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OBJECTIVE Islets after kidney transplantation have been shown to positively affect the quality of life of individuals with type I diabetes (T1D) by reducing the burden of diabetes complications, but [...]

Diabetes is characterized by accelerated atherosclerosis, although molecular mechanisms explaining this phenomenon are still undefined (1,2). We and others have shown that chronic hyperglycemia increases matrix metalloproteinase (MMP) and A [...], OBJECTIVE--Atherosclerosis is accelerated in subjects with type 2 diabetes by unknown mechanisms. We identified tissue inhibitor of metalloproteinase 3 (TIMP3), the endogenous inhibitor of A disintegrin and metalloprotease domain 17 (ADAM17) and other matrix metalloproteinases (MMPs), as a gene modifier for insulin resistance and vascular inflammation in mice. We tested its association with atherosclerosis in subjects with type 2 diabetes and identified Sirtuin 1 (SirT1) as a major regulator of TIMP3 expression. RESEARCH DESIGN AND METHODS--We investigated ADAM10, ADAM17, MMP9, TIMP1, TIMP2, TIMP3, and TIMP4 expression levels in human carotid atherosclerotic plaques (n = 60) from subjects with and without diabetes. Human vascular smooth muscle cells exposed to several metabolic stimuli were used to identify regulators of TIMP3 expression. SirT1 small interference RNA, cDNA, and TIMP3 promoter gene reporter were used to study SirT1-dependent regulation of TIMP3. RESULTS--Here, we show that in human carotid atherosclerotic plaques, TIMP3 was significantly reduced in subjects with type 2 diabetes, leading to ADAM17 and MMP9 overactivity. Reduced expression of TIMP3 was associated in vivo with SirT1 levels. In smooth muscle cells, inhibition of SirT1 activity and levels reduced TIMP3 expression, whereas SirT1 overexpression increased TIMP3 promoter activity. CONCLUSIONS--In atherosclerotic plaques from subjects with type 2 diabetes, the deregulation of ADAM17 and MMP9 activities is related to inadequate expression of TIMP3 via SifT1. Studies in vascular cells confirmed the role of SirT1 in tuning TIMP3 expression.

OBJECTIVE--Fibroblast growth factor (FGF)-21 is highly expressed in the liver and regulates hepatic glucose production and lipid metabolism in rodents. However, its role in the pathogenesis of type 2 diabetes [...]

OBJECTIVE--To quantitate insulin sensitivity in lean and obese nondiabetic baboons and examine the underlying cellular/ molecular mechanisms responsible for impaired insulin action to characterize a baboon model of insulin resistance. RESEARCH DESIGN AND METHODS--Twenty baboons received a hyperinsulinemic-euglycemic clamp with skeletal muscle and visceral adipose tissue biopsies at baseline and at 30 and 120 min after insulin. Genes and protein expression of key molecules involved in the insulin signaling cascade (insulin receptor, insulin receptor substrate-1, p85, phosphatidylinositol 3-kinase, Akt, and AS160) were sequenced, and insulin-mediated changes were analyzed. RESULTS--Overall, baboons show a wide range of insulin sensitivity (6.2 ± 4.8 mg x [kg.sup.-1] x [min.sup.-1]), and there is a strong inverse correlation between indexes of adiposity and insulin sensitivity (r = -0.946, P < 0.001 for % body fat; r = -0.72, P < 0.001 for waist circumference). The genes and protein sequences analyzed were found to have ~98% identity to those of man. Insulin-mediated changes in key signaling molecules were impaired both in muscle and adipose tissue in obese insulin-resistant compared with lean insulin-sensitive baboons. CONCLUSIONS--The obese baboon is a pertinent nonhuman primate model to examine the underlying cellular/molecular mechanisms responsible for insulin resistance and eventual development of type 2 diabetes. Diabetes 57:899-908, 2008, Insulin resistance is characterized by impaired response of target organs (e.g., skeletal muscle, liver, adipose tissue, and heart) to the physiological effects of insulin and results in impaired glucose metabolism. [...]

Aquaporin 7 (AQP7), the gateway protein controlling glycerol release, has recently emerged as a modulator of adipocyte metabolism. AQP7 knockout mice develop obesity and hyperglycemia. The contribution of AQP7 to these abnormalities in humans is unknown. We examined whether common single nucleotide polymorphisms (SNPs) in the AQP7 gene modulate the risk of obesity and related abnormalities. Among several SNPs we identified, A-953G in the AQP7 promoter was associated with type 2 diabetes in 977 (530 female/447 male) Caucasians: odds ratio for XG (i.e., AG+GG) versus AA individuals was 1.36 (95% CI 1.01-1.84), P = 0.04. This finding was entirely due to the association among females (1.8 [1.2-2.6], P = 0.004), which was no longer significant when adjusted for BMI. In fact, BMI was higher in XG than in AA females (30.8 ± 6.6 vs. 28.9 ± 5.2, P = 0.002). This association was confirmed in independent case-control study (n = 299 female subjects) for morbid obesity (1.66 [1.01-2.74], P = 0.04). Luciferase and mobility shift assays showed that, compared with -953A, the -953G promoter had reduced transcriptional activity (P = 0.001) and impaired ability to bind CCAAT/ enhancer binding protein (C/EBP)β transcription factor (P = 0.01). Finally, AQP7 expression in adipose tissue decreased from AA to AG to GG individuals (P = 0.036). These data strongly suggest that AQP7 downregulation is pathogenic for obesity and/or type 2 diabetes., Obesity and related abnormalities, including type 2 diabetes and dyslipidemia, are becoming epidemic, thus representing major health care problems (1). Although these abnormalities have a clear genetic component, the involved [...]

