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Objectives: To assess associations between the activity of hypothalamo-pituitary-adrenal (HPA) axis and liver histology in patients with nonalcoholic fatty liver disease (NAFLD)., Design and Patients: In a cross-sectional study, we enrolled 50 consecutive, overweight, NAFLD patients and 40 control subjects who were comparable for age, sex and body mass index (BMI)., Measurements: NAFLD (by liver biopsy), HPA axis activity (by 24-hour urinary free cortisol [UFC] excretion and serum cortisol levels after 1 mg dexamethasone), insulin resistance (by homeostasis model assessment: HOMA-IR), and metabolic syndrome (MetS) features., Results: NAFLD patients had markedly higher (P < 0.001) 24-h UFC (149 +/- 24 vs. 90 +/- 16 nmol/day) and postdex suppression cortisol concentrations (32 +/- 10 vs. 16 +/- 7 nmol/l) than controls. The MetS and its individual components were more frequent among NAFLD patients. The marked differences in urinary/serum cortisol concentrations that were observed between the groups were little affected by adjustment for age, sex, BMI, waist circumference, systolic blood pressure, triglycerides, homeostasis model assessment for insulin resistance score and presence of diabetes. Importantly, 24-h UFC and postdex cortisol concentrations strongly correlated to hepatic necroinflammatory grade (P < 0.01) and fibrosis stage (P < 0.001) among NAFLD patients. By logistic regression analysis, 24-h UFC (odds ratio (OR) 1.80, 95%CI 1.3-2.8) or postdex cortisol concentrations (OR 1.95, 95%CI 1.4-3.1) independently predicted the severity of hepatic fibrosis, but not necroinflammation, after adjustment for potential confounders., Conclusions: These results suggest that NAFLD patients have a subtle, chronic overactivity in the HPA axis (that is closely associated with the severity of liver histopathology) leading to subclinical hypercortisolism that might be implicated in the development of NAFLD.

Non-alcoholic fatty liver disease (NAFLD) is closely associated with several metabolic syndrome (MetS) features. We assessed whether NAFLD is significantly associated with carotid artery intima-media thickness (IMT), as a marker of subclinical atherosclerosis, and whether such association is independent of classical cardiovascular risk factors and MetS features. We studied 100 diet-controlled Type 2 diabetic patients with ultrasonographically diagnosed NAFLD and 100 diabetic patients without NAFLD who were comparable for age and sex. Main outcome measures were carotid IMT (by ultrasonography), classical risk factors, insulin resistance [as estimated by homeostasis model assessment (HOMA)-IR] and MetS (as defined by the Adult Treatment Panel III criteria). NAFLD patients had a markedly greater carotid IMT (1.24 +/- 0.13 vs 0.95 +/- 0.11 mm; p < 0.001) than those without the condition. The MetS and all its clinical traits were more highly prevalent in those with NAFLD (p < 0.001). Adjustment for age, sex, smoking history, diabetes duration, glycosylated hemoglobin, LDL cholesterol, liver enzymes and microalbuminuria did not really affect the significant differences in carotid IMT that were observed between the groups. Further adjustment for the MetS also had little impact, but additional adjustment for HOMA-IR score consistently attenuated any statistical significance (p = 0.28). In multivariate regression analysis, HOMA-IR score along with age and MetS (principally raised blood pressure values) were independently related to carotid IMT, whereas NAFLD was not. In conclusion, these results suggest that among diet-controlled Type 2 diabetic individuals the significant increase of carotid IMT in the presence of NAFLD is largely explained by HOMA-estimated insulin resistance.

