Diurnal Plasma Cortisol Measurements Utility in Differentiating Various Etiologies of Endogenous Cushing Syndrome.

Cortisol diurnal variation may be abnormal among patients with endogenous Cushing syndrome (CS). The study objective was to compare the plasma cortisol AM/PM ratios between different etiologies of CS. This is a retrospective cohort study, conducted at a clinical research center. Adult patients with CS that underwent adrenalectomy or trans-sphenoidal surgery (n=105) were divided to those with a pathologically confirmed diagnosis of Cushing disease (n=21) and those with primary adrenal CS, including unilateral adrenal adenoma (n=28), adrenocortical hyperplasia (n=45), and primary pigmented nodular adrenocortical disease (PPNAD, n=11). Diurnal plasma cortisol measurements were obtained at 11:30 PM and midnight and at 7:30 and 8:00 AM. The ratios between the mean morning levels and mean late-night levels were calculated. Mean plasma cortisol AM/PM ratio was lower among CD patients compared to those with primary adrenal CS (1.4±0.6 vs. 2.3±1.5, p<0.001, respectively). An AM/PM cortisol ratio≥2.0 among patients with unsuppressed ACTH (>15 pg/ml) excludes CD with a 85.0% specificity and a negative predictive value (NPV) of 90.9%. Among patients with primary adrenal CS, an AM/PM cortisol≥1.2 had specificity and NPV of 100% for ruling out a diagnosis of PPNAD. Plasma cortisol AM/PM ratios are lower among patients with CD compared with primary adrenal CS, and may aid in the differential diagnosis of endogenous hypercortisolemia.

Thumb carpometacarpal joint soft-tissue lesions in an asymptomatic population. An MRI study.

The purpose of this study was to evaluate the prevalence of ligament pathology around the first carpometacarpal joint in an asymptomatic population. We used a compact office-based MRI system and examined the hands of 117 healthy volunteers. We checked the competence of the anterior oblique ligament and of the posterior ligament complex on the MRI images. Our results showed that, in 82% of the study population, both ligaments could be perfectly visualized, with consistent signal from origin to insertion. Examination found signal changes indicating a damaged or torn ligament in the other cases. This study suggests that ligament pathology seen on MRI should be interpreted with caution. In symptomatic patients, pathologic images are to be interpreted in the light of the relevant clinical context. LEVEL OF EVIDENCE: III.

Phenotypic and genetic variation in leptin as determinants of weight regain.

AIMS: Over 75% of obese subjects fail to maintain their weight following weight loss interventions. We aimed to identify phenotypic and genetic markers associated with weight maintenance/regain following a dietary intervention. SUBJECTS AND METHODS: In the 2-year Dietary Intervention Randomized Controlled Trial, we assessed potential predictors for weight changes during the 'weight loss phase' (0-6 months) and the 'weight maintenance/regain phase' (7-24 months). Genetic variation between study participants was studied using single-nucleotide polymorphisms in the leptin gene (LEP). RESULTS: Mean weight reduction was -5.5% after 6 months, with a mean weight regain of 1.2% of baseline weight during the subsequent 7-24 months. In a multivariate regression model, higher baseline high-molecular-weight adiponectin was the only biomarker predictor of greater success in 0- to 6-month weight loss (ß = -0.222, P-value = 0.044). In a multivariate regression model adjusted for 6-month changes in weight and various biomarkers, 6-month plasma leptin reduction exhibited the strongest positive association with 6-month weight loss (ß = 0.505, P-value < 0.001). Conversely, 6-month plasma leptin reduction independently predicted weight regain during the following 18 months (ß = -0.131, P-value < 0.013). Weight regain was higher among participants who had a greater (top tertiles) 6-month decrease in both weight and leptin (+3.4% (95% confidence interval 2.1-4.8)) as compared with those in the lowest combined tertiles (+0.2% (95% confidence interval -1.1 to 1.4)); P-value < 0.001. Weight regain was further significantly and independently associated with genetic variations in LEP (P = 0.006 for both rs4731426 and rs2071045). Adding genetic data to the phenotypic multivariate model increased its predictive value for weight regain by 34%. CONCLUSION: Although greater reduction in leptin concentrations during the initial phase of a dietary intervention is associated with greater weight loss in the short term, plasma leptin reduction, combined with the degree of initial weight loss and with genetic variations in the LEP gene, constitutes a significant predictor of subsequent long-term weight regain.