Obesity (often complicated by type 2 diabetes), arterial hypertension, and hyperlipidemia show insulin resistance (1), metabolic abnormalities (triglycerides, insulin and blood glucose levels, and HDL cholesterol levels [2]), subclinical inflammation [...]

OBJECTIVE--Lifestyle modifications and pharmacological interventions can prevent type 2 diabetes in obese subjects with impaired glucose tolerance. The aim of this study was to compare laparoscopic adjustable gastric banding (LAGB) [...]

OBJECTIVE--The C-174G promoter polymorphism of the interleukin (IL)-6 gene was found to influence transcriptional activity and plasma IL-6 levels in humans. We addressed the question of whether the C-174G IL-6 [...]

OBJECTIVE--Cardiovascular mortality and morbidity are major problems in type 1 diabetic patients with end-stage renal disease (ESRD). The aim of this study was to determine whether islet transplantation can improve [...]

********** OBJECTIVE--Diabetes, hypertension, infections, and nephrotoxicity of certain immunosuppressive drugs (i.e., calcineurin inhibitors) can reduce functional survival of the kidney graft. Our aim was to evaluate survival, hypertrophy, and vascular [...]

To evaluate the effects of kidney-pancreas transplantation on hemostatic abnormalities in uremic type 1 diabetic patients, we conducted a cross-sectional study involving 12 type 1 diabetic patients, 30 uremic type 1 diabetic patients, 27 uremic type 1 diabetic patients who had a kidney-pancreas transplant, 12 uremic type 1 diabetic patients who had a kidney-alone transplant and 13 healthy control subjects. We evaluated platelet and clotting system. Platelets in the group of uremic type 1 diabetic patients were significantly larger than platelets in the other groups. Resting calcium levels were significantly higher in the uremic type 1 diabetic patients and uremic type 1 diabetic patients who had kidney-alone transplant than in the type 1 diabetic patients who had a kidney-pancreas transplant and control subjects. CD41 expression was significantly reduced in platelets from the uremic type 1 diabetic patients compared with the other groups. Levels of hypercoagulability markers in the type 1 diabetic patients who had a kidney-pancreas transplant and, to lesser extent, the uremic type 1 diabetic patients who had a kidney-alone transplant but not the uremic type 1 diabetic patients were similar to those of the control subjects. A reduction in natural anticoagulants was evident in the uremic type 1 diabetic patients, whereas near-normal values were observed in the type 1 diabetic patients who had a kidney-pancreas transplant and uremic type 1 diabetic patients who had a kidney-alone transplant. Hemostatic abnormalities were not ob served in type 1 diabetic patients who had a kidney pancreas transplant. This finding might explain the lower cardiovascular death rate observed in type 1 diabetic patients who had a kidney-pancreas transplant compared with uremic type 1 diabetic patients who had a kidney-alone transplant or uremic type 1 diabetic patients., Hemostatic abnormalities are commonly observed in patients who are affected by type 1 diabetes and in uremic patients (1-10). Enhanced activation of the clotting system has been implicated in the [...]

Uncoupling protein (UCP)-2 is a member of the mitochondrial inner membrane carriers that uncouple proton entry in the mitochondrial matrix from ATP synthesis. The -866G/A polymorphism in the UCP2 gene, which enhances its transcriptional activity, was associated with enhanced risk for type 2 diabetes in obese subjects. We addressed the question of whether the -866G/A polymorphism contributes to variation in insulin sensitivity by genotyping 181 nondiabetic offspring of type 2 diabetic patients. Insulin sensitivity, assessed by the hyperinsulinemic-euglycemic clamp, was reduced in -866A/A carriers compared with -866A/G or -866G/G carriers (P = 0.01). To directly investigate the correlation between UCP2 expression and insulin resistance, UCP2 mRNA levels were measured by real-time RT-PCR in subcutaneous fat obtained from 100 obese subjects who underwent laparoscopic adjustable gastric banding. UCP2 mRNA expression was significantly correlated with insulin resistance as assessed by the homeostasis model assessment index (r = 0.27, P = 0.007). We examined the association of the -866A/A genotype in a case-control study including 483 type 2 diabetic subjects and 565 control subjects. The -866A/A genotype was associated with diabetes in women (odds ratio 1.84, 95% CI 1.03-3.28; P = 0.037), but not in men. These results indicate that the -866A/A genotype of the UCP2 gene may contribute to diabetes susceptibility by affecting insulin sensitivity., The pathophysiology of type 2 diabetes includes two apparently distinct defects, i.e., impairments in insulin action at the level of skeletal muscle, fat, and liver and inadequate compensation by the [...]

Several different tools have been developed to integrate the clinical and biochemical nutritional evaluations in critical care patients. Aims of this study were to evaluate the changes in the Bioelectrical Impedance Vector Analysis (BIVA) and ultrasonographic features of the diaphragm (DTee) and rectus femoris (RFCSA) during the first week of ICU stay.

Type 2 diabetes is characterized by insulin resistance and inadequate insulin secretion. In the advanced stages of the disease, β-cell dysfunction worsens and insulin therapy may be necessary to achieve [...]

A 51-year-old woman affected by advanced aggressive fibromatosis of the abdomen was admitted to the hospital because of diabetes associated with severe insulin resistance. Her medical history was notable for [...]

Type 2 diabetes is characterized by inadequate compensation of pancreatic [Beta]-cells for peripheral insulin resistance. Recent studies suggested that [Beta]-cell failure plays a dominating role in the establishment of type [...]