Aims: To compare plasma biomarkers of inflammation and endothelial dysfunction in individuals with and without non-alcoholic hepatic steatosis (HS), and to evaluate whether such differences were mediated by the adverse metabolic pattern, typically found in these subjects., Methods: HS (by ultrasound and computed tomography), visceral fat (by computed tomography), insulin resistance (by homeostasis model assessment-HOMA), plasma biomarkers of inflammation and endothelial dysfunction (hs-C reactive protein, fibrinogen, von Willebrand factor, plasminogen activator inhibitor-1 activity) were measured in 100 non-smoking, healthy, male volunteers., Results: Plasma hs-CRP, fibrinogen, v-WF and plasminogen activator inhibitor-1 (PAI-1) activity levels were markedly higher (P < 0.01 or less) in subjects with non-alcoholic HS (n = 35) than in those without HS (n = 65). The former also had significantly higher values for body mass index (BMI), visceral fat, diastolic blood pressure, HOMA insulin resistance score, plasma insulin (both fasting and after glucose load), triglycerides, liver enzymes, and lower high-density lipoprotein (HDL)-cholesterol concentration. While the marked differences in these pro-inflammatory biomarkers observed between the groups were little affected by adjustment for age, BMI, blood pressure values, HOMA insulin resistance score, plasma triglyceride and liver enzyme concentrations, they were completely abolished after controlling for visceral fat. Similarly, in multivariate regression analyses, increased visceral fat significantly predicted the pro-inflammatory biomarkers, independently of HS and other potential confounders., Conclusions: These results indicate that, in non-smoking, non-diabetic men, the significant increase of plasma biomarkers of inflammation and endothelial dysfunction in the presence of non-alcoholic HS is largely mediated by abdominal visceral fat accumulation.

Aims: To examine the association of non-alcoholic hepatic steatosis (HS) with the activity of the hypothalamo-pituitary-adrenal (HPA) axis in Type 2 diabetic individuals., Methods: The activity of the HPA axis, as measured by 24-h urinary free cortisol (UFC) excretion and serum cortisol levels after 1.0 mg dexamethasone, was measured in 40 diet-controlled, predominantly overweight, Type 2 diabetic patients with non-alcoholic HS and in 40 diabetic patients without HS who were comparable for age, sex and body mass index (BMI)., Results: Subjects with non-alcoholic HS had significantly higher 24-h UFC excretion (191 +/- 4 vs. 102 +/- 3 nmol/24 h; P < 0.001) and post-dexamethasone cortisol concentrations (29.1 +/- 2 vs. 14.4 +/- 1 nmol/l; P < 0.001) than those without HS. Patients with HS had significantly higher values for HOMA insulin resistance score, plasma triglycerides and liver enzymes. Age, sex, BMI, waist-hip ratio (WHR), diabetes duration, HbA1c, LDL-cholesterol and blood pressure values were not different between the groups. The differences in urinary and serum cortisol concentrations between the groups remained significant after adjustment for age, sex, BMI, WHR, HOMA insulin resistance score, plasma triglycerides, HbA1c and liver enzymes. In multiple logistic regression analyses, 24-h UFC or serum cortisol concentrations (P < 0.05 and P = 0.02, respectively), along with age and HOMA insulin resistance, predicted the presence of HS, independently of potential confounders., Conclusions: These results demonstrate that non-alcoholic HS is closely associated with a subtle, chronic overactivity of the HPA axis in diet-controlled Type 2 diabetic individuals.

Aims: To evaluate whether plasma biomarkers of inflammation and endothelial dysfunction differed in Type 1 diabetic patients as compared with those in non-diabetic subjects, and to examine the association of these biomarkers with early stages of microvascular complications., Methods: Plasma biomarkers of inflammation [fibrinogen, hs-C-reactive protein (hs-CRP)] and endothelial dysfunction [von Willebrand factor (v-WF), intercellular adhesion molecule-1, plasminogen activator inhibitor-1 (PAI-1) activity] were measured in 88 non-smoking young patients with Type 1 diabetes without clinical macrovascular disease and in 40 healthy controls., Results: Plasma levels of hs-CRP, fibrinogen, v-WF, soluble intracellular adhesion molecule-1 (sICAM-1) and PAI-1 activity were markedly higher (P < 0.01 or less) in Type 1 diabetic patients than in healthy controls; these results were essentially unchanged when healthy controls were compared with patients without complications. After stratification by microvascular complication status, plasma biomarkers of inflammation and endothelial dysfunction were significantly increased in those with more advanced disease compared with those with early complications or without complications, respectively. However, while the significant differences in these biomarkers were little affected by adjustment for sex, age, BMI and blood pressure values, they were totally abolished after additional adjustment for diabetes duration and glycaemic control., Conclusions: These results indicate that in Type 1 diabetes there is a subclinical, chronic inflammation which is, at least partly, independent of clinically manifest macro- and microvascular complications, smoking or other traditional cardiovascular risk factors; this subclinical inflammation is closely correlated to the magnitude and duration of hyperglycaemia.