HEREDITARY ENDOCRINE TUMOURS: CURRENT STATE-OF-THE-ART AND RESEARCH OPPORTUNITIES: MEN1-related pancreatic NETs: identification of unmet clinical needs and future directives.

The PanNET Working Group of the 16th International Multiple Endocrine Neoplasia Workshop (MEN2019) convened in Houston, TX, USA, 27-29 March 2019 to discuss key unmet clinical needs related to PanNET in the context of MEN1, with a special focus on non-functioning (nf)-PanNETs. The participants represented a broad range of medical scientists as well as representatives from patient organizations, pharmaceutical industry and research societies. In a case-based approach, participants addressed early detection, surveillance, prognostic factors and management of localized and advanced disease. For each topic, after a review of current evidence, key unmet clinical needs and future research directives to make meaningful progress for MEN1 patients with nf-PanNETs were identified. International multi-institutional collaboration is needed for adequately sized studies and validation of findings in independent datasets. Collaboration between basic, translational and clinical scientists is paramount to establishing a translational science approach. In addition, bringing clinicians, scientists and patients together improves the prioritization of research goals, assures a patient-centered approach and maximizes patient involvement. It was concluded that collaboration, research infrastructure, methodologic and reporting rigor are essential to any translational science effort. The highest priority for nf-PanNETs in MEN1 syndrome are (1) the development of a data and biospecimen collection architecture that is uniform across all MEN1 centers, (2) unified strategies for diagnosis and follow-up of incident and prevalent nf-PanNETs, (3) non-invasive detection of individual nf-PanNETs that have an increased risk of metastasis, (4) chemoprevention clinical trials driven by basic research studies and (5) therapeutic targets for advanced disease based on biologically plausible mechanisms.

Emergency room visit: a red-flag indicator for poor diabetes care.

AIM: To determine the association between emergency room (ER) admission and quality of diabetes care in the community. METHODS: In a nested case-control study of patients with Type 2 diabetes mellitus (DM) within a large health maintenance organization (HMO) in Israel, 919 patients who were admitted to one of West Jerusalem's ERs between 1 May and 30 June 2004 were compared with 1952 control subjects not admitted. Data on study covariates were retrieved from the HMO's computerized database and a subset of the study population was interviewed. Logistic regressions were conducted to estimate the odds ratios of being admitted according to different measures of quality of care, controlling for socio-demographic variables, co-morbidities and type of DM treatment. RESULTS: The main indices of quality of primary care that were inversely associated with visiting an ER during the study period included performance of a cholesterol test in the year prior to the index date [adjusted odds ratio (OR) 0.23, 95% confidence interval (CI) 0.19-0.29, P < 0.001], performance of glycated haemoglobin test (OR 0.26, 95% CI 0.24-0.29, P < 0.001), visiting an ophthalmologist (OR 0.47, 95% CI 0.32-0.68, P = 0.001), and recommendations to stop smoking (OR 0.10, 95% CI 0.05-0.21, P < 0.001). CONCLUSIONS: Admission to the ER can be used as an indicator for poor quality of diabetes care. There is an association between ER admission and poor quality of diabetes care.

Prevalence of asthma in a large group of Israeli adolescents: influence of country of birth and age at migration.