White blood cell (WBC) count is elevated in obesity (1) and is a risk factor for atherosclerosis (2). An elevated WBC count is present in impaired glucose tolerance (IGT) (3), [...]

AbstractBackgroundAtherosclerotic cardiovascular disease (ASCVD) is the leading cause of morbidity and mortality. Hyperlipidemia and vascular subinflammation play critical roles in the pathogenesis of atherosclerosis. Patients with Major Depressive Disorder (MDD) are at higher risk for ASCVD and current antidepressant therapies may carry ASCVD risks. REL-1017 is a novel NMDAR channel blocker which showed rapid, robust, and sustained antidepressant effects, currently in Phase 3 clinical trials for MDD.MethodsWe analyzed total cholesterol (TC), triglycerides (TG), Proprotein Convertase Subtilisin/Kexin 9 (PCSK9), and high-sensitivity C-reactive protein (hs-CRP) from patients enrolled in a Phase-2, multicenter, randomized, double-blind, placebo-controlled, 7-day, 3-arm trial to assess safety, tolerability, pharmacokinetics, and efficacy of REL-1017. Patients were randomized in a 1:1:1 ratio to either placebo, REL-1017 25 mg QD, or REL-1017 50 mg QD. TC, TG, PCSK9, and hs-CRP levels were measured in patients at baseline, day 7 and day 14, 7 days after treatment discontinuation. 6 out of 21 (28% of patients), 6 out of 16 (37%), and 1 out of 19 (5%), were under statin therapy in the placebo, REL-1017 25 and 50 mg groups, respectively.ResultsAt baseline, day 7, day 14 TC levels (mg/dL) were 117.8 ± 30.8, 124.7 ± 34.1, 114.2 ± 18.1; 123.7 ± 59.3, 119.0 ± 28.1, 115.1 ± 14.8; 113.9 ± 22.9, 118.6 ± 29.5, 115.8 ± 25.5 for placebo (n = 21), REL-1017 25 mg (n = 16) and REL-1017 50 mg (n = 19), respectively. Considering the subgroup not on statins, TC levels were 127.5 ± 28.6, 134.2 ± 35.0, 117.8 ± 14.8; 131.0 ± 71.7, 121.1 ± 32.9, 118.6 ± 15.4; 115.5 ± 22.1, 121.4 ± 27.1, 119.0 ± 21.9 for placebo (n = 15, 72% of patients), REL-1017 25 mg (n = 10, 63%) and REL-1017 50 mg (n = 18, 95%), respectively. TG were 57.0 ± 25.6, 55.7 ± 20.0, 58.0 ± 34.1; 62.7 ± 52.1, 56.0 ± 31.0, 59.5 ± 32.6; 48.5 ± 25.3, 50.2 ± 18.4, 55.1 ± 21.9 for placebo (n = 21), REL-1017 25 mg (n = 16) and REL-1017 50 mg (n = 19), respectively. Considering the group not on statins, TG were 52.9 ± 27.6, 55.7 ± 23.1, 51.3 ± 26.8; 70.6 ± 63.3, 57.6 ± 38.9, 65.2 ± 38.9; 47.4 ± 25.4, 48.9 ± 17.9, 52.6 ± 19.5 for placebo (n = 15), REL-1017 25 mg (n = 10), and REL-1017 50 mg (n = 18), respectively. Levels of PCSK9, a key player of LDL cholesterol levels, significantly (P< .05) increased from baseline to day 7 and did not further change by day 14 for placebo, with similar results for REL-1017 25 or 50 mg groups, suggesting fluctuations unrelated to treatment. A total of 30% of the patients had hs-CRP plasma levels higher than 2 mg/L, thus potentially associated with a higher incidence of CV events. However, 7 days of treatment with REL-1017 did not alter hs-CRP plasma levels, neither at 25 mg/day nor at 50 mg/day. In summary, REL-1017 of 25 or 50 mg for 7 days did not affect TC, TG, PCSK9 and hs-CRP levels.ConclusionA 7-day treatment course with REL-1017 did not significantly alter TC, TG, PCSK9, or hs-CRP, suggesting the absence of detrimental effects on ASCVD risk. These data should be confirmed in longer and larger trials.FundingRelmada Therapeutics, Inc.

Oxidative stress contributes to the pathogenesis of protein-energy wasting in maintenance hemodialysis (MHD) patients, but knowledge of specific effectors and mechanisms remains fragmented. Aim of the study was to define whether and how food intake is involved in the causal relationship between oxidative stress and protein-energy wasting.

The role of circulating factors in regulating colonic stem cells (CoSCs) and colonic epithelial homeostasis is unclear. Individuals with long-standing type 1 diabetes (T1D) frequently have intestinal symptoms, termed diabetic enteropathy (DE), though its etiology is unknown. Here, we report that T1D patients with DE exhibit abnormalities in their intestinal mucosa and CoSCs, which fail to generate in vitro mini-guts. Proteomic profiling of T1D+DE patient serum revealed altered levels of insulin-like growth factor 1 (IGF-I) and its binding protein 3 (IGFBP3). IGFBP3 prevented in vitro growth of patient-derived organoids via binding its receptor TMEM219, in an IGF-I-independent manner, and disrupted in vivo CoSC function in a preclinical DE model. Restoration of normoglycemia in patients with long-standing T1D via kidney-pancreas transplantation or in diabetic mice by treatment with an ecto-TMEM219 recombinant protein normalized circulating IGF-I/IGFBP3 levels and reestablished CoSC homeostasis. These findings demonstrate that peripheral IGF-I/IGFBP3 controls CoSCs and their dysfunction in DE.

