Autoantibodies and High-Risk HLA Susceptibility Markers in First-Degree Relatives of Brazilian Patients with Type 1 Diabetes Mellitus: A Progression to Disease Based Study

The objective of this study was to determine the frequencies of autoantibodies to heterogeneous islet-cell cytoplasmic antigens (ICA), glutamic acid decarboxylase(65) (GAD(65)A), insulinoma-associated antigen-2 (IA-2A) and insulin (IAA)-and human leukocyte antigen (HLA) class II markers (HLA-DR and -DQ) in first degree relatives of heterogeneous Brazilian patients with type I diabetes(T1DM). A major focus of this study was to determine the influence of age, gender, proband characteristics and ancestry on the prevalence of autoantibodies and HLA-DR and -DQ alleles on disease progression and genetic predisposition to T1DM among the first-degree relatives. IAA, ICA, GAD(65)A, IA-2A and HLA- class II alleles were determined in 546 first-degree-relatives, 244 siblings, 55 offspring and 233 parents of 178 Brazilian patients with T1DM. Overall, 8.9% of the relatives were positive for one or more autoantibodies. IAA was the only antibody detected in parents. GAD(65) was the most prevalent antibody in offspring and siblings as compared to parents and it was the sole antibody detected in offspring. Five siblings were positive for the IA-2 antibody. A significant number (62.1%) of siblings had 1 or 2 high risk HLA haplotypes. During a 4-year follow-up study, 5 siblings (expressing HLA-DR3 or -DR4 alleles) and 1 offspring positive for GAD(65)A progressed to diabetes. The data indicated that the GAD(65) and IA-2 antibodies were the strongest predictors of T1DM in our study population. The high risk HLA haplotypes alone were not predictive of progression to overt diabetes.

Double-Stranded RNA-Activated Protein Kinase Is a Key Modulator of Insulin Sensitivity in Physiological Conditions and in Obesity in Mice

The molecular integration of nutrient-and pathogen-sensing pathways has become of great interest in understanding the mechanisms of insulin resistance in obesity. The double-stranded RNA-dependent protein kinase (PKR) is one candidate molecule that may provide cross talk between inflammatory and metabolic signaling. The present study was performed to determine, first, the role of PKR in modulating insulin action and glucose metabolism in physiological situations, and second, the role of PKR in insulin resistance in obese mice. We used Pkr(-/-) and Pkr(+/+) mice to investigate the role of PKR in modulating insulin sensitivity, glucose metabolism, and insulin signaling in liver, muscle, and adipose tissue in response to a high-fat diet. Our data show that in lean Pkr(-/-) mice, there is an improvement in insulin sensitivity, and in glucose tolerance, and a reduction in fasting blood glucose, probably related to a decrease in protein phosphatase 2A activity and a parallel increase in insulin-induced thymoma viral oncogene-1 (Akt) phosphorylation. PKR is activated in tissues of obese mice and can induce insulin resistance by directly binding to and inducing insulin receptor substrate (IRS)-1 serine307 phosphorylation or indirectly through modulation of c-Jun N-terminal kinase and inhibitor of kappa B kinase beta. Pkr(-/-) mice were protected from high-fat diet-induced insulin resistance and glucose intolerance and showed improved insulin signaling associated with a reduction in c-Jun N-terminal kinase and inhibitor of kappa B kinase beta phosphorylation in insulin-sensitive tissues. PKR may have a role in insulin sensitivity under normal physiological conditions, probably by modulating protein phosphatase 2A activity and serine-threonine kinase phosphorylation, and certainly, this kinase may represent a central mechanism for the integration of pathogen response and innate immunity with insulin action and metabolic pathways that are critical in obesity. (Endocrinology 153:5261-5274, 2012)

GLUT4 content decreases along with insulin resistance and high levels of inflammatory markers in rats with metabolic syndrome