BACKGROUND: The occurrence of asthma has geographic variations and is lower in developing compared with industrialized countries. Both environmental and genetic factors may influence its prevalence. We aimed to evaluate the importance and effect of immigration (country of birth and age at immigration to Israel) on the prevalence of asthma in a large group of Israeli adolescents. METHODS: Computerized medical records of 17-year-old adolescents, who underwent routine examination before military recruitment, were studied. The sample comprised both native-born Israelis (NBI) and immigrants from Ethiopia, the Former Soviet Union (FSU), and Western countries (WC). Asthma was defined as clinical symptoms and signs compatible with the disease accompanied by abnormal spirometry or documented chronic use of inhaled steroids. RESULTS: Our cohort consisted of 1 466 654 adolescents, including 1 317 556 (89.8%) NBI and 149 098 (10.2%) immigrants. The prevalence of asthma at age 17 was higher in NBI compared with Ethiopian immigrants [4.7% (61 921) vs 2.6% (418), respectively, P < 0.0005], lower compared with immigrants from WC [5.6% (2177), P < 0.0005], and similar to immigrants from the FSU. Further analysis of the association between age at immigration and the risk for developing asthma showed that the younger immigrants from the FSU and Ethiopia arrived to Israel, the higher their prevalence of asthma at the age of 17 was. CONCLUSIONS: Both environmental and genetic factors seem to influence the prevalence of asthma in 17-year-old adolescents. However, the higher risk for developing asthma associated with young age of immigration points toward an environmental predominance.

Activation of mitogen-activated protein kinases in human heart during cardiopulmonary bypass.

Mitogen-activated protein kinases (MAPKs) have been shown to be activated in both in vitro and in vivo models of cardiac tissue in response to ischemia/reperfusion injury. We investigated whether MAPKs are activated in human heart during coronary artery bypass grafting (CABG) surgery. During elective CABG surgery of 8 patients, 3 right atrial appendage biopsies were obtained at baseline, at the end of cross-clamping, and after coronary reperfusion. The expression of the p38-MAPK, c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinases (ERK1/2) MAPKs was not altered during CABG. The phosphorylation and activation of both ERK1/2 and p38-MAPK were increased approximately 2-fold by ischemia and even more (8- and 4-fold, respectively) by reperfusion. Although the ischemic period did not result in a significant activation of JNK, an approximately 6-fold increase in JNK activity could be observed after reperfusion. In conclusion, distinct activation patterns of ERK1/2, p38, and JNK MAPKs can be observed in human heart during CABG.

Prolonged oxidative stress impairs insulin-induced GLUT4 translocation in 3T3-L1 adipocytes.

Prolonged exposure of 3T3-L1 adipocytes to micromolar concentrations of H2O2 results in an impaired response to the acute metabolic effects of insulin. In this study, we further characterized the mechanisms by which oxidative stress impairs insulin stimulation of glucose transport activity. Although insulin induced a 2.5-fold increase in plasma membrane GLUT4 content and a 50% reduction in its abundance in the low-density microsomal (LDM) fraction in control cells, oxidation completely prevented these responses. The net effect of insulin on 2-deoxyglucose uptake activity was reduced in oxidized cells and could be attributed to GLUT1 translocation. Insulin stimulation of insulin receptor substrate (IRS) 1 tyrosine phosphorylation and the association of IRS-1 with phosphatidylinositol (PI) 3-kinase were not impaired by oxidative stress. However, a 1.9-fold increase in the LDM content of the p85 subunit of PI 3-kinase after insulin stimulation was observed in control, but not in oxidized, cells. Moreover, although insulin induced an increase in IRS-1-associated PI 3-kinase activity in the LDM in control cells, this effect was prevented by oxidation. These findings suggest that prolonged low-grade oxidative stress impairs insulin-stimulated GLUT4 translocation, potentially by interfering with compartment-specific activation of PI 3-kinase.

The HIV protease inhibitor nelfinavir induces insulin resistance and increases basal lipolysis in 3T3-L1 adipocytes.

