Abstract: Twenty-five medical centers and the Prader-Willi Syndrome (PWS) Association collaborated on a study which attempted to identify all people with genetically confirmed diagnosis of PWS living in Italy. Investigators of the participating centers contacted PWS subjects and/or their family, filled in a specially developed form with the required data and forwarded this information by email. The study identified 425 subjects (209 males and 216 females, between the ages of 0.4-46.7). Two hundred thirty-eight patients had del15, 104 had UPD15, 4 demonstrated a translocation affecting chromosome 15 and 79 showed a positive methylation test. There were fewer subjects found over the age of 35, probably due to the low rate of identification of older PWS patients as well as the high mortality rate. There were a greater number of male children and adolescents with PWS whilst, amongst adults, there were more females. As expected, the majority of subjects with PWS were obese, especially in adult life. Nevertheless, it is noteworthy that 26% of patients aged between 6 and 17 were normal weight. A total of 212 subjects had received GH treatment, of which 141 were still receiving therapy, while the remaining 71 had stopped. In children and adolescents (233 cases), 89 subjects had never undergone GH therapy. Eighteen PWS patients had died in the past 20 years. Obesity-related cardiovascular and respiratory diseases were the cause of death, both during childhood and after 18 years of age. Three children died suddenly whilst undergoing GH therapy. Respiratory infection and cardiac illness were the causes of death in two cases. There was no definitive cause of death found in the third case. Overall, there was no increase in number of deaths during GH treatment, suggesting that GH administration in patients with PWS, as a group, does not increase the risk of death. (c) 2008 Wiley-Liss, Inc.

Abstract: Regeneration of mesenchymal tissues depends on a resident stem cell population, that in most cases remains elusive in terms of cellular identity and differentiation signals. We here show that primary cell cultures derived from adipose tissue or skeletal muscle differentiate into adipocytes when cultured in high glucose. High glucose induces ROS production and PKCbeta activation. These two events appear crucial steps in this differentiation process that can be directly induced by oxidizing agents and inhibited by PKCbeta siRNA silencing. The differentiated adipocytes, when implanted in vivo, form viable and vascularized adipose tissue. Overall, the data highlight a previously uncharacterized differentiation route triggered by high glucose that drives not only resident stem cells of the adipose tissue but also uncommitted precursors present in muscle cells to form adipose depots. This process may represent a feed-forward cycle between the regional increase in adiposity and insulin resistance that plays a key role in the pathogenesis of diabetes mellitus.

Abstract: BACKGROUND: Prader-Willi syndrome (PWS) is a congenital alteration of chromosome pair 15. It is characterized by short stature, muscular hypotonia, hyperphagia, obesity, behavioural and emotional disturbances, hypogonadism and partial Growth Hormone (GH) deficiency. The aim of this study was to assess the long-term effect of GH treatment on the psychological well-being and Quality of Life (QoL) in an adult PWS group. METHODS: A total of 13 PWS patients, their diagnosis confirmed by genetic tests, and their parents were recruited for this study. The participants were administered the 36-Items Short Form Health Survey (SF-36) and the Psychological General Well-Being Index (PGWBI), for the assessment of QoL and psychological well-being, at the beginning of GH treatment, and at following intervals of 6, 12 and 24 months. Modified versions of the same questionnaires were given to the parents. RESULTS: Significant improvement with respect to the baseline was found, on both scales, in the evaluation of both physical and psychological well-being, although the parents' evaluation was less optimistic than that of the patients. CONCLUSION: Our findings suggest that the amelioration of QoL and psychological status is sustained in patients who continue GH treatment.

Abstract: CONTEXT: In Prader-Willi syndrome (PWS), an altered GH secretion has been related to reduced cardiac mass and systolic function when compared with controls. OBJECTIVES: The objective of the study was to evaluate the cardiovascular response to GH therapy in adult PWS patients. STUDY PARTICIPANTS: Thirteen obese PWS adults (seven males and six females, aged 26.9+/-1.2 yr, body mass index 46.3+/-1.6 kg/m2) participated in the study. METHODS: Determination of IGF-I, metabolic parameters, echocardiography, and cardioscintigraphy with dobutamine stimulation was made during 12 months GH therapy, with results analyzed by repeated-measures ANOVA. RESULTS: GH therapy increased IGF-I (P<0.0001); decreased C-reactive protein levels (P<0.05); and improved lean mass (P<0.001), fat mass (P<0.05), and visceral fat (P<0.001). Echocardiography showed that 6- and 12-month GH therapy increased left ventricle mass in 76 and in 61% of patients, respectively (P<0.05), did not change diastolic function, and slightly decreased the left ventricle ejection fraction (LVEF) (P=0.054). Cardioscintigraphy documented stable values of LVEF throughout the study, whereas right ventricle ejection fraction decreased significantly (P<0.05) being normally responsive to dobutamine infusion. A positive association between IGF-I z-scores and LVEF occurred at the 6- and 12-month follow-up (P<0.05). CONCLUSIONS: In PWS, GH therapy increased cardiac mass devoid of diastolic consequences. The observation of a slight deterioration of right heart function as well as the association between IGF-I and left ventricular function during GH therapy suggest the need for appropriate cardiac and hormonal monitoring in the therapeutic strategy for Prader-Willi syndrome.

Abstract: BACKGROUND: Alström syndrome (ALMS) is a rare recessively inherited progressive disease (OMIM 203800). Among its diverse spectrum of clinical features are phenotypes associated with deficiencies of the GH/IGF-I axis, including short stature, obesity, insulin resistance, hypertriglyceridaemia and heart failure. PATIENTS AND MEASUREMENTS: To characterize the IGF system in ALMS, we evaluated a subset of 15 young adults with ALMS for hepatic, renal and thyroid function. Glycaemic and hormone measurements such as insulin, GH, FSH, LH, testosterone and 17-beta-oestradiol were clinically assessed. In addition, we measured IGF-I, IGF-II, IGF binding-protein-3 (IGFBP-3) and acid labile subunit (ALS - the subunits that constitute the main somatomedin complex in the circulation), and IGFBP-1 and IGFBP-2 (known to influence the bioavailability of the IGFs). RESULTS: A significantly lower height was observed in ALMS patients compared to age-matched controls. ALMS patients were clinically obese (by weight and body mass index (BMI) standards) and leptin levels correlated with BMI. Renal and hepatic dysfunction was implicated in some patients by increased values of blood urea nitrogen (BUN) and creatinine, and transaminases, respectively. One-third of the patients presented with fasting hyperglycaemia and 80% were hyperinsulinaemic. TSH was slightly increased in 20% of patients. Baseline FSH and LH in females were within the normal range, while half of the males had abnormally low testosterone values. Male patients with hypogonadism showed significantly lower testosterone, oestrogen and ALS levels. Baseline GH values were not found to be increased. ALS and IGFBP-1 were significantly reduced and IGFBP-2 was markedly increased in ALMS patients compared to age-matched controls. The IGFs and IGFBPs were not significantly different between males and females affected with ALMS. No significant association was observed between IGFs or IGFBPs levels and weight, height, BMI, glycaemia, hyperinsulinaemia and testosterone levels. However, we found a significant association of gamma-glutamyltransferase (GGT) with IGFBP-2. IGF-I levels were significantly associated with LH in female patients. CONCLUSIONS: In summary, the reduction of ALS and the increase of IGFBP-2 points to a growth hormone deficiency (GHD) condition in ALMS. However, further tests, including GH dynamics, are needed to determine whether, or to what degree disturbances in the GH/IGF axis contribute to the relatively short stature.

Abstract: BACKGROUND AND AIM: The aim of this study was to investigate lipid content and expression of genes involved in lipid metabolism, in lean and obese non-diabetic rats and in obese rats undergoing food restriction and weight loss. METHODS AND RESULTS: We studied lean and genetically obese Zucker rats (fa/fa). Another group of obese rats were food restricted to lose 20% of initial body weight. We measured expression of genes involved in lipid oxidation and synthesis. Tissue triglyceride content, cell apoptosis and tissue fibrosis were also evaluated. The hearts of obese rats have higher triglyceride content compared to lean controls despite an increased expression of genes involved in fatty acid oxidation (PPARalpha, PGC-1alpha, CPT-I, ACO, UCP3). No differences were found in apoptosis and tissue fibrosis between the two phenotypes. Weight loss resulted in a significant reduction in heart lipid content, while the expression of genes involved in fatty acid oxidation remained elevated. CONCLUSION: In contrast to data reported in diabetic rats, pathways of lipid oxidation are increased rather than decreased in insulin-resistant obese rats. Food restriction reduced heart triglyceride content while lipid-oxidizing genes remained elevated, probably as a consequence of lipid oversupply coming from the endogenous source.