Obesity is accompanied by the activation of low-grade inflammatory activity in metabolically relevant tissues. Studies have shown that obesity-associated insulin resistance results from the inflammatory targeting and inhibition of key proteins of the insulin-signaling pathway. At least three apparently distinct mechanisms–endoplasmic reticulum stress, toll-like receptor (TLR) 4 activation, and changes in gut microbiota–have been identified as triggers of obesity-associated metabolic inflammation; thus, they are expected to represent potential targets for the treatment of obesity and its comorbidities. Here, we review the data that place TLR4 in the center of the events that connect the consumption of dietary fats with metabolic inflammation and insulin resistance. Changes in the gut microbiota can lead to reduced integrity of the intestinal barrier, leading to increased leakage of lipopolysaccharides and fatty acids, which can act upon TLR4 to activate systemic inflammation. Fatty acids can also trigger endoplasmic reticulum stress, which can be further stimulated by cross talk with active TLR4. Thus, the current data support a connection among the three main triggers of metabolic inflammation, and TLR4 emerges as a link among all of these mechanisms.

Obesity is a major risk factor for asthma, which is characterized by airway hyperreactivity (AHR). In obesity-associated asthma, AHR may be regulated by non-TH2 mechanisms. We hypothesized that airway reactivity is regulated by insulin in the CNS, and that the high levels of insulin associated with obesity contribute to AHR. We found that intracerebroventricular (ICV)-injected insulin increases airway reactivity in wild-type, but not in vesicle acetylcholine transporter knockdown (VAChT KDHOM−/−), mice. Either neutralization of central insulin or inhibition of extracellular signal-regulated kinases (ERK) normalized airway reactivity in hyperinsulinemic obese mice. These effects were mediated by insulin in cholinergic nerves located at the dorsal motor nucleus of the vagus (DMV) and nucleus ambiguus (NA), which convey parasympathetic outflow to the lungs. We propose that increased insulin-induced activation of ERK in parasympathetic pre-ganglionic nerves contributes to AHR in obese mice, suggesting a drug-treatable link between obesity and asthma.

In this study, we aimed to evaluate the effects of exenatide (EXE) treatment on exocrine pancreas of nonhuman primates. To this end, 52 baboons (Papio hamadryas) underwent partial pancreatectomy, followed by continuous infusion of EXE or saline (SAL) for 14 weeks. Histological analysis, immunohistochemistry, Computer Assisted Stereology Toolbox morphometry, and immunofluorescence staining were performed at baseline and after treatment. The EXE treatment did not induce pancreatitis, parenchymal or periductal inflammatory cell accumulation, ductal hyperplasia, or dysplastic lesions/pancreatic intraepithelial neoplasia. At study end, Ki-67–positive (proliferating) acinar cell number did not change, compared with baseline, in either group. Ki-67–positive ductal cells increased after EXE treatment (P = 0.04). However, the change in Ki-67–positive ductal cell number did not differ significantly between the EXE and SAL groups (P = 0.13). M-30–positive (apoptotic) acinar and ductal cell number did not change after SAL or EXE treatment. No changes in ductal density and volume were observed after EXE or SAL. Interestingly, by triple-immunofluorescence staining, we detected c-kit (a marker of cell transdifferentiation) positive ductal cells co-expressing insulin in ducts only in the EXE group at study end, suggesting that EXE may promote the differentiation of ductal cells toward a β-cell phenotype. In conclusion, 14 weeks of EXE treatment did not exert any negative effect on exocrine pancreas, by inducing either pancreatic inflammation or hyperplasia/dysplasia in nonhuman primates.

Diabetes mellitus is associated to an increased risk of cardiovascular diseases. Hyperglycemia is an important factor in cardiovascular damage, working through different mechanisms such as activation of protein kinase C, polyol and hexosamine pathways, advanced glycation end products production. All of these pathways, in association to hyperglycemia-induced mitochondrial dysfunction and endoplasmic reticulum stress, promote reactive oxygen species (ROS) accumulation that, in turn, promote cellular damage and contribute to the diabetic complications development and progression. ROS can directly damage lipids, proteins or DNA and modulate intracellular signaling pathways, such as mitogen activated protein kinases and redox sensitive transcription factors causing changes in protein expression and, therefore, irreversible oxidative modifications. Hyperglycemia-induced oxidative stress induces endothelial dysfunction that plays a central role in the pathogenesis of micro- and macro-vascular diseases. It may also increase pro-inflammatory and pro-coagulant factors expression, induce apoptosis and impair nitric oxide release. Oxidative stress induces several phenotypic alterations also in vascular smooth-muscle cell (VSMC). ROS is one of the factors that can promote both VSMC proliferation/migration in atherosclerotic lesions and VSMC apoptosis, which is potentially involved in atherosclerotic plaque instability and rupture. Currently, there are contrasting clinical evidences on the benefits of antioxidant therapies in the prevention/treatment of diabetic cardiovascular complications. Appropriate glycemic control, in which both hypoglycemic and hyperglycemic episodes are reduced, in association to the treatment of dyslipidemia, hypertension, kidney dysfunction and obesity, conditions which are also associated to ROS overproduction, can counteract oxidative stress and, therefore, both microvascular and macrovascular complications of diabetes mellitus.