Background: Metabolic syndrome is characterized by insulin resistance, which is closely related to GLUT4 content in insulin-sensitive tissues. Thus, we evaluated the GLUT4 expression, insulin resistance and inflammation, characteristics of the metabolic syndrome, in an experimental model. Methods: Spontaneously hypertensive neonate rats (18/group) were treated with monosodium glutamate (MetS) during 9 days, and compared with Wistar-Kyoto (C) and saline-treated SHR (H). Blood pressure (BP) and lipid levels, C-reactive protein (CRP), interleukin 6 (IL-6), TNF-alpha and adiponectin were evaluated. GLUT4 protein was analysed in the heart, white adipose tissue and gastrocnemius. Studies were performed at 3 (3-mo), 6 (6-mo) and 9 (9-mo) months of age. Results: MetS rats were more insulin resistant (p<0.001, all ages) and had higher BP (3-mo: p<0.001, 6-mo: p = 0.001, 9-mo: p = 0.015) as compared to C. At 6 months, CRP, IL-6 and TNF-alpha were higher (p<0.001, all comparisons) in MetS rats vs H, but adiponectin was lower in MetS at 9 months (MetS: 32 +/- 2, H: 42 +/- 2, C: 45 +/- 2 pg/mL; p<0.001). GLUT4 protein was reduced in MetS as compared to C rats at 3, 6 and 9-mo, respectively (Heart: 54%, 50% and 57%; Gastrocnemius: 37%, 56% and 50%; Adipose tissue: 69%, 61% and 69%). Conclusions: MSG-treated SHR presented all metabolic syndrome characteristics, as well as reduced GLUT4 content, which must play a key role in the impaired glycemic homeostasis of the metabolic syndrome.

Allelic variations in the vitamin D receptor gene, insulin secretion and parents' heights are independently associated with height in obese children and adolescents

Polymorphisms in the VDR gene were reported to be associated with variations in intrauterine and postnatal growth and with adult height, but also with other traits that are strongly correlated such as the BMI, insulin sensitivity, insulin secretion and hyperglycemia. Here, we assessed the impact of VDR polymorphisms on body height and its interactions with obesity- and glucose tolerance-related traits in obese children and adolescents. We studied 173 prepubertal (Tanner's stage 1) and 146 pubertal (Tanner's stages 2-5) obese children who were referred for a weight-loss program. Three single nucleotide polymorphisms were genotyped: rs1544410 (BsmI), rs7975232 (ApaI) and rs731236 (TaqI). BsmI and TaqI genotypes were significantly associated with height in pubertal children, but the associations did not reach statistical significance in prepubertal children. In stepwise regression analyses, the lean body mass, insulin secretion, BsmI or TaqI genotypes and the father's and the mother's height were independently and positively associated with height in pubertal children. These covariables accounted for 46% of the trait variance. The height of homozygous carriers of the minor allele of BsmI was 0.65 z-scores (4 cm) higher than the height of homozygous carriers of the major allele (P=.0006). Haplotype analyses confirmed the associations of the minor alleles of BsmI and TaqI with increased height. In conclusion, VDR genotypes were significantly associated with height in pubertal obese children. The associations were independent from the effects of confounding traits, such as the body fat mass, insulin secretion, insulin sensitivity and glucose tolerance. (C) 2012 Elsevier Inc. All rights reserved.

Sunflower Oil Supplementation Has Proinflammatory Effects and Does Not Reverse Insulin Resistance in Obesity Induced by High-Fat Diet in C57BL/6Mice

High consumption of polyunsaturated fatty acids, such as sunflower oil has been associated to beneficial effects in plasma lipid profile, but its role on inflammation and insulin resistance is not fully elucidated yet. We evaluated the effect of sunflower oil supplementation on inflammatory state and insulin resistance condition in HFD-induced obese mice. C57BL/ 6 male mice (8 weeks) were divided in four groups: (a) control diet (CD), (b) HFD, (c) CD supplemented with n-6 (CD + n-6), and (d) HFD supplemented with n-6 (HFD + n-6). CD + n-6 and HFD + n-6 were supplemented with sunflower oil by oral gavage at 2 g/ Kg of body weight, three times per week. CD and HFD were supplemented with water instead at the same dose. HFD induced whole andmuscle-specific insulin resistance associated with increased inflammatory markers in insulin-sensitive tissues andmacrophage cells. Sunflower oil supplementation was not efficient in preventing or reducing these parameters. In addition, the supplementation increased pro-inflammatory cytokine production by macrophages and tissues. Lipid profile, on the other hand, was improved with the sunflower oil supplementation in animals fed HFD. In conclusion, sunflower oil supplementation improves lipid profile, but it does not prevent or attenuate insulin resistance and inflammation induced by HFD in C57BL/ 6 mice.