Abstract:

Abstract: BACKGROUND AND AIMS: We evaluated whether hyperinsulinemia and/or insulin resistance are independently associated with plasma lipids, uric acid and blood pressure in non-diabetic subjects. METHODS AND RESULTS: A database of non-diabetic Italian subjects has recently been set up using data from hyperinsulinemic euglycemic clamp studies carried out using the standard technique (40mU per min per square meter of body surface area). In this database we evaluated the relationships between fasting plasma insulin (FPI), glucose metabolized during clamp (M) and plasma levels of triglycerides (TG), high density lipoprotein cholesterol (HDL-C), uric acid (UA) as well as blood pressure (BP) in non-diabetic subjects with fasting plasma glucose <6.1mmol/l. Parallel analyses were conducted in all subjects in the database (n=1093) and in those with all variables available (n=309). In the univariate analysis both FPI and M were significantly correlated with TG, HDL-C, UA and BP (systolic, diastolic and mean). Multivariate regression analyses including center, sex, age, body mass index (BMI), FPI and M as independent variables showed that: (1) TG and UA were positively correlated with FPI and negatively correlated with M; (2) HDL-C was negatively correlated with FPI and positively correlated with M; and (3) BP was negatively correlated with both FPI and M. Analyses of covariance showed that, after adjusting for center, sex, age and BMI, subjects with isolated hyperinsulinemia or isolated insulin resistance had higher TG and UA and lower HDL-C. Subjects with isolated insulin resistance had also higher BP whereas subjects with isolated hyperinsulinemia had lower BP. Subjects with both defects had a worse profile. CONCLUSIONS: Hyperinsulinemia and insulin resistance might contribute with distinct and independent mechanisms to the development of several metabolic and hemodynamic disorders often clustering in the same individual. In particular, hypertriglyceridemia, low HDL-cholesterol and hyperuricemia seem to be related to both hyperinsulinemia and insulin resistance, whereas hypertension seems to be related only to insulin resistance.

Abstract: BACKGROUND: The endogenous cannabinoid system participates in the regulation of energy balance, and its dysregulation may be implicated in the pathogenesis of obesity. Adipose tissue endocannabinoids may produce metabolic and endocrine effects, but very few data are available in human adipose tissue and in primary human fat cells. EXPERIMENTAL DESIGN: We measured expression of type 1 and type 2 cannabinoid receptors (CNR), enzymes of cannabinoids synthesis and degradation in human omental, sc abdominal, and gluteal adipose tissue from lean and obese subjects. Furthermore, we assessed the effect of CNR1 stimulation on glucose uptake and intracellular transduction mechanisms in primary human adipocytes. Then we assessed the reciprocal regulation between CNR1 and peroxisome proliferator-activated receptor-gamma (PPARgamma). Finally, we tested whether leptin and adiponectin are regulated by CNR1 in human adipocytes. RESULTS: We found that most genes of the endocannabinoid system are down-regulated in gluteal fat and up-regulated in visceral and sc abdominal adipose tissue of obese patients. Treatment of adipocytes with rosiglitazone markedly down-regulated CNR1 expression, whereas Win 55,212 up-regulated PPARgamma. Win 55,212 increased (+50%) glucose uptake, the translocation of glucose transporter 4, and intracellular calcium in fat cells. All these effects were inhibited by SR141716 and wortmannin and by removing extracellular calcium. Win 55,212 and SR141716 had no effect on expression of adiponectin and leptin. CONCLUSIONS: These results indicate a role for the local endocannabinoids in the regulation of glucose metabolism in human adipocytes and suggest a role in channeling excess energy fuels to adipose tissue in obese humans.

Abstract: OBJECTIVE: We assessed the accuracy of the American Diabetes Association (ADA)(2003) definition of impaired fasting glucose (IFG) in identifying subjects with low insulin sensitivity, and determined cardiovascular risk factors in ADA(2003) IFG subjects. RESEARCH DESIGN AND METHODS: This study included 930 non-diabetic Italian Caucasians from the GISIR database in which subjects underwent a hyperinsulinaemic-euglycaemic clamp performed with a standard technique. Low insulin sensitivity was defined as being in the lower quartile of glucose metabolized during the last hour of the clamp (M). Subjects were stratified in the following groups: normal fasting glucose (NFG) (<100 mg/dL), IFG100 (100-109 mg/dL), ADA(1997) IFG110 (110-125 mg/dL), and ADA(2003) IFG (100-125 mg/dL). RESULTS: The sensitivity of identifying subjects with low insulin sensitivity increased adopting the ADA(2003) criterion. After Bonferroni correction for multiple comparisons, both IFG100 and ADA(1997) IFG110 showed significantly higher body mass index (BMI), waist, systolic blood pressure (SBP) and diastolic blood pressure (DBP), triglyceride, fasting plasma insulin (FPI) and fasting plasma glucose (FPG), and lower insulin sensitivity as compared with NFG. As compared with IFG100, ADA(1997) IFG110 showed significantly higher BMI, waist, SBP, FPI, FPG, and lower insulin sensitivity. ADA(2003) IFG group showed significantly higher BMI, waist, SBP and DBP, triglyceride, cholesterol, FPI, and FPG, but lower HDL levels and insulin sensitivity compared with NFG subjects. CONCLUSIONS: Although neither the ADA(2003) nor the ADA(1997) definition of IFG appears to be particularly efficacious for the identification of subjects' low insulin sensitivity, lowering the criterion to the ADA(2003) glucose threshold increased the sensitivity without affecting the specificity. ADA(2003) IFG showed a worse cardiovascular risk profile compared with NFG.Copyright (c) 2007 John Wiley & Sons, Ltd.

Abstract: BACKGROUND: A high prevalence of metabolic syndrome has been reported in primary aldosteronism. Low levels of adiponectin, an adipokine with insulin-sensitizing properties, are considered a hallmark of the metabolic syndrome. We evaluated the relationship between adiponectin and insulin sensitivity in primary aldosteronism, with and without metabolic syndrome, compared with essential hypertension. METHODS: Forty patients with primary aldosteronism and 40 matched patients with low-renin essential hypertension (LREH) were studied. Patients with type 2 diabetes were excluded. Each group was divided into two subsets: one including patients with metabolic syndrome and one including patients without metabolic syndrome (ie, hypertension alone or associated with another component of the syndrome). RESULTS: Insulin resistance, defined by increased homeostasis model assessment (HOMA index), was higher in patients with primary aldosteronism than in those with LREH only in the absence of metabolic syndrome (P<.01), whereas in the subsets bearing the syndrome it was similar. Adiponectin levels were lower in primary aldosteronism than in patients with LREH (P<.01). Like HOMA index, the difference was maintained (P<.01) only in the subsets without metabolic syndrome. Adiponectin levels were inversely correlated with HOMA index and positively correlated with potassium levels both in primary aldosteronism (P<.001) or in LREH (P<.05) groups. CONCLUSIONS: Lower adiponectin as well as lower insulin sensitivity in primary aldosteronism compared with LREH seem to result from both direct (aldosterone excess) and indirect (hypokalemia) mechanisms. Therapeutic interventions aimed at correcting both potassium and adiponectin levels by specific antihypertensive agents might improve insulin sensitivity, providing better cardiovascular protection in primary aldosteronism.

Abstract: Alström syndrome is a monogenic recessive disorder featuring an array of clinical manifestations, with systemic fibrosis and multiple organ involvement, including retinal degeneration, hearing loss, childhood obesity, diabetes mellitus, dilated cardiomyopathy (DCM), urological dysfunction, and pulmonary, hepatic, and renal failure. We evaluated a large cohort of patients with Alström syndrome for mutations in the ALMS1 gene. In total, 79 disease-causing variants were identified, of which 55 are novel mutations. The variants are primarily clustered in exons 8, 10, and 16, although we also identified novel mutations in exons 12 and 18. Most alleles were identified only once (45/79), but several were found recurrently. Founder effects are likely in families of English and Turkish descent. We also identified 66 SNPs and assessed the functional significance of these variants based on the conserved identity of the protein and the severity of the resulting amino acid substitution. A genotype-phenotype association study examining 18 phenotypic parameters in a subset of 58 patients found suggestive associations between disease-causing variants in exon 16 and the onset of retinal degeneration before the age of 1 year (P = 0.02), the occurrence of urological dysfunction (P = 0.02), of DCM (P = 0.03), and of diabetes (P = 0.03). A significant association was found between alterations in exon 8 and absent, mild, or delayed renal disease (P = 0.0007). This data may have implications for the understanding of the molecular mechanisms of ALMS1 and provides the basis for further investigation of how alternative splicing of ALMS1 contributes to the severity of the disease.

Abstract: Patients with primary hyperparathyroidism (PHPT) have impaired vasodilation both dependent and independent of endothelium. The aims of our study were to measure three different biochemical markers of endothelial activation, i. e., plasma thrombomodulin, soluble(s) E-selectin, and von Willebrand factor, in PHPT patients before and one year after successful parathyroidectomy, and to distinguish the potential effect of hypercalcemia and/or high parathyroid hormone from that of major cardiovascular risk factors (diabetes mellitus, hyperlipidemia, hypertension, obesity, smoking habit) on endothelial function. Twenty consecutive patients with PHPT subdivided into two groups according to the absence (n = 8) or presence (n = 12) of one or more risk factors, and fifteen healthy normocalcemic subjects were studied. Baseline thrombomodulin levels were similar in the groups with and without risk factors, and in controls. In contrast, sE-selectin and von Willebrand factor were higher in PHPT patients with risk factors than in those without risk factors (p < 0.05 and p < 0.01, respectively) and controls (p < 0.01). Neither thrombomodulin nor sE-selectin changed after parathyroidectomy in either PHPT group. Plasma von Willebrand factor decreased (p < 0.01) in patients without risk factors, while persisting at high levels in patients with risk factors. In conclusion, in spite of a limitation due to the small number of patients, our study suggests that classic cardiovascular risk factors seem to be the main determinants for the high plasma levels of sE-selectin and vWF in PHPT. Together with unaltered thrombomodulin and sE-selectin levels, a plasma vWF decrease after parathyroidectomy might reflect a specific mechanism of its endothelial calcium- and/or PTH-stimulated secretion in some PHPT patients without risk factors. Whether a vWF reduction after parathyroidectomy may be used as a biochemical index for improved endothelial function in PHPT patients without risk factors has yet to be demonstrated in larger studies.