Cardiometabolic risk comprises a cluster of traditional and emerging factors that are good indicators of a patients overall risk for type 2 diabetes and cardiovascular disease. The insulin resistance, a key feature common to obesity and type 2 diabetes, is associated with impaired vascular response and contributes to increased cardiovascular risk. Abnormal vascular insulin signalling induces endothelial dysfunction, the initial step of atherosclerotic process, characterized by attenuated nitric oxide-mediated vasodilatation and atherogenic response. Insulin resistance and endothelial dysfunction are two pathological conditions that can co-exist, even if their cause-effect relationship is not yet clarified. Multiple signaling pathways shared by insulin resistance and endothelial dysfunction include hyperinsulinemia, glucotoxicity, lipotoxicity, and inflammation. These mechanisms selectively impair PI3K-dependent insulin in vascular endothelium harming endothelial balance and strengthening the evidence of the close association between metabolic and cardiovascular disease. The present review analyzes the close relationship between endothelial dysfunction and insulin resistance and explores the common mechanisms, with clinical considerations and pharmacological strategies.

The reasons for the long-term complete or partial loss of islet graft function are unknown, but there are obviously other reasons than just pure allogeneic graft rejection. Earlier studies have shown that deposition of islet amyloid polypeptide amyloid in transplanted islets may indicate a mechanism for loss of cells.

Introduction:Obesity is a major health threat in the Western world because of its high incidence and prevalence, and its association with metabolic and cardiovascular disease as well as cancer. The reduction of food intake in obese patients can be achieved only transiently (generally for no longer than 6 months), in the absence of concomitant pharmacological therapy. Only bariatric surgery provides a means to increase satiety and/or decrease nutrient absorption in obese patients, in the long term.Areas covered:This article reviews the available pharmacological treatments for obesity as well as the pharmacology and mechanism of action of exenatide in obese type 2 diabetic patients.Expert opinion:Exenatide is a potential new candidate treatment for obesity, possibly in combination with other hormones that increase satiety (leptin) and slow gastric emptying (amylin).

Obesity, insulin resistance, glucose intolerance/type 2 diabetes and hypertension are clustered in the metabolic syndrome representing critical risk factors for increased incidence cardio-cerebro-vascular diseases, kidney failure and cancer. Ectopic fat accumulation, i.e., accumulation in the mediastinum, liver and the abdomen, as well as generalized fat accumulation are associated with arterial hypertension, either systolic or diastolic. Several mechanisms including insulin resistance, sub-inflammatory state, increased Renin- Angiotensin-Aldosterone System (RAAS) system activity, oxidative stress, autonomic dysregulation as well as mechanical compression on the kidneys are all activated by obesity. Interestingly angiotensin-converting enzyme (ACE) inhibitors and angiotensin II (ATII) receptor blockers, while correcting arterial hypertension, also have a positive effect on glucose metabolism and diabetes prevention, in high risk patients. The implementation of dietary, medical and surgical strategies to prevent and treat obesity, are cornerstones for the primary prevention as well as treatment of arterial hypertension.

A growing body of evidence suggests that oxidative stress plays a key role in the pathogenesis of micro- and macrovascular diabetic complications. The increased oxidative stress in subjects with type 2 diabetes is a consequence of several abnormalities, including hyperglycemia, insulin resistance, hyperinsulinemia, and dyslipidemia, each of which contributes to mitochondrial superoxide overproduction in endothelial cells of large and small vessels as well as the myocardium. The unifying pathophysiological mechanism that underlies diabetic complications could be explained by increased production of reactive oxygen species (ROS) via: (1) the polyol pathway flux, (2) increased formation of advanced glycation end products (AGEs), (3) increased expression of the receptor for AGEs, (4) activation of protein kinase C isoforms, and (5) overactivity of the hexosamine pathway. Furthermore, the effects of oxidative stress in individuals with type 2 diabetes are compounded by the inactivation of two critical anti-atherosclerotic enzymes: endothelial nitric oxide synthase and prostacyclin synthase. Of interest, the results of clinical trials in patients with type 2 diabetes in whom intensive management of all the components of the metabolic syndrome (hyperglycemia, hypercholesterolemia, and essential hypertension) was attempted (with agents that exert a beneficial effect on serum glucose, serum lipid concentrations, and blood pressure, respectively) showed a decrease in adverse cardiovascular end points. The purpose of this review is (1) to examine the mechanisms that link oxidative stress to micro- and macrovascular complications in subjects with type 2 diabetes and (2) to consider the therapeutic opportunities that are presented by currently used therapeutic agents which possess antioxidant properties as well as new potential antioxidant substances.

One of the critical points in the pathogenesis of aortic aneurysms (AAs) is the disruption of the balance between vascular extracellular matrix (ECM) deposition and degradation. AAs are common features in some genetically determined diseases of the connective tissue, such as Marfan and Ehlers-Danlos. Acquired factors determining an enhanced inflammatory state of the arterial wall also play a key role. Previous studies have determined the role of tumor growth factor (TGF-); as a principal mediator of the pathogenesis of the alterations of the arterial wall homeostasis in AAs. The medical management of any AA is mainly focused on the use of pharmacological agents that reduce hemodynamic stress of the aortic wall, since hypertension is the major risk factor for the enlargement and rupture of the AAs. However, this is far from being a comprehensive pathophysiology-based therapeutic approach. Drugs potentially able to reduce the release of TGF- may play a role in the pathogenesis of the AAs. They work by improving matrix repair, decreasing the proteolytic pattern and inhibition of angiotensin-converting enzyme (ACE) as well as preventing angiotensin II-induced angiotensin type-1 receptor (AT1R) activation. A new pathophysiology-based therapeutic approach, involving the mechanisms leading to the rupture of the AAs, could represent an additional tool in combination with the current established antihypertensive therapy.