Objectives: To examine the association between nonalcoholic hepatic steatosis (HS) and the activity of the hypothalamo-pituitary-adrenal (HPA) axis, as evaluated by a low-dose dexamethasone suppression test, in obese subjects., Design and Patients: In a cross-sectional study, we examined 54 obese, otherwise healthy, individuals with a negligible or zero daily alcohol consumption., Measurements: HS (by ultrasonography), glucose tolerance status (by oral glucose load), insulin resistance [by homeostasis model assessment (HOMA)] and 1.0 mg postdexamethasone (postdex) suppression cortisol levels were measured., Results: Subjects with nonalcoholic HS (n = 39) had markedly less suppressed circulating cortisol levels than those without HS (n = 15) (21.9 +/- 2.6 vs. 11.0 +/- 1.4 nmol/l, P < 0.001). In addition, subjects with nonalcoholic HS had significantly higher values of HOMA-insulin resistance score and circulating liver enzymes than their counterparts without HS. Age, body mass index (BMI), waist/hip ratio, plasma glucose concentration (both at fasting and after glucose load), lipids and blood pressure values did not differ significantly between the groups. Females were more represented among those without HS. The marked differences in postdex suppression cortisol levels that were observed between the groups were little affected by adjustment for sex, age, BMI, waist/hip ratio, HOMA-insulin resistance score, plasma lipids and liver enzyme levels. Similarly, in a logistic regression analysis, cortisol levels significantly predicted the presence of nonalcoholic HS (P = 0.032), independently of potential confounders., Conclusions: These results suggest that nonalcoholic hepatic steatosis is closely correlated with a subtle, chronic activation of the HPA axis in obese, otherwise healthy, individuals.

Background and Aim: Hepatic steatosis has recently been associated with insulin resistance and other metabolic abnormalities as a possible feature of the metabolic syndrome, but it is still uncertain how hepatic steatosis and insulin sensitivity are connected. Furthermore, obesity is a well characterized insulin resistant condition that is often associated with hepatic steatosis. The aim of this study was to verify whether hepatic steatosis further worsens insulin sensitivity in obese subjects by comparing the degree of insulin sensitivity in obese subjects with normal glucose tolerance on the basis of the presence or absence of hepatic steatosis., Methods and Results: We analyzed 86 obese patients whose alcohol intake was less than 20 g/day and who showed no signs of viral hepatopathy. All of the subjects had normal glucose tolerance as shown by an oral glucose tolerance test. Insulin resistance was estimated using the homeostasis model assessment (HOMA) method and the diagnosis of steatosis was determined by an ultrasound scan of the liver. The subjects were comparable in terms of body mass index (BMI), lipid profile and serum uric acid levels; those with hepatic steatosis were slightly older and tended to have higher systolic blood pressure and fasting glycemia levels. The HOMA values were significantly higher in the group with hepatic steatosis (4.48 +/- 2.22 vs 3.11 +/- 1.47, p=0.002). There was no linear correlation between HOMA and alanine aminotransferase (ALT) levels, but a close linear correlation between HOMA and BMI (r=0.40; p<0.001). The effect of hepatic steatosis on HOMA remained significant after adjusting for age and gender (covariance analysis, p=0.006). When BMI was added to the covariance analysis, hepatic steatosis retained its statistical significance., Conclusion: Our data suggest that hepatic steatosis can increase insulin resistance independently of obesity.