Abstract: CONTEXT: The recently discovered hormone resistin is linked to the development of insulin resistance, but direct evidence of resistin levels in humans with nonalcoholic fatty liver disease (NAFLD) is lacking. METHODS: We conducted this study to assess the relationship between serum resistin and NAFLD. We measured serum resistin and biochemical, hormonal, and histological correlates in 28 NAFLD patients, 33 controls, and 30 obese patients [body mass index (BMI), >30 kg/m2] without NAFLD. RESULTS: Resistin and adiponectin expression were measured in sc adipose tissue by quantitative RT-PCR. Resistin was higher in NAFLD patients compared with controls (5.87 +/- 0.49 vs. 4.30 +/- 0.20 ng/ml; P = 0.002) and obese patients (4.37 +/- 0.27 ng/ml; P = 0.002). Increased resistin mRNA was also found in the adipose tissue of NAFLD patients compared with controls and obese subjects. CONCLUSIONS: Both NAFLD and obese patients had lower adiponectin levels, whereas leptin was increased only in the obese group. No correlation was found between resistin and high-sensitivity C-reactive protein, BMI, homeostasis model assessment, insulin, glucose, transaminases, and lipid values. A positive correlation was found between resistin and histological inflammatory score. These data report increased resistin in NAFLD patients that is related to the histological severity of the disease, but do not support a link between resistin and insulin resistance or BMI in these patients.

Abstract: OBJECTIVE: To explore the changes of ghrelin circulating levels induced by a mixed meal and their relationship with postprandial substrate oxidation rates in overweight and obese children with different levels of insulin sensitivity. METHODS: A group of ten boys (age 9-12 years) with different levels of overweight (standard deviation score of body mass index: 1.6-3.2) was recruited. Body composition was measured by dual-energy X-ray absorptiometry. Insulin sensitivity was assessed by a frequently sampled i.v. glucose tolerance test. Pre-prandial and postprandial (3 h) substrate oxidation was measured by indirect calorimetry. The energy content of the test meal (16% protein, 36% carbohydrate and 48% fat) was 40% of pre-prandial energy expenditure (kJ/day). RESULTS: Pre-prandial serum concentration of total ghrelin was 701.4+/-66.9 pg/ml (S.E.M.). The test meal induced a rapid decrease in ghrelin levels and maximal decrease was 27.3+/-2.7% below baseline. Meal intake induced a progressive increase of the carbohydrate oxidation rate for 45 min after food ingestion, followed by a slow decrease without returning to pre-prandial values. Postprandial cumulative carbohydrate oxidation was 16.9+/-0.8 g/3 h. Insulin sensitivity and postprandial maximal decrease of ghrelin concentration showed a significant correlation (r = 0.803, P < 0.01). Moreover, the postprandial carbohydrate oxidation rate correlated with the area under the curve for both insulin (r = 0.673, P < 0.03) and ghrelin (r = -0.661, P < 0.04). CONCLUSIONS: A relevant association between postprandial insulin-mediated glucose metabolism and ghrelin secretion in children with different levels of overweight was found. It is possible that the maintenance of an adequate level of insulin sensitivity and glucose oxidation may affect appetite regulation by favoring a more efficient postprandial ghrelin reduction.

Abstract: The metabolic syndrome (visceral obesity, dyslipidaemia, hyperglycaemia, and hypertension), has become one of the major public-health challenges worldwide. There has been growing interest in this constellation of closely related cardiovascular risk factors. Recently, definition and relevance of this syndrome have been questioned. In the present review we analyze in detail the epistemological issues regarding definition and the methodological approaches to definition of the metabolic syndrome.

Abstract: AIMS/HYPOTHESIS: Recent studies suggest that wingless-type MMTV integration site family, member 10B (WNT10B) may play a role in the negative regulation of adipocyte differentiation in vitro and in vivo. In order to determine whether mutations in WNT10B contribute to human obesity, we screened two independent populations of obese subjects for mutations in this gene. SUBJECTS AND METHODS: We studied 96 subjects with severe obesity of early onset (less than 10 years of age) from the UK Genetics of Obesity Study and 115 obese Italian subjects of European origin. RESULTS: One proband with early-onset obesity was found to be heterozygous for a C256Y mutation, which abrogated the ability of WNT10B to activate canonical WNT signalling and block adipogenesis and was not found in 600 control alleles. All relatives of the proband who carried this allele were either overweight or obese. Three other rare missense variants were found in obese probands, but these did not clearly cosegregate with obesity in family studies and one (P301S), which was found in three unrelated subjects with early-onset obesity, had normal functional properties. CONCLUSIONS/INTERPRETATION: These mutations represent the first naturally occurring missense variants of WNT10B. While the pedigree analysis in the case of C256Y WNT10B does not provide definitive proof of a causal link of this variant with obesity, the finding of a non-functioning WNT10B allele in a human family affected by obesity should encourage further study of this gene in other obese populations.

Abstract: CONTEXT: Visfatin was recently identified as a protein highly expressed and secreted in adipose tissue with insulin-mimetic effect and is a candidate hormone to help explain the association among adipose tissue expansion, insulin resistance, and type 2 diabetes. OBJECTIVE: The objective of the study was to assess expression of visfatin in lean and obese subjects and in sc and visceral adipose tissue and moreover to explore the role of visfatin on insulin resistance in humans. DESIGN: We measured circulating visfatin and its mRNA expression in sc adipose tissue (SAT) in lean and obese subjects. Furthermore, we measured visfatin mRNA in visceral adipose (VAT) and SAT by quantitative RT-PCR. Finally, plasma visfatin and its mRNA in SAT were measured under free fatty acid-induced insulin resistance in healthy subjects. RESULTS: Plasma visfatin and its mRNA in SAT were significantly lower in obese subjects, compared with normal-weight controls. Both circulating visfatin and SAT visfatin mRNA were negatively correlated with body mass index, whereas no correlation was found with homeostasis model assessment. Significantly higher visfatin mRNA was found in VAT of obese subjects, compared with lean controls. Interestingly, visfatin mRNA in VAT was positively correlated with BMI. Elevation of free fatty acid induced a condition of insulin resistance but did not affect either circulating visfatin or its mRNA. CONCLUSIONS: Our findings show that, in human obesity, plasma visfatin is reduced, whereas visfatin mRNA is differentially regulated in SAT and VAT. Visfatin is not related to insulin resistance either as assessed by homeostasis model assessment or during lipid infusion.

Abstract: Regulation of adipocyte differentiation is an important process in the control of adipose tissue development. So far, adipogenesis has been investigated through the use of various experimental models. In this work, we used human mesenchymal stem cells (hMSCs) obtained from amniotic fluid (AF) as an alternative model more representative of what naturally happens in vivo. In our opinion, these hMSCs are still not influenced by differentiation stimuli and could act in a way more correspondent to the physiological process of adipogenesis, representing also an ethically acceptable alternative to totipotent human embryonic stem cells (ES). Adipocyte differentiation was monitorated following the expressions of key genes. We measured the expression levels of PPARgamma2, PPARgamma-C1alpha, UCP-1, adipsin, and leptin genes using quantitative real-time PCR. We tested our experimental model with two different media. Understanding in vivo adipogenesis mechanisms will shed light on the pathophysiology of many diseases.

Abstract: OBJECTIVE: To elucidate the pathogenesis of HAART-associated lipodystrophy, by investigating the effects of antiretroviral drugs on adipocyte differentiation and gene expression profile. DESIGN AND METHODS: Analysis of gene expression profile by DNA microarrays and quantitative RT-PCR of 3T3-L1 preadipocytes treated with the nucleoside reverse transcriptase inhibitors (NRTI) lamivudine, zidovudine, stavudine, and zalcitabine, and with the protease inhibitors (PI) indinavir, saquinavir, and lopinavir during maturation into adipocytes. RESULTS: Under standard adipogenic differentiation protocols, PI significantly inhibited adipocyte differentiation, as demonstrated by cell viability assay and Oil Red O staining and quantification, whereas NRTI had mild effects on adipogenesis. Gene expression profile analysis showed that treatment with NRTI modulated the expression of transcription factors, such as Aebp1, Pou5f1 and Phf6, which could play a key role in the determination of the adipocyte phenotype. PI also modulated gene expression toward inhibition of adipocyte differentiation, with up-regulation of the Wnt signaling gene Wnt10a and down-regulation of the expression of genes encoding master adipogenic transcription factors (e.g., C/EBPalpha and PPARgamma), oestrogen receptor beta, and adipocyte-specific markers (e.g., Adiponectin, Leptin, Mrap, Cd36, S100A8). CONCLUSIONS: This study identifies new genes modulated by PI and NRTI in differentiating adipocytes. Abnormal expression of these genes, which include master adipogenic transcription factors and genes involved in lipid metabolism and cell cycle control, could contribute to the understanding of the pathogenesis of HAART-associated lipodystrophy.

Abstract: AIMS/HYPOTHESIS: Satellite cells are responsible for postnatal skeletal muscle regeneration. It has been demonstrated that mouse satellite cells behave as multipotent stem cells. We studied the differentiation capacities of human satellite cells and evaluated the effect of the insulin sensitiser rosiglitazone, a well known peroxisome proliferative activated receptor gamma (PPARG) agonist, on their adipogenic conversion. SUBJECTS, MATERIALS AND METHODS: We obtained human satellite cells from human muscle biopsies of healthy subjects by single-fibre isolation and cultured them under myogenic, osteogenic and adipogenic conditions. Moreover, we compared the morphological features and the adipose-specific gene expression profiling, as assessed by quantitative PCR, between adipocytes differentiated from human satellite cells and those obtained from the stromal vascular fraction of human visceral fat. RESULTS: We proved by morphological analysis, mRNA expression and immunohistochemistry that human satellite cells are able to differentiate into myotubes, adipocytes and osteocytes. The addition of rosiglitazone to the adipogenic medium strongly activated PPARG expression and enhanced adipogenesis in human satellite cells, but did not in itself trigger the complete adipogenic programme. Moreover, we observed a decrease in wingless-type MMTV integration site family member 10B and an upregulation of growth differentiation factor 8 expression, both being independent of PPARG activation. CONCLUSIONS/INTERPRETATION: Human satellite cells possess a clear adipogenic potential that could explain the presence of mature adipocytes within skeletal muscle in pathological conditions such as obesity, type 2 diabetes and ageing-related sarcopenia. Rosiglitazone treatment, while enhancing adipogenesis, induces a more favourable pattern of adipocytokine expression in satellite-derived fat cells. This could partially counteract the worsening effect of intermuscular adipose tissue depots on muscle insulin sensitivity.