Tobacco smoking is the most important preventable cause of cardiovascular disease. In this paper we review current epidemiological and pathophysiological evidence linking smoking with cardiovascular and metabolic diseases. Among the effects of smoking there is the alteration of lipid metabolism through the increase in lipolysis, insulin resistance and tissue lipotoxicity. Smoking is both prothrombotic and atherogenic. As an effect, the risk of acute myocardial infarction, sudden cardiac death, stroke, aortic aneurysm and peripheral vascular disease is increased. Even very low doses of exposure increase the risk of cardiovascular disease (CVD) and metabolic alterations. On the other hand, smoking cessation restores, at least in part, lipid metabolism and the benefits can be observed already after a short period of abstinence from smoking, although it occurs several years before the risks approach those of the never-smoker.

To learn more about nonimmune-mediated islet graft failure, we transplanted different preparations (preps) of isolated human islets under the kidney capsule of streptozotocin (STZ)-diabetic nude mice. One month after the implantation of 1,000 or 2,000 islets, grafts were harvested for morphological, immunohistochemical, and ultrastructural analysis. Only a single islet prep cured the diabetes out of all the recipients, while the remaining preps showed only partial function after the implantation of 2,000 islets. Transplanted mice showed high circulating proinsulin levels but, with the exclusion of those bearing curative grafts, relatively low mature insulin levels. Engrafted β-cells showed positive carboxypeptidase E (CPE) and prohormone convertase 1 (PC1) staining, while prohormone convertase 2 (PC2) was undetectable. In contrast, PC2 was abundantly expressed by engrafted α-cells. Moreover, engrafted β-cells did not show evidence of replication, and preapoptotic β-cells, with intra- and extracellular amyloid deposition, were detected with electron microscopy. Cell cycle inhibitors p16INK4, p21WAF1, and p27Kip1were abundantly expressed in the islet grafts and showed a predominant nuclear localization. In conclusion, diabetic nude mice transplanted with human islets showed disproportionate hyperproinsulinemia and graft evidence of β-cell restricted PC2 depletion, amyloid deposition and β-cell death, and lack of β-cell replication with nuclear translocation of p27Kip1and p21WAF1that together may contribute to delayed graft failure.

Essential hypertension is an insulin resistant state. Early insulin signaling steps are impaired in essential hypertension and a large body of data suggests that there is a crosstalk at multiple levels between the signal transduction pathways that mediate insulin and angiotensin II actions. At the extracellular level the angiotensin converting enzyme (ACE) regulates the synthesis of angiotensin II and bradykinin that is a powerful vasodilator. At early intracellular level angiotensin II acts on JAK-2/IRS1-IRS2/PI3-kinase, JNK and ERK to phosphorylate serine residues of key elements of insulin signaling pathway therefore inhibiting signaling by the insulin receptor. On another level angiotensin II inhibits the insulin signaling inducing the regulatory protein SOCS 3. Angiotensin II acting through the AT1 receptor can inhibit insulininduced nitric oxide (NO) production by activating ERK 1/2 and JNK and enhances the activity of NADPH oxidase that leads to an increased reactive oxygen species generation. From the clinical standpoint, the inhibition of the renin angiotensin system improves insulin sensitivity and decreases the incidence of Type 2 Diabetes Mellitus (T2DM). This might represent an alternative approach to prevent type 2 diabetes in patients with hypertension and metabolic syndrome, (i.e. insulin resistant patients). This review will discuss: a) the molecular mechanisms of the crosstalk between the insulin and angiotensin II signaling systems b) the results of clinical studies employing drugs targeting the renin-angiotensin IIaldosterone systems and their role in glucose metabolism and diabetes prevention.

The aim of this study was to evaluate the effects of islet transplantation on patient survival and diabetic vascular complications.

We recently developed two rat pituitary GH3 cell clones engineered to secrete human insulin (InsGH3). InsGH3 cells convert proinsulin into mature insulin, which is partially stored into a readily releasable pool of secretory granules. The efficiency of these processes, however, is relatively low in these cells, either in vitro or in vivo. This study was aimed at determining whether carboxypeptidase E (Cpe) overexpression can increase proinsulin processing and regulated secretion by InsGH3 clones. Indeed, in its membrane-bound form Cpe works as sorting receptor for the regulated secretory pathway of many hormones while, in its soluble form, Cpe takes part to the late step of insulin maturation. We obtained two Cpe-overexpressing cell lines from two different InsGH3 clones (InsGH3/C1 and C7). In the Cpe-overexpressing cell lines, derived from InsGH3 of clone 1 (InsGH3/C1-HACpe), in which the membrane-bound form of exogenous Cpe is accounted for by 90% of total Cpe immunoreactivity, we observed an increase in proinsulin gene expression, and in basal and stimulated insulin secretion compared with the original clone. In contrast, in the Cpe-overexpressing cell line derived from InsGH3 of clone 7 (InsGH3/C7-HACpe), where the exogenous membrane-bound form was only 60% of total Cpe, we detected a decrease in basal insulin release and a modest, albeit significant, increase in intracellular proinsulin processing. In conclusion, Cpe overexpression can increase regulated insulin secretion and proinsulin processing in InsGH3 cells; however, such improvements appear quantitatively and qualitatively modest.