Objective: The aim of the present work was to compare mortality from site-specific malignancies in type 2 diabetic patients with those in the general population., Research Design and Methods: Mortality from site-specific cancers was assessed in a population-based cohort of 7,148 type 2 diabetic patients from Verona (Northern Italy) during a 10-year follow-up (1987-1996) by reviewing death certificates. Standardized mortality ratio (SMR) data were computed using as reference mortality rates in the general population of Verona., Results: During follow-up, 641 patients (378 men and 263 women) died of malignancies. The most common causes of death among site-specific malignancies were digestive tumors both in men (140 of 378, 37.0%) and women (105 of 263, 39.9%), respiratory tumors in men (103 of 378, 27.2%), and tumors of the reproductive system in women (79 of 263, 30.0%). A slight increase in the overall mortality from malignancies was observed in diabetic patients and achieved statistical significance in women (observed/expected = 1.16, 95% CI 1.02-1.30; P = 0.019) but not in men (observed/expected = 1.07, 0.97-1.19; P = 0.163). Excess mortality from hepatic cancer (SMR = 1.86, 1.44-2.38) was observed in both men and women. In addition, women with diabetes experienced a higher mortality from pancreatic tumors (observed/expected = 1.78, 1.13-2.67) and breast tumors (observed/expected = 1.40, 1.06-1.81). Excess mortality from breast cancer was confined to obese women with diabetes., Conclusions: Mortality from site-specific malignancies is different in type 2 diabetic patients than in the general population. Better control of body weight seems necessary to prevent the excess mortality from breast cancer in women.

Objective: The relation between body weight and mortality in type II diabetic patients has not been fully elucidated. The aim of the present study was to evaluate the impact of body weight on mortality in a well-characterized type II diabetic cohort., Research Design and Methods: We examined a cohort of 3398 type II diabetic patients, alive on December 1986 and followed up for 10 years, to assess mortality and its causes and to investigate the relationship between body mass index (BMI) and mortality from all and specific causes. For this purpose, survival in the different quartiles of BMI was evaluated by a Cox model, controlling for sex, age, treatment, smoking, duration of diabetes, hypertension, and fasting plasma glucose. The Cox model was applied either excluding (model 1) or including (model 2) the last three variables., Results: During the 10 ys of follow-up, 1212 deaths (639 women, 573 men) occurred in the cohort under study. Since the interaction between BMI and age was statistically significant (P = 0.002), survival was studied separately in people aged <65 and > or =65 y (median age of the cohort = 65.9 y). Under 65 y, a significantly higher all-cause mortality was observed in obese patients, that is, in the IV quartile (BMI> or =30.9 kg/m(2); RR = 1.74; CI 95% = 1.26-2.40), in model 1. The inclusion of hypertension, duration of diabetes, and fasting plasma glucose in the model (model 2) slightly decreased the relative risk (RR = 1.52; CI 95% = 1.10-2.11). After 65 y, higher body weight was associated with a better outcome, especially in patients belonging to the IV quartile of BMI (RR = 0.74; CI 95% = 0.62-0.90)., Conclusions: In older type II diabetic patients, a moderate excess weight predicts a better survival, while obesity is a negative prognostic factor in patients younger than 65 y. In the latter patients, the effect of obesity on mortality seems to be partly mediated by hypertension, duration of diabetes, and fasting plasma glucose.

Evaluation of quality of life is important in treatment strategies of chronic illnesses, such as diabetes mellitus. The present study aimed at measuring the impact of self-reported exercise on quality of life in T1DM patients. Fifty-three young T1DM patients on intensive insulin-treatment regime who were regularly attending the Diabetes Clinic were selected. At the end of the scheduled clinic visit, they were asked to fill in an Italian version of the Diabetes Quality of Life (DQOL) questionnaire; 30 patients exercised regularly, whereas 23 were sedentary. Patients with clinical and laboratory evidence of major eye, renal and cardiovascular diseases were excluded. A significant difference in the scale of satisfaction (1.9 +/- 0.3 vs 2.2 +/- 0.5; p = 0.009), along with a better HbA1c level (7.0 +/- 1.0 vs 7.8 +/- 1.2; p = 0.014), was observed in physically active patients as compared to sedentary ones, whereas there were no differences between the 2 groups with respect to the impact and worry scale. Moreover, there were no significant correlations between glucose control parameters and the quality of life scores. In conclusion, we showed that self-reported exercise is associated with both a better quality of life and a better metabolic control in young T1DM patients. Further efforts should however be made to recognize all factors that can help motivate patients to exercise.