Abstract: BACKGROUND: Adiponectin is an adipocyte-derived protein with insulin-sensitizing and antiatherogenic properties. Failure to decrease blood pressure (BP) normally during night in hypertensive patients has been independently associated with left ventricular hypertrophy. METHODS: We examined the relationship between adiponectin levels, insulin sensitivity, and left ventricular structure in 40 newly diagnosed never-treated patients with essential hypertension, including 20 patients with a normal night-time pressure decrease (ie, dippers) and 20 patients with BP persistently elevated throughout the 24-h period (ie, nondippers). All subjects had grade 1-2 hypertension, aged 18 to 65 years, no diabetes mellitus, no obesity, no hyperlipidemia, and no cardiopulmonary, renal, or hepatic disease. RESULTS: The two groups of patients were similar for age, sex, body mass index, and had no differences for clinic, 24-h, and diurnal BP, and 24-h, diurnal, and nocturnal heart rate, as well as glucose, total cholesterol, and triglyceride levels. Plasma insulin and homeostasis model assessment (HOMA index) were higher (P < .01), and adiponectin levels were lower (P < .005) in nondippers than in dippers. Adiponectin correlated inversely with HOMA index and insulin levels (r = -0.58, and r = -0.62, respectively, P < .001) in the entire population. Nondippers showed left ventricular mass, relative wall thickness, and measure of early and late diastolic peak flow velocity ratio similar to those of dippers. CONCLUSIONS: In the absence of major cardiovascular risk factors, nondipper essential hypertensive patients show more prominent insulin resistance and lower adiponectin compared to dippers. Therapeutic modulation of adiponectin or insulin resistance might provide additional benefit to the conventional antihypertensive treatment.

Abstract: Epidemiological data show an increased risk of HIV-associated lipodystrophy in women, and sex hormone abnormalities have been reported with highly active antiretroviral therapy (HAART). This study, which demonstrates that oestrogen receptor beta expression is significantly reduced in the subcutaneous adipose tissue of HIV-infected lipodystrophic patients, downregulated by HAART regimens including protease inhibitors (PI), and restored after switching from PI, opens perspectives for the investigation of selective oestrogen receptor modifiers for the management of this syndrome.

Abstract: OBJECTIVES: Nonalcoholic fatty liver disease (NAFLD) is a major cause of liver-related morbidity and is frequently associated with obesity and metabolic syndrome. The recently discovered hormone adiponectin is produced by adipose tissue, and low plasma adiponectin is considered a key factor in the development of the insulin resistance underlying metabolic syndrome. Animal studies suggest that adiponectin may protect against non-alcoholic steatohepatitis, but direct evidence in humans is lacking. We therefore conducted this study to assess the relationship between plasma adiponectin and nonalcoholic fatty liver disease to explore its role in the pathogenesis of this disease. DESIGN AND METHODS: We measured plasma adiponectin and anthropometric, biochemical, hormonal and metabolic correlates in a group of 17 NAFLD patients with diagnosis confirmed by biopsy, and 20 controls with comparable age, body-mass index and sex. Furthermore we compared plasma adiponectin in patients with simple steatosis and steatohepatitis. RESULTS: Plasma adiponectin was significantly lower in NAFLD patients than controls (5.93+/-0.45 vs 15.67+/-1.60ng/ml). Moreover, NAFLD patients were significantly more insulin resistant while having similar serum leptin. Adiponectin was similar in simple steatosis and in steatohepatitis (6.16+/-0.78 vs 5.69+/-0.49ng/ml). An inverse correlation was observed between adiponectin and homeostatic model assessment (HOMA) of insulin resistance (P = 0.008), while adiponectin did not correlate with serum transaminases and lipid values. CONCLUSIONS: These data support a role for low circulating adiponectin in the pathogenesis of NAFLD and confirm the strict association between reduced adiponectin production by adipose tissue, NAFLD and insulin resistance.

Abstract: OBJECTIVE: The aim of this study was to provide a comprehensive meta-analysis of randomized controlled clinical studies on the effects of sibutramine on weight loss and glycemic control in obese subjects with type 2 diabetes. RESEARCH DESIGN AND METHODS: Controlled clinical trials assessing the effect sizes of sibutramine on weight loss effects on glycemia in obese subjects with type 2 diabetes were identified and reviewed using the Cochrane Library, Medline, EMBASE, and a manual search. RESULTS: Eight placebo-controlled, double-blind, randomized trials of sibutramine were included. After sibutramine treatment, the decrease in body weight and waist circumference was significantly greater than in the placebo group. Fasting blood glucose and HbA(1c) significantly decreased after sibutramine treatment. Treatment benefits were seen in plasma triglycerides and HDL, without significant variations in serum total and LDL cholesterol. No differences in systolic blood pressure between the sibutramine and the placebo groups were seen, while recording of diastolic blood pressure and heart rate showed that sibutramine produced a small increase relative to placebo. CONCLUSIONS: A pharmacological approach in a weight management program for patients with type 2 diabetes may be helpful in glycemic control and in the management of other risk factors. Sibutramine may help improve glucose control because it is conducive to weight loss. The reviewed data on the effect of sibutramine further enforce the recommendations that weight management may be the most important therapeutic task for most obese subjects with type 2 diabetes.

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Abstract: CONTEXT: Determinants of insulin resistance in Prader-Willi syndrome (PWS) are not completely understood. The discovery of several adipokines with relevant effects on insulin resistance and cardiovascular complications of metabolic syndrome offered new tools of investigation of insulin resistance in PWS. OBJECTIVE: The purpose of this study was to measure serum resistin and mRNA in adipose tissue of patients with PWS, those with simple obesity, and healthy controls and correlate resistin levels with anthropometric and biochemical features. DESIGN: Twenty-eight adult PWS patients, 29 obese patients, and 25 healthy controls were studied. Anthropometric variables were measured and fasting serum and plasma were collected for measurement of resistin, adiponectin, leptin, lipid profile, glucose, and insulin. RESULTS: Serum resistin and resistin mRNA expression in adipose tissue was significantly higher in PWS patients, compared with both healthy lean controls and obese patients. Moreover, on regression analysis resistin was significantly correlated with body mass index, whereas no significant association was found between resistin and homeostasis model assessment index. A weak association between resistin and adiponectin was found in the PWS group only. However, on multivariate analysis only the correlation between resistin and body mass index remained significant. CONCLUSIONS: These results support a link between circulating resistin and obesity in humans but do not support a role for resistin in human insulin resistance.

Abstract: BACKGROUND: Sudden death and increased prevalence of ventricular arrhythmias have already been described in acromegaly. Although late potentials (LPs) have been proved to be a new technique in detecting patients at risk for ventricular tachyarrhythmias its use in acromegaly is still unknown. METHODS: We studied 70 acromegalic patients [32 males, 38 females; age 49+/-12 years (mean+/-S.D.)] and 70 control subjects age- and sex-matched [(35 males and 35 females; 46+/-12 years (mean+/-S.D.)]. Besides hormonal tests, we performed the following cardiovascular investigations: ECG, 24-h ECG Holter monitoring, echocardiography, and signal-averaged ECG (SAECG) time-domain analysis. RESULTS: LPs occurrence was significantly higher in acromegalic patients as compared to the control group (22.9% vs. 2.9%; p=0.001). A greater duration of disease in patients with positive LPs compared to negative ones was pointed out (18 vs. 12 years; p=0.024). In the group of acromegalic patients with positive LPs we observed a significant association with premature ventricular complexes (PVCs) detected by means of 24-h Holter ECG recording (13 out of 15 patients: 86.7%; p=0.024). The positivity or negativity of LPs proved to be significantly associated with Lown scale PVC trends recorded by 24-h Holter ECG (p=0.014). In the group of patients with left ventricular hypertrophy a significant and pathological worsening of SAECG signals (QRS, LAS, RMS) was documented. CONCLUSIONS: We observed a higher prevalence of LPs in acromegaly which significantly correlated with Lown scale of PVCs.

Abstract: Insulin resistance has been implicated as one possible factor that links visceral obesity to unfavourable metabolic and cardiovascular consequences. However, the mechanism whereby adipose tissue causes alterations in insulin action remains unclear. White adipose tissue is secreting several hormones, particularly leptin and adiponectin, and a variety of other protein signals: the adipocytokines. They include proteins involved in the regulation of energy balance, lipid and glucose metabolism as well as angiogenesis, vascular and blood pressure regulation. Visceral obesity and inflammation within white adipose tissue may be a crucial step contributing to the emergence of insulin resistance, type 2 diabetes and atherosclerosis. A growing list of adipocytokines involved in inflammation (IL-1beta, IL-6, IL-8, IL-10, TNF-alpha, TGF-beta,) and the acute-phase response (serum amyloid A, PAI-1) have been found to be increased in the metabolic syndrome. It is, however, unclear as to the extent adipose tissue contributes quantitatively to the elevated circulating levels of these factors in obesity and how they may affect the insulin-dependent tissues. This review describes the role of the currently known adipocytokines and hormones released by adipose tissue in generating the insulin resistance state and the chronic inflammatory profile which frequently goes together with visceral obesity.