Evaluation of the Effects of a Preoperative 2-Hour Fast With Maltodextrine and Glutamine on Insulin Resistance, Acute-Phase Response, Nitrogen Balance, and Serum Glutathione After Laparoscopic Cholecystectomy: A Controlled Randomized Trial

Background: Prolonged preoperative fasting increases insulin resistance (IR). The authors investigated whether an abbreviated preoperative fast with glutamine (GLN) plus a carbohydrate (CHO)-based beverage would improve the organic response after surgery. Methods: Forty-eight female patients (19-62 years) were randomized to either standard fasting (control group) or to fasting with 1 of 3 different beverages before video-cholecystectomy. Beverages were consumed 8 hours (400 mL; placebo group: water; GLN group: water with 50 g maltodextrine plus 40 g GLN; and CHO group: water with 50 g maltodextrine) and 2 hours (200 mL; placebo: water; GLN: water with 25 g maltodextrine plus 10 g GLN; and CHO: water with 25 g maltodextrine) before anesthesia. Blood samples were collected pre- and postoperatively. Results: The mean (SEM) postoperative homeostasis model assessment-insulin resistance was greater (P < .05) in control patients (4.3 [1.3]) than in the other groups (placebo, 1.6 [0.3]; CHO, 2.3 [0.4]; and GLN, 1.5 [0.1]). Glutathione was significantly higher (P < .01) in the GLN group than in both CHO and control groups. Interleukin-6 increased in all groups except the GLN group. The C-reactive protein/albumin ratio was higher (P < .05) in controls than in CHO and GLN groups. The nitrogen balance was less negative in GLN (-2.5 [0.8] gN) than in both placebo (-9.0 [2] gN; P = .001) and control (-6.6 [0.4] gN; P = .04) groups. Conclusions Preoperative intake of a GLN-enriched CHO beverage appears to improve IR and antioxidant defenses and decreases the inflammatory response after video-cholecystectomy. (JPEN J Parenter Enteral Nutr. 2012; 36: 43-52)

Up-regulation of fas and fasL pro-apoptotic genes expression in type 1 diabetes patients after autologous haematopoietic stem cell transplantation

Type 1 diabetes (T1D) is a chronic autoimmune disease characterized by T cell-mediated destruction of pancreatic beta cells, resulting in insulin deficiency and hyperglycaemia. Recent studies have described that apoptosis impairment during central and peripheral tolerance is involved in T1D pathogenesis. In this study, the apoptosis-related gene expression in T1D patients was evaluated before and after treatment with high-dose immunosuppression followed by autologous haematopoietic stem cell transplantation (HDI-AHSCT). We also correlated gene expression results with clinical response to HDI-AHSCT. We observed a decreased expression of bad, bax and fasL pro-apoptotic genes and an increased expression of a1, bcl-xL and cIAP-2 anti-apoptotic genes in patients' peripheral blood mononuclear cells (PBMCs) compared to controls. After HDI-AHSCT, we found an up-regulation of fas and fasL and a down-regulation of anti-apoptotic bcl-xL genes expression in post-HDI-AHSCT periods compared to pre-transplantation. Additionally, the levels of bad, bax, bok, fasL, bcl-xL and cIAP-1 genes expression were found similar to controls 2 years after HDI-AHSCT. Furthermore, over-expression of pro-apoptotic noxa at 540 days post-HDI-AHSCT correlated positively with insulin-free patients and conversely with glutamic acid decarboxylase autoantibodies (GAD65) autoantibody levels. Taken together, the results suggest that apoptosis-related genes deregulation in patients' PBMCs might be involved in breakdown of immune tolerance and consequently contribute to T1D pathogenesis. Furthermore, HDI-AHSCT modulated the expression of some apoptotic genes towards the levels similar to controls. Possibly, the expression of these apoptotic molecules could be applied as biomarkers of clinical remission of T1D patients treated with HDI-AHSCT therapy.

Endurance exercise training increases APPL1 expression and improves insulin signaling in the hepatic tissue of diet-induced obese mice, independently of weight loss