The purposes of this study were 1) to compare soluble tumor necrosis factor-alpha receptors, which are thought to reflect the degree of TNF-alpha activation, in nondiabetic subjects and type 1 diabetic patients, and 2) to evaluate the effects of smoking and microvascular complications on soluble tumor necrosis factor-alpha receptor levels in type 1 diabetic individuals. Plasma soluble tumor necrosis factor-alpha receptor levels (R1 and R2) were measured in 50 young type 1 diabetic patients without clinical macroangiopathy and in a matched group of 20 healthy volunteers. When diabetic patients were grouped according to smoking and microvascular complication status, the groups of patients had similar values of age, sex, body mass index, blood pressure, lipids, creatinine, and glycometabolic control. Nevertheless, soluble tumor necrosis factor-alpha receptor-R1 levels but not R2 levels, were markedly elevated (P < 0.05 or less) in complicated vs. uncomplicated (2.40 +/- 0.3 vs. 1.80 +/- 0.1 ng/ml) patients and in smokers vs. nonsmokers (2.66 +/- 0.4 vs. 1.76 +/- 0.1 ng/ml). In a two-factor ANOVA, both smoking (P < 0.01) and microvascular complications (P < 0.05) were independent predictors of soluble tumor necrosis factor-alpha receptor-R1. Soluble tumor necrosis factor-alpha receptor levels of diabetic patients who did not smoke or without complications were similar to those of healthy controls. In conclusion, smoking and microvascular complications seem to exert an additive and deleterious impact on TNF-alpha activation, as reflected by levels of soluble tumor necrosis factor-alpha receptors, in young adults with type 1 diabetes.

Monocytes bear insulin receptors similar to those expressed in other tissues, but insulin action in these cells remains unclear. There is evidence that adhesion, by generating a complex array of irreversible transformations, may modify the response of cells to other stimuli, such as hormones. The present study aimed to characterise the pattern of insulin induced tyrosine phosphorylation of monocytes in suspension. Monocytes in suspension were obtained by sequential gradient centrifugation and the tyrosine phosphoproteins were analyzed by immunoblot with antiphosphotyrosine antibodies. The major result of the study is that in suspended monocytes insulin induced a dose and time dependent dephosphorylation of a protein with a molecular mass of about 92 kDa without stimulating the tyrosine phosphorylation of the Insulin Receptor Substrat-1 (IRS-1). In conclusion, we showed that in monocytes in suspension insulin seems to activate a tyrosine phosphatase, which, in turn, dephosphorylates a protein with an apparent molecular weight of 92 kDa.

To study the role of membrane lipids in signal transduction by the insulin receptor, we have studied the effect of phospholipase C (Clostridium perfringens) and a phosphatidylinositol-specific phospholipase (Staphylococcus aureus) on insulin binding, a function of the alpha-subunit, and tyrosine kinase activity, a function of the beta-subunit in IM-9 lymphocytes and NIH 3T3 fibroblasts transfected with the human insulin receptor. Treatment of the cells with phospholipase C at concentrations up to 3.4 U/ml did not affect specific insulin binding, but reduced insulin-stimulated receptor phosphorylation by 50%. This effect of phospholipase C was observed within 10 min of treatment and occurred with no change in the basal level of phosphorylation. Pre-treatment of cells with insulin for 5 min prior to enzyme addition prevented any change in kinase activity. Insulin-stimulated phosphorylation of pp 185, the presumed endogenous substrate for the insulin receptor kinase, was also reduced following phospholipase C treatment, with an almost complete loss of insulin stimulation after exposure of cells to enzyme at concentrations as low as 0.6 U/ml. In contrast to these effects of phospholipase C on intact cells, receptor autophosphorylation was not affected in insulin receptors purified on wheat germ agglutinin-agarose from phospholipase C treated cells. Likewise, the phospholipase C effect was reduced by the addition of phosphatidylcholine, but not by the addition of the protease inhibitors, aprotinin and phenylmethylsulfonyl fluoride, to the incubation indicating its dependence on phospholipid hydrolysis. Treatment of cells with the phosphatidylinositol-specific phospholipase C did not affect any of the parameters studied.(ABSTRACT TRUNCATED AT 250 WORDS)

