3.9 variation in IDDM incidence across the registries was found when the IR for St. Croix was compared to the IR for Barbados (IR 2.57 per 100,000; 95% CI 0.90–4.64). CONCLUSIONS The variation in childhood-onset IDDM incidence rates among the black populations of the eastern Caribbean islands is consistent with the geographic variation in IDDM incidence seen among African Americans in the U.S. Variation in incidence rates of childhood diabetes in black populations may reflect differences in level of white genetic admixture or exposure to environmental diabetogenic agents.