Abstract: CONTEXT: Adult patients with Prader-Willi syndrome (PWS) are prone to develop obesity, GH deficiency (GHD), and their related complications, with cardiopulmonary failure explaining more than half of PWS fatalities. OBJECTIVE AND STUDY PARTICIPANTS: This study was undertaken to examine the effect of GHD and sleep breathing disorders on cardiovascular risk factors and heart features of 13 PWS (age 26.9 +/- 1.2 yr) and 13 age-, gender-, and body mass index-matched obese individuals (age 26.2 +/- 0.8 yr). RESULTS: Compared with controls, PWS patients had lower GH response to arginine+GHRH, IGF-I levels, triglycerides, total and LDL-cholesterol, insulin, and insulin resistance measured by a homeostatic model approach. Dual-energy x-ray absorptiometry, abdominal computed tomography scans, and polysomnography revealed a greater fat mass, similar abdominal fat, but greater sleep breathing disorders in PWS than obese subjects. Echocardiography showed no systolic or diastolic alteration, although PWS had lower left ventricle (LV) mass (135.7 +/- 7.7 vs. 163.5 +/- 8.4 g, P < 0.05) and near significantly lower values of LV end-diastole diameter (P = 0.08), compared with obese controls. Baseline radionuclide angiography documented comparable values of systolic and diastolic values between groups. However, adrenergic stimulation with dobutamine caused a lower increase of LV ejection fraction (71.9 +/- 1.9 vs. 76.3 +/- 1.2%, P < 0.05) and heart rate (103 +/- 6.9 vs. 128 +/- 2.8 beats/min, P < 0.05) in PWS than obese individuals. By multivariate analysis, nocturnal oxygen desaturation and IGF-I levels were main significant predictors of LV mass and heart rate in PWS patients. CONCLUSIONS: PWS differs from simple obesity by a healthier metabolic profile, impaired nocturnal breathing, decreased heart geometry, and systolic and chronotropic performance. GHD and the predictive role of IGF-I on structural and functional heart parameters suggest a GH/IGF-I-mediated control of cardiac risk in PWS.

Abstract: Myostatin is a member of transforming growth factor-beta superfamily that plays an important inhibitory role during muscle development; in fact mutations of myostatin gene result in a hypermuscular phenotype. Moreover myostatin-deficient mice have a significant reduction in fat depots and a depression of adipogenesis. Little is known about myostatin function in muscle growth regulation in humans and in particular during caloric restriction. In the present work we quantified by real-time RT-PCR myostatin expression in muscle biopsies of a group of morbidly obese patients before and after weight loss obtained by biliopancreatic diversion (BPD). The patients reduced body weight by 38.9%, mostly due to fat-mass loss, showing also a significant reduction in the 24-hour EE as assessed by the respiratory chamber. Myostatin mRNA levels result clearly decreased after weight loss, suggesting a role in counteracting the progressive decline of muscle mass after BPD. Myostatin may provide therefore another mechanistic explanation for the control of energy partitioning between protein and fat, working against muscle wasting. Our data suggest that myostatin might represent an important regulator of skeletal muscle size also in conditions of food restriction in obese subjects.

Abstract: Glucose transporter 4 (GLUT4) seems to be involved in the mechanism of insulin resistance in polycystic ovary syndrome (PCOS) patients (PCOSs) in both muscular and adipose tissue. The observation that insulin stimulates glucose oxidation in endometrial cells led us to investigate the presence of GLUT4 in this tissue and whether a defect of GLUT4 is present at the endometrial level in PCOSs. We also investigated whether body weight influences GLUT4 expression in this syndrome. GLUT4 mRNA content was examined by real-time quantitative RT-PCR and immunostaining reaction in the endometrial tissue of nine normal subjects, nine lean and eight obese hyperinsulinemic (h-INS), and eight lean and 10 obese normoinsulinemic (n-INS) PCOSs. GLUT4 mRNA and its positive immunostaining reaction were present in epithelial cell level in the endometrium of both normal and PCOS subjects. Significantly higher levels of GLUT4 were observed in normal and lean n-INS PCOSs in comparison with other groups. In both n-INS and h-INS obese PCOSs, GLUT4 was significantly lower than in lean subjects. However, obese n-INS and lean h-INS PCOSs showed a similar low GLUT4 expression, whereas obese h-INS PCOSs showed the lowest expression when compared with other groups. In conclusion, our data demonstrate that GLUT4 is present in the endometrium of normal and PCOS subjects and that hyperinsulinism and obesity seem to have a negative effect on endometrial GLUT4 expression in PCOS.

Abstract: The bradykinin B2 receptor (B2R) gene is a candidate in the pathogenesis of insulin resistance, which often clusters with other abnormalities in metabolic syndrome. We investigated the distribution of the C-58T B2R gene polymorphism within a population of overweight/obese patients (BMI > or = 25 kg/m2) potentially characterised by different levels of insulin resistance. Patients with type 2 diabetes, dyslipidemia and hypertension were excluded in order to distinguish the effect of obesity on insulin sensitivity from that of confounding factors. Ninety-two unrelated adults (41 men and 51 women, aged 33.7 +/- 11.6 years) were recruited by random sampling from a general population evaluated for cardiovascular risk stratification. Measurements included BMI, waist circumference, body composition, blood pressure, serum leptin, and lipid profile. Insulin sensitivity was calculated according to the homeostasis model assessment (HOMA) method. C-58T genotypes--CC (n = 20), CT (n = 47) and TT (n = 25)--were determined by restriction fragment-length polymorphism PCR. Patients subdivided on the basis of C-58T polymorphism, showed no difference in any of the parameters examined, including HOMA index values, after adjustment for age, sex, BMI and waist circumference. The results indicate that the C-58T B2R gene polymorphism is not associated with different levels of insulin resistance within a population of obese patients.

Abstract: OBJECTIVE: Glucocorticoids were found to inhibit adiponectin gene expression and secretion both in vitro and in animal models. We evaluated first the acute effect of i.v. glucocorticoids on adiponectin in normal subjects and secondly plasma adiponectin levels in a series of patients with Cushing's syndrome compared with controls. DESIGN AND METHODS: Hydrocortisone (25 mg) was administered i.v. to five healthy volunteers, with blood samples taken at -15, 0, 30, 60, 120 and 180 min. Twenty-one patients with Cushing's syndrome were divided in two groups: one with 11 obese and the other with 10 non-obese Cushing's patients. Each group was compared with controls that were matched for sex, age, body mass index, waist circumference, glucose, insulin, lipid levels and blood pressure. RESULTS: In normal subjects, hydrocortisone produced a decrease in adiponectin at 30 and 60 min, compared with placebo (P<0.05). Adiponectin was lower in non-obese Cushing's patients than in non-obese controls (P<0.004). In contrast, there was no difference in adiponectin levels in obese Cushing's patients and in obese controls. Adiponectin was inversely correlated (P<0.05) with homeostasis model assessment index in both obese and non-obese Cushing's patients; in non-obese Cushing's patients only, adiponectin was inversely correlated with urinary cortisol (P<0.05). CONCLUSIONS: Glucocorticoids inhibit adiponectin in man, as shown by both exogenous administration to healthy subjects and endogenous cortisol hyperproduction. Similar levels of adiponectin in obese Cushing's patients and their obese controls indicate that obesity per se may act as a predominant factor in masking the relationship between adiponectin and cortisol.

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Abstract: A reduced lipid oxidative capacity is considered a risk factor for the development of obesity, but a further impairment of lipid oxidative capacity is observed after weight loss. We aimed to define the mechanisms underlying this phenomenon in skeletal muscle and in particular to study the mitochondrial and peroxisomal lipid oxidative pathways. Thus we measured intramyocellular triglyceride content (IMTG) and the expression of genes of lipid oxidation [peroxisome proliferator-activated receptor-alpha, carnitine palmitoyltransferase 1B, and acyl-coenzyme A (acyl-CoA) oxidase 1] and synthesis (acetyl-CoA carboxylase B) using RT-PCR analysis in muscle biopsies of morbidly obese patients before and after biliopancreatic diversion. Weight reduction significantly decreased IMTG while increasing insulin sensitivity, measured by euglycemic hyperinsulinemic clamp. Moreover, an increase in glucose and a decline in lipid oxidation, as assessed by respiratory chamber, were observed. Weight loss reduced the expression of peroxisome proliferator-activated receptor-alpha (-46.7%), carnitine palmitoyltransferase 1B (-43.1%), acyl-CoA oxidase 1 (-37.8%), and acetyl-CoA carboxylase B (-48.7%). Our results indicate that a defect of both peroxisomal and mitochondrial oxidative pathways at the muscular level may contribute to the reduced fat oxidation in obese subjects after biliopancreatic diversion. They also suggest that a depression of the de novo lipogenesis may account for IMTG depletion.