The mechanism of action of sulphonylureas is not completely understood. In the present study we evaluated the effects of gliquidone, a second-generation compound, on several metabolic parameters in 22 patients with untreated newly-diagnosed type II (noninsulin-dependent) diabetes mellitus. After either 1 or 6 months of treatment with gliquidone plus isocaloric diet we observed: 1) a significant decrease in fasting plasma glucose and glycemic profile after oral glucose load; 2) unchanged fasting and postglucose plasma insulin levels; 3) no change in fasting C-peptide levels but a significant increase in C-peptide concentrations after glucose challenge; 4) a significant increase in glucose disappearance rate from plasma following iv insulin injection; 5) an increase in the insulin-induced reduction of plasma levels of free-fatty acids; 6) no change in plasma C-peptide levels following iv insulin injection; 7) a significant increase in specific insulin binding to monocytes. After 6 but not 1 month of gliquidone therapy we also found an increase in the activity of hexokinase in circulating mononuclear leukocytes. These results suggest that the hypoglycemic effect of gliquidone occurs through either an increased beta cell response to glucose stimulus or an enhanced insulin sensitivity. The latter effect seems to depend on both receptor and postreceptor mechanisms.

The exact relationship between EGF-stimulated tyrosine phosphorylation, induction of the cellular proto-oncogenes c-myc and c-fos, and DNA synthesis remains uncertain. Madin-Darby Canine Kidney (MDCK) cells possess EGF receptor sites with high binding capacity, and in contrast to A431 cells, respond to EGF by increasing DNA synthesis. Following EGF stimulation of intact MDCK cells, there was a rapid and marked increase in the autophosphorylation of the EGF receptor. This was associated with an increase in the tyrosine phosphorylation of a 120 kDa phosphoprotein believed to be an endogenous substrate of this receptor kinase. The ED50 for stimulation of phosphorylation of pp120 was approximately 0.05 nM versus 1.0 nM for receptor autophosphorylation, consistent with amplification of signalling at this step in EGF action. Stimulation of DNA synthesis occurred after 12 to 24 hours and revealed even further amplification with an ED50 of about 0.1 nM. Intermediate between these events was a time-dependent activation of c-fos and c-myc gene expression. However, the ED50 for these processes was approximately 10 nM, indicating a relatively lower sensitivity of EGF for stimulation of proto-oncogene expression. Tyrphostin (RG 50864), a compound reported to inhibit specifically the EGF receptor kinase, completely blocked EGF stimulation of proto-oncogene induction. Interestingly, under the same experimental conditions, EGF receptor autophosphorylation was decreased only 60%. These data, along with the dose-response studies, indicate that proto-oncogene induction requires near maximal stimulation of EGF receptor autophosphorylation. They also suggest that, in MDCK cells, the EGF dependent induction of the c-fos and c-myc genes is not strictly correlated to the extent of EGF receptor autophosphorylation or EGF-stimulated DNA synthesis, and that EGF stimulation of DNA synthesis likely involves additional rate-limiting intermediate steps.

The aim of the present study was to evaluate whether the inhibitory effect on pancreatic A-cell exerted by hyperglycemic hyperinsulinemia and/or by somatostatin administration is impaired in human obesity. For this purpose plasma glucagon concentrations were measured in 8 obese and 8 nonobese nondiabetic subjects during a 4-h hyperglycemic clamp. Synthetic cyclic somatostatin-14 was infused at the rate of 2.5 nmol/min during the third hour of the study. Fasting plasma glucagon was higher in obese than in nonobese subjects (242 +/- 32 vs 163 +/- 15 pg/ml, p less than 0.05) (mean +/- SEM). In the last 20 min of the glucose infusion period preceding somatostatin administration (100-120 min of the study) plasma glucagon averaged 195 +/- 26 pg/ml in obese and 122 +/- 13 pg/ml in nonobese subjects (p less than 0.05), with a reduction of 19 +/- 3% in the former and 28 +/- 4% in the latter (p = n.s.). In both groups somatostatin infusion did not result in a further decrease in plasma glucagon, which averaged 192 +/- 27 pg/ml in obese and 123 +/- 16 pg/ml in nonobese subjects (p less than 0.05) in the 160-180 min period of the study. Also after discontinuing somatostatin infusion plasma glucagon levels did not change. These results suggest that in human obesity hyperglycemic hyperinsulinemia has a normal inhibitory effect on pancreatic A-cell and that somatostatin administration has no additive effect on hyperglycemia and hyperinsulinemia in either obese or nonobese nondiabetic subjects.