HIV protease inhibitors (HPIs) are potent antiretroviral agents clinically used in the management of HIV infection. Recently, HPI therapy has been linked to the development of a metabolic syndrome in which adipocyte insulin resistance appears to play a major role. In this study, we assessed the effect of nelfinavir on glucose uptake and lipolysis in differentiated 3T3-L1 adipocytes. An 18-h exposure to nelfinavir resulted in an impaired insulin-stimulated glucose uptake and activation of basal lipolysis. Impaired insulin stimulation of glucose up take occurred at nelfinavir concentrations >10 micromol/l (EC(50) = 20 micromol/l) and could be attributed to impaired GLUT4 translocation. Basal glycerol and free fatty acid (FFA) release were significantly enhanced with as low as 5 micromol/l nelfinavir, displaying fivefold stimulation of FFA release at 10 micromol/l. Yet, the antilipolytic action of insulin was preserved at this concentration. Potential underlying mechanisms for these metabolic effects included both impaired insulin stimulation of protein kinase B Ser 473 phosphorylation with preserved insulin receptor substrate tyrosine phosphorylation and decreased expression of the lipolysis regulator perilipin. Troglitazone pre- and cotreatment with nelfinavir partly protected the cells from the increase in basal lipolysis, but it had no effect on the impairment in insulin-stimulated glucose uptake induced by this HPI. This study demonstrates that nelfinavir induces insulin resistance and activates basal lipolysis in differentiated 3T3-L1 adipocytes, providing potential cellular mechanisms that may contribute to altered adipocyte metabolism in treated HIV patients.

Lipoic acid reduces glycemia and increases muscle GLUT4 content in streptozotocin-diabetic rats.

Alpha lipoic acid (lipoate [LA]), a cofactor of alpha-ketodehydrogenase, exhibits unique antioxidant properties. Recent studies suggest a direct effect of LA on glucose metabolism in both human and experimental diabetes. This study examines the possibility that LA positively affects glucose homeostasis in streptozotocin (STZ)-induced diabetic rats by altering skeletal muscle glucose utilization. Blood glucose concentration in STZ-diabetic rats following 10 days of intraperitoneal (i.p.) injection of LA 30 mg/kg was reduced compared with that in vehicle-treated diabetic rats (495 +/- 131 v 641 +/- 125 mg/dL in fed state, P = .003, and 189 +/- 48 v 341 +/- 36 mg/dL after 12-hour fast, P = .001). No effect of LA on plasma insulin was observed. Gastrocnemius muscle crude membrane GLUT4 protein was elevated both in control and in diabetic rats treated with LA by 1.5- and 2.8-fold, respectively, without significant changes in GLUT4 mRNA levels. Gastrocnemius lactic acid was increased in diabetic rats (19.9 +/- 5.5 v 10.4 +/- 2.8 mumol/g muscle, P < .05 v nondiabetic rats), and was normal in LA-treated diabetic rats (9.1 +/- 5.0 mumol/g muscle). Insulin-stimulated 2-deoxyglucose (2 DG) uptake into isolated soleus muscle was reduced in diabetic rats compared with the control group (474 +/- 15 v 568 +/- 52 pmol/mg muscle 30 min, respectively, P = .05). LA treatment prevented this reduction, resulting in insulin-stimulated glucose uptake comparable to that of nondiabetic animals. These results suggest that daily LA treatment may reduce blood glucose concentrations in STZ-diabetic rats by enhancing muscle GLUT4 protein content and by increasing muscle glucose utilization.

Regulation of glucose transporters--implications for insulin resistance states.

Altered glucose homeostasis in the different diabetic states often results from a combination of insulin deficiency (absolute or relative), and impaired hormone action. The latter involves alterations in the expression and/or function of glucose transporters in insulin responsive peripheral tissues - skeletal muscle and adipose tissue. Since whole body glucose utilization depends mainly on controlled changes in glucose transport in these tissues, this review focuses on the role of glucose transporters in the regulation of insulin-stimulated glucose transport activity. The molecular mechanisms by which several inducers of insulin resistance inhibit insulin action on glucose uptake are also discussed. Better understanding of the complex regulation of glucose transport and transporters will hopefully shed light on potential sites for new pharmaceutical interventions. Several excellent reviews have been published in the past 2 years detailing various aspects which are discussed only briefly in this review. They are mentioned in the text to allow further reading.