Abstract: BACKGROUND: Leptin has been shown to be linked to adiposity and insulin resistance in middle-aged participants. However, the association between leptin and metabolic syndrome independently of body fat and body fat distribution has not been evaluated in healthy elderly people. METHODS: We studied the independent relation between leptin and the components of the metabolic syndrome in 107 women aged 67-78 years with body mass index (BMI) ranging from 18.19 to 36.16 kg/m2. In all participants, we evaluated BMI, waist and hip circumferences, body composition by dual energy X-ray absorptiometry, fasting, and 2-hour glucose, lipids, insulin, homeostasis model assessment of insulin resistance (HOMA), systolic (SBP), diastolic blood pressure (DBP), and leptin. RESULTS: Significant correlation was found between leptin, BMI, waist circumference, fat mass, DBP, SBP, cholesterol, triglycerides, insulin, and HOMA. After adjusting for age and waist circumference, as well for age and fat mass, leptin was significantly related to insulin levels, HOMA, and cholesterol. In a stepwise multiple regression analysis using insulin levels or HOMA as dependent variables and age, waist circumference, fat mass, leptin, SBP, DBP, cholesterol, and triglycerides as independent variables, leptin entered the regression first, waist circumference second, and age third. CONCLUSION: Our study shows that leptin is significantly related to indices of adiposity in elderly women, and leptin is significantly associated with insulin levels, HOMA, and cholesterol independent of age, body fat, and fat distribution. Leptin, waist circumference, and age together explained 31% and 33% of insulin levels and HOMA variance, respectively, in healthy elderly women.

Abstract: OBJECTIVE: It is under debate whether free fatty acids (FFAs) play an independent role in the regulation of adipose cell functions. In this study, we evaluated whether leptin secretion induced by FFA is due directly to an increased FFA availability or whether it is mediated by insulin levels. RESEARCH METHODS AND PROCEDURES: To test this hypothesis, we compared the effects of six different experimental designs, with different FFA and insulin levels, on plasma leptin: euglycemic clamp, euglycemic clamp + FFA infusion, FFA infusion alone, FFA + somatostatin infusion, somatostatin infusion alone, and saline infusion. RESULTS: Our results showed that euglycemic clamp, FFA infusion, or both in combination induced a similar increment of circulating leptin (3.31 +/- 0.30, 3.40 +/- 0.90, and 3.35 +/- 0.80 ng/mL, respectively). Moreover, the inhibition of FFA-induced insulin increase by means of somatostatin infusion completely abolished the rise of leptin in response to FFA (1.05 +/- 0.30 vs. 3.40 +/- 0.90 ng/mL, p < 0.001). DISCUSSION: In conclusion, our data showed that the effects of high FFA levels on plasma leptin were mediated by the rise of insulin concentration. These data confirm a major role for insulin in the regulation of leptin secretion from rat adipose tissue and support the hypothesis that leptin secretion is coupled to net triglyceride synthesis in adipose tissue.

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Abstract: Polycystic ovary syndrome (PCOS) is one of the most common endocrine metabolic diseases and is characterized by obesity in approximately 50% of those affected. Adiponectin is an adipocyte-derived protein that possesses an antiatherosclerotic action and improves insulin sensitivity. Peroxisome proliferator-activated receptor-gamma (PPAR-gamma) regulates the transcription of several adipocyte-specific genes. The aim of this study was to investigate the putative influence of the PPAR-gamma gene Pro12Ala polymorphism on the adiponectin levels in PCOS and healthy women. One hundred twenty women with PCOS and 120 healthy women whose ages and body mass indexes matched those of the PCOS patients were investigated. The genetic analysis of PPAR-gamma gene Pro12Ala polymorphism was performed by restriction fragment of polymorphisms. Serum adiponectin levels were evaluated, and the homeostasis model assessment score was also calculated. No subject was homozygous for the Ala12 allele of the PPAR-gamma gene. No significant differences in body mass index, plasma glucose and lipid levels, and homeostasis model assessment scores were observed between and within genotype groups in PCOS and control women. No significant differences in serum adiponectin concentrations were observed between and within genotype groups in PCOS and control women.In conclusion, our results confirm that adiponectin concentrations are similar in PCOS and controls and demonstrate no effect of the PPAR-gamma gene Pro12Ala polymorphism on serum adiponectin levels.

Abstract: The aim of this study was to evaluate the morphometric changes of adipose tissue of lean and obese rats as assessed by computerized image analysis (IA) system in experimental conditions, with different degrees of adiposity. Moreover, to validate measures obtained by image analysis by correlation with direct measures of adiposity (body weight, epididimal fat, mean fat cell size and serum leptin). Finally to correlate these changes to expression of genes involved in lipid deposition and mobilization in adipose tissue. Lean (Fa/?) and genetically obese (fa/fa) Zucker rats were studied. Obese rats were food-restricted or treated with retinoic acid (ATRA) in order to reduce body weight and fat content. Moreover, gene expression of two key enzymes involved in fat metabolism (HSL and DGAT) were assessed in adipose tissue by RT-PCR. Our results show that HSL expression in adipose tissue was lower in obese compared to lean rats (1.47+/-0.02 vs 0.35+/-0.03, p<0.005) and was upregulated during food restriction in obese rats. DGAT expression was similar in lean and obese rats and was reduced by treatment with ATRA in obese rats. Tissue texture assessed by IA was significantly higher in lean compared to obese rats (23.2+/-0.6 vs 11.6+/-2.4%; p=0.01). Tissue structure highly correlated with adiposity in obese rats with different amount of body fat (area fraction vs epididimal fat depot: p=0.001). Distribution of measures for each sample, an index of spread of adipose tissue texture, as expressed by the standard deviation, correlated with adiposity (standard deviation vs epididimal fat depot: p=0.002) thus suggesting that adipose tissue texture increases its heterogeneity when adiposity is lower. This observation is in agreement with the hypothesis that the process of lipid mobilization from adipose tissue is not uniform, but a subpopulation of slimming adipocytes undergoes a complete release of their fat content while the rest of the tissue is much less affected. Moreover, image analysis system seems a reliable quantitative tool for assessment of adipose tissue texture.

Abstract: GLUT4 is the most important glucose transporter in insulin-dependent tissues. A decrease of its expression by the adipocytes was reported in polycystic ovary syndrome (PCOS), regardless of obesity and glucose tolerance. In PCOS, abnormal menstrual cycles, abnormal insulin secretory patterns, and obesity, which are risk factors for endometrial diseases, frequently coexist. The endometrial effects of insulin are direct through specific insulin receptors. However, it is unknown whether the endometrium expresses GLUT4 and can be considered an insulin-regulated tissue. In this study, we investigated this question, and we investigated whether obesity modulates this expression in PCOS normoinsulinemic patients. We assayed GLUT4 in the endometrial samples from 18 normoinsulinemic PCOS patients and 9 controls in the advanced follicular phase of the cycle; 10 patients were lean and 8 obese, and all were aged between 23 and 32 years. Most tissue was immediately frozen for RT-PCR; some tissue was saved for histology and immunohistochemistry. GLUT4 mRNA expression was measured in three samples for every patient and expressed as mean +/- SE of an arbitrary unit. In obese PCOS subjects, endometrial GLUT4 expression was significantly lower than in the lean ones (24.0 +/- 6.8 vs. 65.2 +/- 24.4; P < 0.005) and the controls (53.2 +/- 10.7). Lean PCOS and control subjects showed similar values. GLUT4 immunostaining was strong in the epithelial and absent in the stromal cells. We demonstrated endometrial GLUT4 expression. The similar results in lean PCOS and control subjects suggest that endometrial GLUT4 expression is not affected by PCOS itself, whereas it is reduced by obesity in PCOS patients.

Abstract: BACKGROUND: The aim of this study was to evaluate the relation between adiponectin and leptin, fat distribution, and insulin resistance in elderly men and women. METHODS: 68 elderly participants (28 men and 40 women) aged 66-77 years, with body mass index (BMI) ranging from 19.83 to 37.18 kg/m2, participated in the study. In all participants, we evaluated BMI, waist and hip circumferences, sagittal abdominal diameter (SAD), fat mass (FM) by dual energy X-ray absorptiometry, fasting and 2-hour glucose, insulin, homeostasis model assessment of insulin resistance (HOMA), leptin, and adiponectin. RESULTS: Elderly women had significantly higher circulating levels of adiponectin and leptin compared to men even after adjusting for age, FM, or waist circumference. In men and women, leptin was positively associated, whereas adiponectin was negatively associated, with BMI, indices of body fat distribution, as well as FM and FM%. Both fasting insulin and HOMA showed significant positive correlation with leptin and negative correlation with adiponectin in both sexes. In a step-wise multiple regression model with HOMA as the dependent variable and age, gender, waist circumference, FM, leptin, and adiponectin as independent variables, waist entered the regression first, explaining 19.7% of HOMA variance, leptin was second, and adiponectin was third, explaining each one an additional 10% of variance. In a multiple linear regression analysis, leptin and adiponectin alone explained up to 38% of HOMA variance. CONCLUSION: Leptin and adiponectin together seem to be strictly related to insulin resistance in elderly people, independently of body fat and body fat distribution.

Abstract: OBJECTIVE: Little is known about the physiological role and the regulation of uncoupling proteins-2 and -3 (UCP-2 and -3) in adipose tissue. We investigated whether the expression of UCP-2 and -3 in adipose tissue was affected by weight loss due to a biliopancreatic diversion (BPD) and related to the daily energy expenditure (24-h EE). DESIGN: Ten morbidly obese subjects (mean body mass index +/- s.e.m.=49.80 +/- 2.51 kg/m(2)) were studied before and 18+/-2 Months after BPD. METHODS: We determined body composition using tritiated water and 24-h EE in a respiratory chamber. Adipose tissue UCP-2 and -3 mRNA, plasma insulin, glucose, free fatty acids (NEFA), free triiodothyronine (FT3), free thyroxine (FT4) and leptin were assayed before and after BPD. RESULTS: BPD treatment resulted in a marked weight loss (P<0.001) mainly due to a fat mass reduction. A significant decrease in 24-h EE/fat-free mass (FFM) (P<0.05) and in UCP-2 (P<0.05) and UCP-3 (P<0.05) mRNA was observed. A significant reduction in plasma insulin, glucose, NEFA, FT3, FT4 and leptin was seen after BPD. The decline in plasma leptin and FFA was tightly correlated with the decrease in both UCP-2 and -3. A significant correlation was found between changes in FT3 and variations in 24-h EE (r=0.64, P<0.05). In a multiple-regression analysis changes in 24-h EE/FFM after BPD were significantly correlated with changes in UCP-3 expression (P<0.05). CONCLUSION: These findings suggest that UCPs in adipose tissue may play a role in the reduction in 24-h EE observed in post-obese individuals.