Hepatic insulin resistance is the major contributor to fasting hyperglycemia in type 2 diabetes. The protein kinase Akt plays a central role in the suppression of gluconeogenesis involving forkhead box O1 (Foxo1) and peroxisome proliferator-activated receptor gamma co-activator 1 alpha (PGC-1a), and in the control of glycogen synthesis involving the glycogen synthase kinase beta (GSK3 beta) in the liver. It has been demonstrated that endosomal adaptor protein APPL1 interacts with Akt and blocks the association of Akt with its endogenous inhibitor, tribbles-related protein 3 (TRB3), improving the action of insulin in the liver. Here, we demonstrated that chronic exercise increased the basal levels and insulin-induced Akt serine phosphorylation in the liver of diet-induced obese mice. Endurance training was able to increase APPL1 expression and the interaction between APPL1 and Akt. Conversely, training reduced both TRB3 expression and TRB3 and Akt association. The positive effects of exercise on insulin action are reinforced by our findings that showed that trained mice presented an increase in Foxo1 phosphorylation and Foxo1/PGC-1a association, which was accompanied by a reduction in gluconeogenic gene expressions (PEPCK and G6Pase). Finally, exercised animals demonstrated increased at basal and insulin-induced GSK3 beta phosphorylation levels and glycogen content at 24?h after the last session of exercise. Our findings demonstrate that exercise increases insulin action, at least in part, through the enhancement of APPL1 and the reduction of TRB3 expression in the liver of obese mice, independently of weight loss. J. Cell. Physiol. 227: 29172926, 2012. (C) 2011 Wiley Periodicals, Inc.

The Regulation of Rasd1 Expression by Glucocorticoids and Prolactin Controls Peripartum Maternal Insulin Secretion

The transition from gestation to lactation is characterized by a robust adaptation of maternal pancreatic beta-cells. Consistent with the loss of beta-cell mass, glucose-induced insulin secretion is down-regulated in the islets of early lactating dams. Extensive experimental evidence has demonstrated that the surge of prolactin is responsible for the morphofunctional remodeling of the maternal endocrine pancreas during pregnancy, but the precise molecular mechanisms by which this phenotype is rapidly reversed after delivery are not completely understood. This study investigated whether glucocorticoid-regulated expression of Rasd1/Dexras, a small inhibitoryGprotein, is involved in this physiological plasticity. Immunofluorescent staining demonstrated that Rasd1 is localized within pancreatic beta-cells. Rasd1 expression in insulin-secreting cells was increased by dexamethasone and decreased by prolactin. In vivo data confirmed that Rasd1 expression is decreased in islets from pregnant rats and increased in islets from lactating mothers. Knockdown of Rasd1 abolished the inhibitory effects of dexamethasone on insulin secretion and the protein kinase A, protein kinase C, and ERK1/2 pathways. Chromatin immunoprecipitation experiments revealed that glucocorticoid receptor (GR) and signal transducer and activator of transcription 5b (STAT5b) cooperatively mediate glucocorticoid-induced Rasd1 expression in islets. Prolactin inhibited the stimulatory effect of GR/STAT5b complex on Rasd1 transcription. Overall, our data indicate that the stimulation of Rasd1 expression by glucocorticoid at the end of pregnancy reverses the increased insulin secretion that occurs during pregnancy. Prolactin negatively regulates this pathway by inhibiting GR/STAT5b transcriptional activity on the Rasd1 gene. (Endocrinology 153: 3668-3678, 2012)

Maternal Moderate Physical Training during Pregnancy Attenuates the Effects of a Low-Protein Diet on the Impaired Secretion of Insulin in Rats: Potential Role for Compensation of Insulin Resistance and Preventing Gestational Diabetes Mellitus

The effects of pregestational and gestational low-to-moderate physical training on insulin secretion in undernourished mothers were evaluated. Virgin female Wistar rats were divided into four groups as follows: control (C, n = 5); trained (T, n = 5); low-protein diet (LP, n = 5); trained with a low-protein diet (T + LP, n = 5). Trained rats ran on a treadmill over a period of 4 weeks before mate (5 days week(-1) and 60 min day(-1), at 65% of VO2max). At pregnancy, the intensity and duration of the exercise were reduced. Low-protein groups were provided with an 8% casein diet, and controls were provided with a 17% casein diet. At third day after delivery, mothers and pups were killed and islets were isolated by collagenase digestion of pancreas and incubated for a further 1 h with medium containing 5.6 or 16.7 mM glucose. T mothers showed increased insulin secretion by isolated islets incubated with 16.7 mM glucose, whereas LP group showed reduced secretion of insulin by isolated islets when compared with both C and LP + T groups. Physical training before and during pregnancy attenuated the effects of a low-protein diet on the secretion of insulin, suggesting a potential role for compensation of insulin resistance and preventing gestational diabetes mellitus.