Peripheral blood leucocyte subclasses as potential biomarkers of adipose tissue inflammation and obesity subphenotypes in humans.

While obesity is clearly accepted as a major risk factor for cardio-metabolic morbidity, it is also apparent that some obese patients largely escape this association, forming a unique obese subphenotype(s). Current approaches to define such subphenotypes include clinical biomarkers that largely reflect already manifested comorbidities, such as markers of dyslipidaemia, hyperglycaemia and impaired regulation of vascular tone, and anthropometric or imaging-based assessment of adipose tissue distribution. Low-grade inflammation, evident both systemically and within adipose tissue (particularly intra-abdominal fat depots), seems to characterize the more cardio-metabolically morbid forms of obesity. Indeed, several systemic inflammatory markers (C-reactive protein), adipokines (retinol-binding protein 4, adiponectin) and cytokines have been shown to correlate in humans with adipose tissue inflammation and with obesity-associated health risks. Circulating leucocytes constitute a diverse group of cells that form a major arm of the immune system. They are both major sources of cytokines and likely also of infiltrating adipose tissue immune cells in obesity. In the present review, we summarize currently available literature on 'classical' blood white cell classes and on more specific leucocyte subclasses present in the circulation in human obesity. We critically raise the possibility that leucocytes may constitute clinically available markers for the more morbidity-associated obesity subphenotype(s), and when available, for intra-abdominal adipose tissue inflammation.

25 hydroxyvitamin D levels in patients undergoing coronary artery catheterization.

INTRODUCTION: A growing body of evidence suggests that vitamin D deficiency is associated with increased cardiovascular morbidity and mortality. The present study assessed the association between low serum 25-hydroxyvitamin D (25(OH)D) and coronary artery disease status defined by coronary catheterization findings. METHODS: An observational study of 101 consecutive patients admitted to Assaf Harofeh Medical Center during 2009, and scheduled to undergo coronary catheterization was undertaken. Blood was collected for parathyroid hormone, 25(OH)D and high sensitivity C reactive protein (hsCRP). 25(OH)D deficiency was defined as <20 ng/ml. Patients were divided into two groups: patients with normal or non-significant coronary artery disease and patients with a significant coronary artery disease as found during cardiac catheterization. Logistic regression model was used to compare pathological coronary catheterization findings, including 25(OH)D levels dichotomized to low (serum 25(OH)D levels<20 ng/ml) vs. high (serum 25(OH)D levels ≥ 20 ng/ml) and other confounders. RESULTS: Patients with pathological coronary catheterization had 25(OH)D deficiency (75% vs 55.1%, p=0.036). Pathological coronary catheterization was more prevalent among patients with 25(OH)D deficiency (Odds ratio (OR) 2.44, 95% confidence interval (CI) 1.05-5.68, p=0.038). This difference was more pronounced after controlling for sex, age, BMI, ethnicity and present smoking (OR 2.92, 95% CI 1.01-8.46, p=0.016). CONCLUSIONS: 25(OH)D deficiency is significantly associated with pathological cardiac catheterization findings. This association is strengthened further by controlling for other cardiovascular disease risk factors.

Oxidative stress impairs insulin but not platelet-derived growth factor signalling in 3T3-L1 adipocytes.