Increase in adipose mass results in obesity and modulation of several factors in white adipose tissue (WAT). Two important examples are tumor necrosis factor α (TNFα) and leptin, both of which are upregulated in adipose tissue in obesity. In order to isolate genes differentially expressed in the WAT of genetically obese db/db mice compared to their lean littermates, we performed RNA fingerprinting and identified haptoglobin (Hp), which is significantly upregulated in the obese animals. Hp is a glycoprotein induced by a number of cytokines, LPS (Lipopolysaccharide), and more generally by inflammation. A significant upregulation of WAT Hp expression was also evident in several experimental obese models including the yellow agouti (/) A^y, ob/ob and goldthioglucose-treated mice (10-, 8-, and 7-fold, respectively). To identify the potential signals for an increase in Hp expression in obesity, we examined leptin and TNFα in vivo. Wild type animals treated with recombinant leptin did not show any alteration in WAT Hp expression compared to controls that were food restricted to the level of intake of the treated animals. On the other hand, Hp expression was induced in mice transgenically expressing TNFα in adipose tissue. Finally, a significant downregulation of WAT Hp mRNA was observed in ob/ob mice deficient in TNFα function, when compared to the ob/ob controls. These results demonstrate that haptoglobin expression in WAT is increased in obesity in rodents and TNFα is an important signal for this regulation. J. Cell. Physiol. 190: 251–258, 2002. © 2002 Wiley-Liss, Inc.

Phosphatidylinositol 3kinase PI3K controls important intracellular steps involved in inflammation, immunity, and cell growth. PI3K also modulates leukocyte integrin adhesiveness. In this study we evaluated the role of PI3K on neutrophil adhesion to intercellular adhesion molecule1 ICAM1transfected cells. Nformylmethionylleucylphenylalanine fMLPstimulated neutrophil adhesion was inhibited by wortmannin and LY294002, two unrelated PI3K inhibitors, whereas phorbol myristate acetate PMAinduced neutrophil adhesion was not inhibited by them. After fMLP stimulation, a rapid activation of AKT and ERK was observed. However, only activation of AKT was reversed by the PI3K inhibitors. Neutrophil expression of the 2integrins Mac1, lymphocyte functionassociated antigen1LFA1, and gp150.95 was not affected by wortmannin, nor was expression of the activation epitope recognized by MAB24. We conclude that a PI3K is involved in fMLPactivated neutrophil adhesion to ICAM1transfected cells, b the mechanism involved is not mediated by the modulation of 2integrin expression or activation, and c another mechanism seems to involve the adhesion to ICAM1 when a cellular system of adhesion is used.

Background & Aims:We investigated the expression of estrogen receptor (ER) α and β subtypes in cholangiocytes of normal and bile duct–ligated (BDL) rats and evaluated the role and mechanisms of estrogens in the modulation of cholangiocyte proliferation. Methods:ER-α and ER-β were analyzed by immunohistochemistry, reverse-transcription polymerase chain reaction, and Western blotting in normal and BDL rats. The effects of the ER antagonists tamoxifen and ICI 182,780 on cholangiocyte proliferation were evaluated. Results:Cholangiocytes expressed both ER-α and ER-β subtypes, whereas hepatocytes expressed only ER-α. In association with a marked cholangiocyte proliferation and with enhanced estradiol serum levels, the immunoreactivity for ER-α involved a 3-fold higher percentage of cholangiocytes in 3-week BDL than in normal rats; immunoreactivity for ER-β showed a 30-fold increase. Western blot analysis showed that during BDL, the total amount of ER-β in cholangiocytes was markedly increased (5-fold), whereas that of ER-α decreased slightly (−25%). Treatment with tamoxifen or ICI 182,780 of 3-week BDL rats inhibited cholangiocyte proliferation and induced overexpression of Fas antigen and apoptosis in cholangiocytes. In vitro, 17β estradiol stimulated proliferation of cholangiocyte, an effect blocked to the same extent by tamoxifen or ICI 182,780. Conclusions:This study suggests that estrogens and their receptors play a role in the modulation of cholangiocyte proliferation.

In a companion article, we describe the engineering and characterization of pituitary GH3 cell clones stably transfected with a furin-cleavable human insulin cDNA (InsGH3 cells). This article describes the performance of InsGH3 (clones 1 and 7) cell grafts into streptozotocin (STZ)-induced diabetic nude mice. Subcutaneous implantation of 2 × 106InsGH3 cells resulted in the progressive reversal of hyperglycemia and diabetic symptoms, even though the progressive growth of the transplanted cells (clone 7) eventually led to glycemic levels below the normal mouse range. Proinsulin transgene expression was maintained in harvested InsGH3 grafts that, conversely, lose the expression of the prolactin (PRL) gene. Elevated concentrations of circulating mature human insulin were detected in graft recipients, demonstrating that proinsulin processing by InsGH3 cells did occur in vivo. Histologic analysis showed that transplanted InsGH3 grew in forms of encapsulated tumors composed of cells with small cytoplasms weakly stained for the presence of insulin. Conversely, intense insulin immunoreactivity was detected in graft-draining venules. Compared to pancreatic βTC3 cells, InsGH3 cells showed in vitro a higher rate of replication, an elevate resistance to apoptosis induced by serum deprivation and proinflammatory cytokines, and significantly higher antiapoptotic Bcl-2 protein levels. Moreover, InsGH3 cells were resistant to the streptozotocin toxicity that, in contrast, reduced βTC3 cell viability to 50–60% of controls. In conclusion, proinsulin gene expression and mature insulin secretion persisted in transplanted InsGH3 cells that reversed hyperglycemia in vivo. InsGH3 cells might represent a potential β-cell surrogate because they are more resistant than pancreatic β cells to different apoptotic insults and might therefore be particularly suitable for encapsulation.