Metabolic Disorders and Adipose Tissue Insulin Responsiveness in Neonatally STZ-Induced Diabetic Rats Are Improved by Long-Term Melatonin Treatment

Diabetes mellitus is a product of low insulin sensibility and pancreatic beta-cell insufficiency. Rats with streptozotocin-induced diabetes during the neonatal period by the fifth day of age develop the classic diabetic picture of hyperglycemia, hypoinsulinemia, polyuria, and polydipsia aggravated by insulin resistance in adulthood. In this study, we investigated whether the effect of long-term treatment with melatonin can improve insulin resistance and other metabolic disorders in these animals. At the fourth week of age, diabetic animals started an 8-wk treatment with melatonin (1 mg/kg body weight) in the drinking water at night. Animals were then killing, and the sc, epididymal (EP), and retroperitoneal (RP) fat pads were excised, weighed, and processed for adipocyte isolation for morphometric analysis as well as for measuring glucose uptake, oxidation, and incorporation of glucose into lipids. Blood samples were collected for biochemical assays. Melatonin treatment reduced hyperglycemia, polydipsia, and polyphagia as well as improved insulin resistance as demonstrated by constant glucose disappearance rate and homeostasis model of assessment-insulin resistance. However, melatonin treatment was unable to recover body weight deficiency, fat mass, and adipocyte size of diabetic animals. Adiponectin and fructosamine levels were completely recovered by melatonin, whereas neither plasma insulin level nor insulin secretion capacity was improved in diabetic animals. Furthermore, melatonin caused a marked delay in the sexual development, leaving genital structures smaller than those of nontreated diabetic animals. Melatonin treatment improved the responsiveness of adipocytes to insulin in diabetic animals measured by tests of glucose uptake (sc, EP, and RP), glucose oxidation, and incorporation of glucose into lipids (EP and RP), an effect that seems partially related to an increased expression of insulin receptor substrate 1, acetyl-coenzyme A carboxylase and fatty acid synthase. In conclusion, melatonin treatment was capable of ameliorating the metabolic abnormalities in this particular diabetes model, including insulin resistance and promoting a better long-term glycemic control. (Endocrinology 153: 2178-2188, 2012)

Carbohydrate- and lipid-enriched meals acutely disrupt glycemic homeostasis by inducing transient insulin resistance in rats

Chronic intake of high-carbohydrate or high-lipid diets is a well-known insulin resistance inducer. This study investigates the immediate effect (1-6 h) of a carbohydrate-or lipid-enriched meal on insulin sensitivity. Fasted rats were refed with standard, carbohydrate-enriched (C), or lipid-enriched (L) meal. Plasma insulin, glucose, and non-esterified fatty acids (NEFA) were measured at 1, 2, 4, and 6 h of refeeding. The glucose-insulin index showed that either carbohydrates or lipids decreased insulin sensitivity at 2 h of refeeding. At this time point, insulin tolerance tests (ITTs) and glucose tolerance tests (GTTs) detected insulin resistance in C rats, while GTT confirmed it in L rats. Reduced glycogen and phosphorylated AKT and GSK3 content revealed hepatic insulin resistance in C rats. Reduced glucose uptake in skeletal muscle subjected to the fatty acid concentration that mimics the high NEFA level of L rats suggests insulin resistance in these animals is mainly in muscle. In conclusion, carbohydrate-or lipid-enriched meals acutely disrupt glycemic homeostasis, inducing a transient insulin resistance, which seems to involve liver and skeletal muscle, respectively. Thus, the insulin resistance observed when those types of diets are chronically consumed may be an evolution of repeated episodes of this transient insulin resistance.

Lyophilized bovine colostrum as a source of immunoglobulins and insulin-like growth factor for newborn goat kids