Activation of phosphatidylinositol 3-kinase (PI 3-kinase) is a common event in both insulin and platelet-derived growth factor (PDGF) signalling, but only insulin activates this enzyme in the high-speed pellet (HSP), and induces GLUT4 translocation. Recently, we have demonstrated that exposure of 3T3-L1 adipocytes to oxidative stress impairs insulin-stimulated GLUT4 translocation and glucose transport, associated with impaired PI 3-kinase translocation and activation in the HSP [Tirosh, Potashnik, Bashan and Rudich (1999) J. Biol. Chem. 274, 10595-10602]. In this study the effect of a 2 h exposure to approximately 30 microM H(2)O(2) on insulin versus PDGF-BB signalling and metabolic effects was compared. PDGF-stimulated p85-associated PI 3-kinase activity in total cell lysates, as well as co-precipitation of the PDGF receptor, were unaffected by oxidative stress. Additionally, the increase in p85 association with the plasma-membrane lawns by PDGF remained intact following oxidation, whereas the insulin effect was decreased. PDGF significantly increased protein kinase B (PKB) activity in early differentiated cells, and that of p70 S6-kinase in both early and fully differentiated 3T3-L1 adipocytes. Following oxidation the effect of PDGF on PKB and p70 S6-kinase activation remained intact, whereas significant inhibition of insulin-stimulated activation of those enzymes was observed. In accordance, in both early and fully differentiated cells, oxidative stress completely blunted insulin- but not PDGF-stimulated protein synthesis. In conclusion, oxidative stress impairs insulin, but not PDGF, signalling and metabolic actions in both early and fully differentiated 3T3-L1 adipocytes. This emphasizes compartment-specific activation of PI 3-kinase as an oxidation-sensitive step specifically leading to insulin resistance.

Oxidative stress disrupts insulin-induced cellular redistribution of insulin receptor substrate-1 and phosphatidylinositol 3-kinase in 3T3-L1 adipocytes. A putative cellular mechanism for impaired protein kinase B activation and GLUT4 translocation.

In a recent study we have demonstrated that 3T3-L1 adipocytes exposed to low micromolar H2O2 concentrations display impaired insulin stimulated GLUT4 translocation from internal membrane pools to the plasma membrane (Rudich, A., Tirosh, A., Potashnik, R., Hemi, R., Kannety, H., and Bashan, N. (1998) Diabetes 47, 1562-1569). In this study we further characterize the cellular mechanisms responsible for this observation. Two-hour exposure to approximately 25 microM H2O2 (generated by adding glucose oxidase to the medium) resulted in disruption of the normal insulin stimulated insulin receptor substrate (IRS)-1 and phosphatidylinositol (PI) 3-kinase cellular redistribution between the cytosol and an internal membrane pool (low density microsomal fraction (LDM)). This was associated with reduced insulin-stimulated IRS-1 and p85-associated PI 3-kinase activities in the LDM (84 and 96% inhibition, respectively). The effect of this finding on the downstream insulin signal was demonstrated by a 90% reduction in insulin stimulated protein kinase B (PKB) serine 473 phosphorylation and impaired activation of PKBalpha and PKBgamma. Both control and oxidized cells exposed to heat shock displayed a wortmannin insensitive PKB serine phosphorylation and activity. These data suggest that activation of PKB and GLUT4 translocation are insulin signaling events dependent upon a normal insulin induced cellular compartmentalization of PI 3-kinase and IRS-1, which is oxidative stress-sensitive. These findings represent a novel cellular mechanism for the induction of insulin resistance in response to changes in the extracellular environment.

Are chronic digestive complaints the result of abnormal dietary patterns? Diet and digestive complaints in children at 22 and 40 months of age.

We studied 149 healthy children at 22 months of age and 74 at 40 months of age, employing a 24-hour dietary record based on premailed food measurement guide and telephone questionnaire. Parents cooperated in 94% of contacts. Chronic digestive complaints decreased from 27% to 5% of the sample over the study period: constipation from 16% to 3%, chronic diarrhea from 8% to 1%, and abdominal pain from 5% to 1%. Excessive fluid intake (1470 +/- 600 vs 840 +/- 300 mL/d) correlated most strongly for seven children at 22 months experiencing alternating symptoms of chronic diarrhea and constipation or abdominal pain. Many other children tolerated dietary extremes without complaint. All macronutrient categories except dietary fiber intake increased over the study period. Thus, excessive fluid intake may provoke symptoms suggesting the irritable bowel syndrome in a susceptible group of younger children. Failure to increase fiber intake from 22 to 40 months of age leaves children on an immature diet whose effects require further study.