Successful β-cell replacement therapy in insulin-dependent (type I) diabetes is hindered by the scarcity of human donor tissue and by the recurrence of autoimmune destruction of transplanted β cells. Availability of non-β cells, capable of releasing insulin and escaping autoimmune recognition, would therefore be important for diabetes cell therapy. We developed rat pituitary GH3 cells stably transfected with a furin-cleavable human proinsulin cDNA linked to the rat PRL promoter. Two clones (InsGH3/clone 1 and 7) were characterized in vitro with regard to basal and stimulated insulin release and proinsulin transgene expression. Mature insulin secretion was obtained in both clones, accounting for about 40% of total released (pro)insulin-like products. Immunocytochemistry of InsGH3 cells showed a cytoplasmic granular insulin staining that colocalized with secretogranin II (SGII) immunoreactivity. InsGH3 cells/clone 7 contained and released in vitro significantly more insulin than clone 1. Secretagogue-stimulated insulin secretion was observed in both InsGH3 clones either under static or dynamic conditions, indicating that insulin was targeted also to the regulated secretory pathway. Proinsulin mRNA levels were elevated in InsGH3 cells, being significantly higher than in PTC3 cells. Moreover, proinsulin gene expression increased in response to various stimuli, thereby showing the regulation of the transfected gene at the transcriptional level. In conclusion, these data point to InsGH3 cells as a potential β-cell surrogate even though additional engineering is required to instruct them to release insulin in response to physiologic stimulations.

Although nitric oxide (NO) has potent antiplatelet actions, the signaling pathways affected by NO in the platelet are poorly understood. Since NO can induce platelet disaggregation and phosphoinositide 3-kinase (PI3-kinase) activation renders aggregation irreversible, we tested the hypothesis that NO exerts its antiplatelet effects at least in part by inhibiting PI3-kinase. The results demonstrate that the NO donorS-nitrosoglutathione (S-NO-glutathione) inhibits the stimulation of PI3-kinase associated with tyrosine-phosphorylated proteins and of p85/PI3-kinase associated with the SRC family kinase member LYN following the exposure of platelets to thrombin receptor-activating peptide. The activation of LYN-associated PI3-kinase was unrelated to changes in the amount of PI3-kinase physically associated with LYN signaling complexes but did require the activation of LYN and other tyrosine kinases. The cyclic GMP-dependent kinase activator 8-bromo-cyclic GMP had similar effects on PI3-kinase activity, consistent with a model in which the cyclic nucleotide mediates the effects of NO. Additional studies showed that wortmannin and S-NO-glutathione have additive inhibitory effects on thrombin receptor-activating peptide-induced platelet aggregation and the surface expression of platelet activation markers. These data provide evidence of a distinct and novel mechanism for the inhibitory effects of NO on platelet function.

Incubation of rat liver particulate fraction with insulin causes the release of a substance that stimulates lipid synthesis and down regulates the insulin receptor in primary cultures of isolated rat hepatocytes. This substance may be similar to putative mediator(s) of insulin action which has been shown to modulate the activity of key enzymes of lipid and carbohydrate metabolism in various cell free systems. Our data demonstrate that the mediator of insulin is also biologically active in an intact cell system. Down regulation of the insulin receptor by the mediator supports the concept that this phenomenon is a post binding event of insulin action.

In vivo vanadate and vanadyl have been shown to mimic the action of insulin and to be effective treatment for animal models of both Type I and Type II diabetes. The molecular mechanism of action of the vanadium salts on insulin sensitivity remains uncertain, and several potential sites proposed for the insulin-like effects are reviewed. In human trials, insulin sensitivity improved in patients with NIDDM, as well as in some patients with IDDM after two weeks of treatment with sodium metavanadate. This increase in insulin sensitivity was primarily due to an increase in non-oxidative glucose disposal, whereas oxidative glucose disposal and both basal and insulin stimulated suppression of hepatic glucose output (HGP) were unchanged. Clinically, oral vanadate was associated with a small decrease in insulin requirements in IDDM subjects. Of additional benefit, there was a decrease in total cholesterol levels in both IDDM and NIDDM subjects. Furthermore, there was an increase in the basal activities of MAP and S6 kinases to levels similar to the insulin-stimulated levels in controls, but there was little or no further stimulation with insulin was seen. Further understanding of the mechanism of vanadium action may ultimately be useful in the design of drugs that improve glucose tolerance.

Recent evidence suggests that insulin signaling through the insulin receptor A type (Ex11−), regulates insulin gene transcription. Because chronic hyperglycemia negatively affects insulin receptor function and regulates alternative splicing of the insulin receptor, we inquired whether chronic exposure of pancreatic β‐cells to high glucose results in alterations in insulin signaling due to changes in insulin receptor expression and relative abundance of its spliced isoforms. Our results demonstrate that the insulin receptor is localized in insulin secretory vescicles in human pancreatic β‐cells. Furthermore, we find that alterations in insulin expression and secretion caused by chronic exposure to high glucose are paralleled by decreased insulin receptor expression and increased relative abundance of the Ex11+ isoform in both human islets and RIN β‐cells. PDX‐1 and HMGI(Y) transcription factors are down‐regulated by high glucose. These changes are associated with defects in insulin signaling involving insulin receptor‐associated PI 3‐kinase/Akt/PHAS‐I pathway in RIN β‐cells. Re‐expression in RIN β‐cells chronically exposed to high glucose of the Ex11−, but not the Ex11+, isoform restored insulin mRNA expression. These data suggest that changes in early steps of insulin receptor signaling may play a role in determining β‐cell dysfunction caused by chronic hyperglycemia.