Abstract: The aim of the present study was to examine the effects of anorexia nervosa (AN) on adipocytokines (leptin and adiponectin) plasma concentrations and insulin-stimulated glucose disposal in adolescent and young adult women. Adiponectin and leptin plasma levels, along with insulin-stimulated glucose disposal (as measured by the euglycemic-hyperinsulinemic glucose clamp) and oxidative and nonoxidative glucose metabolism (as measured by indirect calorimetry during the last 60 min of the insulin clamp), were measured in 11 anorectic patients and 26 normal-weight healthy female controls. Leptin levels were significantly lower in AN patients, according to the reduced body mass index and their respective fat mass. On the contrary, adiponectin plasma levels were significantly higher in AN patients than in control women. Likewise, insulin-stimulated glucose disposal and nonoxidative glucose metabolism were significantly lower in AN patients. In conclusion, our study shows that young women affected by AN have higher adiponectin plasma levels than healthy female controls of similar age, despite the presence of an impairment of insulin-stimulated glucose disposal, with a prevalent failure of nonoxidative glucose metabolism. Taken together, these data suggest that the reduction of fat mass may play the major role in the control of adiponectin release, with respect to changes in insulin sensitivity.

Abstract: Serum adiponectin levels were evaluated in 60 women with polycystic ovary syndrome (PCOS), 30 normal-weighted and 30 obese women, and 60 healthy women age and body mass index (BMI) matched with the patients. The homeostasis model assessment (HOMA) score was also calculated. Both in PCOS and controls, serum adiponectin levels were significantly (P < 0.05) lower in obese than normal-weight women, without any difference between PCOS and controls. The HOMA score was significantly (P < 0.05) higher in obese than normal-weight women both in PCOS and controls; additionally, the HOMA score was significantly (P < 0.05) higher in normal-weight PCOS than normal-weight controls. Both in PCOS and controls, adiponectin levels were significantly correlated with BMI (r = -0.51, P < 0.01 in PCOS; r = -0.45, P < 0.01 in controls) and HOMA values (r = -0.39, P < 0.05 in PCOS; r = -0.35, P < 0.05 in controls); HOMA was correlated with BMI (r = 0.51, P < 0.01 in PCOS, r = 0.61, P < 0.001 in controls). In conclusion, our results confirm that adiponectin concentrations change according to variations of fat mass. They further suggest that insulin sensitivity per se probably does not play any pivotal role in the control of adiponectin levels in PCOS women.

Abstract: Eating behavior is a complex phenomenon, resulting from the interaction in the hypothalamus and other brain regions, of many factors, including olfactory, visual, emotional and higher cognitive inputs, as well as several nutritional signals coming from the periphery. These signals modulate the expression of neurotransmitters and neuropeptides with orexigenic and anorexigenic activity. Observations performed more than 5 decades ago with brain lesioning and stimulation experiments led to the proposal of the dual centre hypothesis in the central control of energy balance. On the basis of these studies the "satiety centre" was located in the ventromedial hypothalamic nucleus, since lesions of this region caused overfeeding, while its electrical stimulation suppressed eating. On the contrary, lesioning or stimulation of the lateral hypothalamus elicited the opposite set of responses, thus leading to the conclusion that this area represented the "feeding centre". The subsequent expansion of our knowledge of specific neuronal subpopulations involved in energy homeostasis has replaced the notion of specific "centres" controlling energy balance with that of discrete neuronal pathways fully integrated in a more complex neuronal network. This review will focus on the central and peripheral factors thought to be involved in the neuroendocrine control of feeding behavior.

Abstract: OBJECTIVE: To assess the relationship between circulating leptin levels, bone mineral content and density in the elderly. DESIGN: A cross-sectional study. PATIENTS: A cohort of 92 men and 171 women, with ages ranging from 68 to 75 years, selected as a healthy and normal functioning group, in the city centre of Verona. MEASUREMENTS: Plasma leptin levels were determined in each participant. Body composition was evaluated with dual energy X-ray absorptiometry (DXA). Bone mineral content (BMC) and bone mineral density (BMD) were measured at whole-body, hip and femoral neck level in all subjects. RESULTS: In both men and women a significant relationship between fat mass and whole-body BMC or BMD was found. The strength of this association was consistently reduced after adjustment for plasma leptin. A significant association between circulating leptin levels, whole-body, total hip and femoral neck BMC and BMD was found in both sexes. This association retained the statistical significance after adjustment for fat mass percentage, especially in women. In stepwise multiple linear regression analyses, leptin was shown to be a significant predictor of whole-body, total hip and femoral neck BMC and BMD, independently of age and the percentage of body fat in both sexes. The circulating levels of leptin accounted for a variance in whole-body BMC of 8.9% in men and 18.2% in women, and in whole-body BMD of 10.6% in women. CONCLUSION: Our data show a significant relationship between leptin, bone mineral mass and density in healthy elderly men and women.

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Abstract: Moderate alcohol consumption has a cardioprotective effect on coronary artery disease. Among the beneficial effects of alcohol, a suppression of the plasma free fatty acid (FFA) concentration has been shown but the mechanism which accounts for this action is not clear. We assessed whether moderate alcohol intake affects plasma adiponectin levels and tumor necrosis factor (TNF)-alpha, two regulators of lipolysis. Oral glucose tolerance tests were performed twice on 22 volunteers: "the alcohol study" and "control study". In the former, red wine was sipped to maintain steady state alcohol concentration. Samples for plasma glucose, insulin, FFA, adiponectin, and TNF-alpha concentrations were obtained. In the latter, tap water was sipped. Insulin action has been assessed by the Oral Glucose Insulin Sensitivity (OGIS) Model. The mean blood alcohol concentration was 5+/-2 mg/dl. No differences were observed between the two studies in the OGIS (406+/-19 ml x min(-1) x m(-2) with alcohol and 402+/-20 without, respectively). Baseline FFA levels were lower in the alcohol study; however, post-glucose inhibition was comparable. No differences in the TNF-alpha and adiponectin responses were observed. A significant correlation was observed between the OGIS index and the fasting adiponectin level (r=0.589, p<0.0001). Moderate red wine intake improves post-glucose FFA profiles but does not modify the plasma concentrations of both TNF-alpha and of adiponectin concentrations: the latter is significantly and positively associated to the insulin action. Further studies are needed to clarify the antilipolytic effect of moderate alcohol intake.

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Abstract: BACKGROUND: Our aim was to investigate the regulation of the gene expression of leptin in subcutaneous adipose tissue biopsies in morbid obesity before and after biliopancreatic diversion (BPD). METHODS: Longitudinal study in morbidly obese subjects investigated twice: before and 6 months after BPD. Fourteen morbidly obese women, 37+/-13 years old and with a body mass index of 51.6+/-8.2 kg/m2, were studied before and 6 months after BPD (40.6+/-8.0 kg/m2). Using reverse transcriptase polymerase chain reaction analysis, the mRNA expression of leptin was investigated in adipose tissue. Plasma leptin was measured by radioimmunoassay; plasma insulin was measured by microparticle enzyme immunoassay. Free fatty acids (FFA) were measured using a colorimetric kit. RESULTS: A significant decrease in leptin mRNA level was observed in comparison with pretreatment in BPD patients (59+/-34 vs 143+/-85 arbitrary units, P<0.01). A strict relationship between adipose tissue leptin mRNA and plasma leptin either before (R2=0.80, P<0.0001) or after BPD (R2=0.86, P<0.0001) and between plasma FFA concentration and insulin either before (R2=0.65, P<0.001) or after BPD (R2=0.92, P<0.0001) was observed. Finally, a significant correlation was found between changes in FFA and insulin (R2=0.64, P<0.001), insulin and leptin (R2=0.88, P<0.0001), and insulin and leptin mRNA (R2=0.83, P<0.0001). CONCLUSION: These data demonstrate a high correlation between leptin mRNA expression in adipose tissue and plasma leptin in postobese subjects after BPD. The significant relationship between both leptin mRNA and plasma leptin with insulin suggests that circulating insulin might regulate leptin expression. It might be hypothesized that plasma FFA concentration can act on the insulin secretion and subsequently on the leptin secretory pathway.

Abstract: Obesity is a frequent cause of insulin resistance and poses a major risk for diabetes. Abnormal fat deposition within skeletal muscle has been identified as a mechanism of obesity-associated insulin resistance. We tested the hypothesis that dietary lipid deprivation may selectively deplete intramyocellular lipids, thereby reversing insulin resistance. Whole-body insulin sensitivity (by the insulin clamp technique), intramyocellular lipids (by quantitative histochemistry on quadriceps muscle biopsies), muscle insulin action (as the expression of Glut4 glucose transporters), and postprandial lipemia were measured in 20 morbidly obese patients (BMI = 49 +/- 8 [mean +/- SD] kg x m(-2)) and 7 nonobese control subjects. Patients were restudied 6 months later after biliopancreatic diversion (BPD; n = 8), an operation that induces predominant lipid malabsorption, or hypocaloric diet (n = 9). At 6 months, BPD had caused the loss of 33 +/- 10 kg through lipid malabsorption (documented by a flat postprandial triglyceride profile). Despite an attained BMI still in the obese range (39 +/- 8 kg x m(-2)), insulin resistance (23 +/- 3 micromol/min per kg of fat-free mass; P < 0.001 vs. 53 +/- 13 of control subjects) was fully reversed (52 +/- 11 micromol/min per kg of fat-free mass; NS versus control subjects). In parallel with this change, intramyocellular-but not perivascular or interfibrillar-lipid accumulation decreased (1.63 +/- 1.06 to 0.22 +/- 0.44 score units; P < 0.01; NS vs. 0.07 +/- 0.19 of control subjects), Glut4 expression was restored, and circulating leptin concentrations were normalized. In the diet group, a weight loss of 14 +/- 12 kg was accompanied by very modest changes in insulin sensitivity and intramyocellular lipid contents. We conclude that lipid deprivation selectively depletes intramyocellular lipid stores and induces a normal metabolic state (in terms of insulin-mediated whole-body glucose disposal, intracellular insulin signaling, and circulating leptin levels) despite a persistent excess of total body fat mass.