Lipoic acid protects against oxidative stress induced impairment in insulin stimulation of protein kinase B and glucose transport in 3T3-L1 adipocytes.

AIMS/HYPOTHESIS: Oxidative stress has been shown to impair insulin-stimulated glucose transporter 4 translocation in 3T3-L1 adipocytes. This study explores the potential of the antioxidant lipoic acid to protect the cells against the induction of insulin resistance when given before exposure to oxidative stress. METHODS: 3T3-L1 were exposed for 16 h to lipoic acid after which cells were exposed for 2 h to continuous production of H2O2 by adding glucose oxidase to the culture medium. RESULTS: These conditions resulted in a 50-70% reduction in insulin-stimulated glucose transport activity associated with a decrease in reduced glutathione content from 37.4 +/- 3.1 to 26.4 +/- 4.9 nmol/mg protein, (p < 0.005). Lipoic acid pretreatment increased insulin-stimulated glucose transport following oxidative stress, reaching 84.8 +/- 4.4% of the control, associated with an increase in reduced glutathione content. Oxidation impaired the 4.89 +/- 0.36-fold insulin-stimulated increase in glucose transporter 4 content in plasma membrane lawns of control cells. Lipoic acid pretreatment was, however, associated with preserved insulin-induced glucose transporter 4 translocation in cells exposed to oxidation, yielding 80% of its content in controls. Although tyrosine phosphorylation patterns were not affected by lipoic acid pretreatment, insulin-stimulated protein kinase B/Akt serine 473 phosphorylation and activity were considerably impaired by oxidation but protected by lipoic acid pretreatment. A protective effect was not observed with either troglitazone, its isolated vitamin E moiety, or with vitamin C. CONCLUSION/INTERPRETATION: This study shows the ability of lipoic acid to provide partial protection against the impaired insulin-stimulated glucose transporter 4 translocation and protein kinase B/Akt activation induced by oxidative stress, potentially by its capacity to maintain intracellular redox state.

Nelfinavir-induced insulin resistance is associated with impaired plasma membrane recruitment of the PI 3-kinase effectors Akt/PKB and PKC-zeta.

AIMS/HYPOTHESIS: Chronic exposure of 3T3-L1 adipocytes to the HIV protease inhibitor nelfinavir induces insulin resistance, recapitulating key metabolic alterations of adipose tissue in the lipodystrophy syndrome induced by these agents. Our goal was to identify the defect in the insulin signal transduction cascade leading to nelfinavir-induced insulin resistance. METHODS: Fully differentiated 3T3-L1 adipocytes were exposed to 30 micro mol/l nelfinavir for 18 h, after which the amount, the phosphorylation and the localisation of key proteins in the insulin signalling cascade were evaluated. RESULTS: Insulin-induced interaction of phosphatidylinositol 3'-kinase (PI 3-kinase) with IRS proteins was normal in cells treated with nelfinavir, as was IRS-1-associated PI 3-kinase activity. Yet insulin-induced phosphorylation of Akt/protein kinase B (PKB), p70S6 kinase and extracellular signal-regulated kinase 1/2 was significantly impaired. This could not be attributed to increased protein phosphatase 2A activity or to increased expression of phosphoinositide phosphatases (SHIP2 or PTEN). However, insulin failed to induce translocation of the PI 3-kinase effectors Akt/PKB and protein kinase C-zeta (PKC-zeta) to plasma membrane fractions of nelfinavir-treated adipocytes. CONCLUSIONS/INTERPRETATION: We therefore conclude that nelfinavir induces a defect in the insulin signalling cascade downstream of the activation of PI 3-kinase. This defect manifests itself by impaired insulin-mediated recruitment of Akt/PKB and PKC-zeta to the plasma membrane.