The objective of this study was to evaluate lyophilized bovine colostrum as an alternative source of passive immunity and insulin like growth factor I (IGF-I) for goat kids, considering newborns consuming non-maternal colostrum. Twenty-nine male newborns received 5% of body weight of lyophilized bovine (LBC) or goat colostrum (GC), both with 55 mg/mL of IgG, at 0, 7 and 14 h of life. Blood samples were collected at 0, 7, 14, 18, 24, 36, 48, 72 and 96 h of life to determine serum IgG, total protein (TP), IGF-I and apparent efficiency of IgG absorption at 7, 14, 18 and 24 h (AEA(7hr), AEA(14hr), AEA(18hr), AEA(total), respectively). In LBC, the values of serum IgG at 14, 18,24 and 48 h (13.1, 13.4, 14.1 and 14.6 mg/mL, respectively) were higher than the values at 0 and 7 h (0.04 and 6.9 mg/mL, respectively). In GC, the serum IgG at 18 h (9.3 mg/mL) was higher than the value at 7 h (5.5 mg/mL). AEA(7hr) and AEA(14hr) in LBC were the same (19.2 and 18.5%, respectively, P>0.05) and the values of AEA(18hr) and AEA(total), 9.3 and 9.5%, respectively, were equal and smaller than AEA(7hr), and AEA(14hr). In GC, AEA(7hr), 20.8%, was higher than AEA(14hr), 16.1% (P<0.05) and AEA(18hr), and AEA(total), 9.2 and 8.0%, respectively, were equal and smaller than AEA(7hr) and AEA(14hr). The serum TP and IGF-I were not affected by colostrum feeding. Considering the variables study in the present work, lyophilized bovine colostrum constitutes a promising alternative substitute to goat colostrum in newborn goat kids, since the supply of immunoglobulins and IGF-I was suitable for the kids. (c) 2012 Elsevier B.V. All rights reserved.

Glucose Control in Acute Myocardial Infarction: A Pilot Randomized Study Controlled by Continuous Glucose Monitoring System Comparing the Use of Insulin Glargine with Standard of Care

Background: This pilot study aimed to verify if glycemic control can be achieved in type 2 diabetes patients after acute myocardial infarction (AMI), using insulin glargine (iGlar) associated with regular insulin (iReg), compared with the standard intensive care unit protocol, which uses continuous insulin intravenous delivery followed by NPH insulin and iReg (St. Care). Patients and Methods: Patients (n = 20) within 24 h of AMI were randomized to iGlar or St. Care. Therapy was guided exclusively by capillary blood glucose (CBG), but glucometric parameters were also analyzed by blinded continuous glucose monitoring system (CGMS). Results: Mean glycemia was 141 +/- 39 mg/dL for St. Care and 132 +/- 42 mg/dL for iGlar by CBG or 138 +/- 35 mg/dL for St. Care and 129 +/- 34 mg/dL for iGlar by CGMS. Percentage of time in range (80-180 mg/dL) by CGMS was 73 +/- 18% for iGlar and 77 +/- 11% for St. Care. No severe hypoglycemia (<= 40 mg/dL) was detected by CBG, but CGMS indicated 11 (St. Care) and seven (iGlar) excursions in four subjects from each group, mostly in sulfonylurea users (six of eight patients). Conclusions: This pilot study suggests that equivalent glycemic control without increase in severe hyperglycemia may be achieved using iGlar with background iReg. Data outputs were controlled by both CBG and CGMS measurements in a real-life setting to ensure reliability. Based on CGMS measurements, there were significant numbers of glycemic excursions outside of the target range. However, this was not detected by CBG. In addition, the data indicate that previous use of sulfonylurea may be a potential major risk factor for severe hypoglycemia irrespective of the type of insulin treatment.

Progression to Adrenocortical Tumorigenesis in Mice and Humans through Insulin-Like Growth Factor 2 and beta-Catenin

Dysregulation of the WNT and insulin-like growth factor 2 (IGF2) signaling pathways has been implicated in sporadic and syndromic forms of adrenocortical carcinoma (ACC). Abnormal beta-catenin staining and CTNNB1 mutations are reported to be common in both adrenocortical adenoma and ACC, whereas elevated IGF2 expression is associated primarily with ACC. To better understand the contribution of these pathways in the tumorigenesis of ACC, we examined clinicopathological and molecular data and used mouse models. Evaluation of adrenal tumors from 118 adult patients demonstrated an increase in CTNNB1 mutations and abnormal beta-catenin accumulation in both adrenocortical adenoma and ACC. In ACC, these features were adversely associated with survival. Mice with stabilized beta-catenin exhibited a temporal progression of increased adrenocortical hyperplasia, with subsequent microscopic and macroscopic adenoma formation. Elevated Igf2 expression alone did not cause hyperplasia. With the combination of stabilized beta-catenin and elevated Igf2 expression, adrenal glands were larger, displayed earlier onset of hyperplasia, and developed more frequent macroscopic adenomas (as well as one carcinoma). Our results are consistent with a model in which dysregulation of one pathway may result in adrenal hyperplasia, but accumulation of a second or multiple alterations is necessary for tumorigenesis. (Ant J Pathol 2012, 181:1017-1033; http://dx.doi.org/10.1016/j.ajpath.2012.05.026)