Abstract: There is increasing evidence that the abdominal obesity phenotype may be associated with multiple alterations of the hypothalamic-pituitary-adrenocortical (HPA) axis activity in both sexes. Our hypothesis is that the lack of adequate cortisol suppression after the dexamethasone test may constitute an indirect marker of HPA axis hyperactivity in the presence of the abdominal obesity phenotype. A total of 34 normal-weight (13 men and 21 women) and 87 obese (36 men and 51 women), healthy, nondepressed subjects therefore underwent four different dexamethasone suppression tests randomly performed at varying intervals of at least 1 wk between each test. After a standard overnight 1-mg dexamethasone test, which served as a reference, three other tests were randomly performed at 1-wk intervals by administering 0.0035, 0.0070, and 0.015 mg oral dexamethasone per kilogram of body weight overnight. Blood samples were obtained for cortisol, ACTH, and dexamethasone. Results were analyzed separately in men and women as well as in normal-weight [body mass index (BMI) < or = 25 kg/m(2)] and overweight or obese (BMI > 25 kg/m(2)) subjects. The waist circumference and the waist to hip ratio (WHR) were used as markers of body fat distribution. After the standard 1-mg test, cortisol suppression was greater than 90% in all subjects. However, after each test, obese women had significantly higher values of percent cortisol and percent ACTH suppression than normal-weight women without any difference between obese and normal-weight men. Considering the response to the three variable-dose tests, a clear dose- response pattern (P < 0.001 for trend analysis) in percent cortisol and percent ACTH suppression was found in all subjects. After each test men had significantly higher dexamethasone levels than women, regardless of BMI. However, obese women, but not men, had significantly higher dexamethasone levels after each test than their normal-weight counterpart. Plasma dexamethasone concentrations were dose related (P < 0.001 for trend analysis) in all subjects, but the dose-related increase was significantly higher in normal-weight men than normal-weight women, whereas it was similar in obese subjects of both sexes. Stepwise multiple regression analysis revealed that both percent cortisol and percent ACTH variations were significantly and negatively influenced by dexamethasone levels, as well as by waist circumference values in men, and independently by BMI and waist circumference in women. However, in contrast to what has been found in men, a divergent contribution of BMI and waist circumference was found in women indicating that, with increasing waist values, a smaller suppression of the HPA axis was found with respect to that expected on the basis of BMI values. In conclusion, this study provides data of both physiological and physiopathological relevance. Overall, our data indicated that adjustment of the dexamethasone dose to body weight does not seem to substantially improve the sensitivity of the test, even in obese individuals, particularly when near-maximal doses are administered. However, this study demonstrated a highly significant effect of dexamethasone blood level concentrations on cortisol and ACTH suppression to low-dose dexamethasone tests. In addition, a significant effect of gender on postdexamethasone cortisol concentrations, suppression of the HPA axis, and dexamethasone levels were found, which may be dependent on related differences in both cortisol and dexamethasone metabolism. We showed that pituitary sensitivity to feedback inhibition by dexamethasone is preserved in obesity in both sexes even at low dosages. On the other hand, our data suggest that, at least in women, abdominal fat distribution may partially counteract the progressively greater suppressibility of the HPA axis that would be expected according to increasing BMI.

Abstract: OBJECTIVE: It has been reported that an increased availability of free fatty acids (NEFA) not only interferes with glucose utilization in insulin-dependent tissues, but may also result in an uncoupling effect of heart metabolism. We aimed therefore to investigate the effect of an increased availability of NEFA on gene expression of proteins involved in transmembrane fatty acid (FAT/CD36) and glucose (GLUT4) transport and of the uncoupling proteins UCP2 and 3 at the heart and skeletal muscle level. STUDY DESIGN: Euglycemic hyperinsulinemic clamp was performed after 24 h Intralipid(R) plus heparin or saline infusion in lean Zucker rats. Skeletal and heart muscle glucose utilization was calculated by 2-deoxy-[1-(3)H]-D-glucose technique. Quantification of FAT/CD36, GLUT4, UCP2 and UCP3 mRNAs was obtained by Northern blot analysis or RT-PCR. RESULTS: In Intralipid(R) plus heparin infused animals a significant decrease in insulin-mediated glucose uptake was observed both in the heart (22.62+/-2.04 vs 10.37+/-2.33 ng/mg/min; P<0.01) and in soleus muscle (13.46+/-1.53 vs 6.84+/-2.58 ng/mg/min; P<0.05). FAT/CD36 mRNA was significantly increased in skeletal muscle tissue (+117.4+/-16.3%, P<0.05), while no differences were found at the heart level in respect to saline infused rats. A clear decrease of GLUT4 mRNA was observed in both tissues. The 24 h infusion of fat emulsion resulted in a clear enhancement of UCP2 and UCP3 mRNA levels in the heart (99.5+/-15.3 and 80+/-4%) and in the skeletal muscle (291.5+/-24.7 and 146.9+/-12.7%). CONCLUSIONS: As a result of the increased availability of NEFA, FAT/CD36 gene expression increases in skeletal muscle, but not at the heart level. The augmented lipid fuel supply is responsible for the depression of insulin-mediated glucose transport and for the increase of UCP2 and 3 gene expression in both skeletal and heart muscle.

Abstract: AIMS/HYPOTHESIS: The aim of the present study was to investigate the relationship between intramyocytic triglycerides levels, muscle TNF-alpha and GLUT4 expression and insulin resistance. METHODS: Insulin sensitivity was studied in 14 severely obese women (BMI>40 kg/m(2)), before and 6 months after low-dietary intake or bariatric malabsorptive surgery (bilio-pancreatic diversion, BPD), by the euglycaemic hyperinsulinaemic clamp technique, while the amount of intramyocytic triglycerides was chemically measured in needle muscle biopsies. Using reverse transcriptase-polymerase chain reaction analysis, the muscle mRNA expression of TNF-alpha and GLUT4 was also investigated. RESULTS: The weight loss after surgery was 25.98+/-5.81 kg (P<0.001), while that obtained with the diet was 5.07+/-5.99 kg (P=NS). Marked decrease in TNF-alpha mRNA levels (76.67+/-12.59 to 14.01+/-5.21 AU, P<0.001) were observed in comparison with pre-treatment, whereas GLUT4 was significantly increased (62.25+/-11.77-124.25+/-21.01 AU, P<0.001) only in BPD patients. Increased glucose uptake (M) was accompanied by a significant decrease of TNF-alpha mRNA (76.67+/-12.59-14.01+/-5.21 AU, P<0.01) and an increase of GLUT4. The amounts of TNF-alpha mRNAs in skeletal muscle correlated inversely with GLUT4 mRNAs and directly with intramyocytic triglycerides levels. In a step-down regression analysis (r(2)=0.95) TNFalpha mRNA (P=0.0014), muscular TG levels (P=0.018), and GLUT4 mRNA (P=0.028) resulted to be the most powerful independent variables for predicting M values. CONCLUSION/INTERPRETATION: These findings suggest that insulin resistance in morbidly obese patients is positively associated to the intramyocytic triglycerides content and to TNF-alpha gene expression and inversely correlated to GLUT4 expression.

Abstract: OBJECTIVE: Obesity-related insulin resistance is closely associated with visceral fat accumulation. Several adipocyte-secreted molecules have been implicated in the development of type 2 diabetes, among them, the recently discovered adiponectin and resistin proteins. Some of these adipocytokines are also present in the immune system, thus suggesting an intriguing functional connection. RESEARCH METHODS AND PROCEDURES: We determined adiponectin and resistin expressions in visceral (VAT) and subcutaneous adipose tissue of lean and obese Zucker (fa/fa) rats using reverse-transcription polymerase chain reaction. Moreover, we analyzed the variations after body-weight reduction in food-restricted obese rats. RESULTS: Resistin and adiponectin expression was significantly lower in VAT of genetically obese in comparison with lean rats; no differences were observed when subcutaneous adipose tissues of the same animals were compared. Weight loss resulted in an increase of adiponectin expression in VAT, whereas a further significant decrease in resistin mRNA level was observed. Resistin is also present and equally expressed in splenocytes of lean and obese rats. DISCUSSION: Adiponectin and resistin are down-regulated in VAT of obese rats. Adiponectin expression is restored to normal levels after body-weight reduction, supporting its link with obesity-related insulin resistance. On the contrary, the further decrease of resistin mRNA after weight loss does not support the hypothesis that resistin may play a causative role in insulin resistance in obese rats. Moreover, we demonstrated the presence of resistin in immunocompetent cells in both humans and rats, thus adding another factor to the list of molecules that adipose tissue shares with the immune system.

Abstract: The dual center hypothesis in the central control of energy balance originates from the first observations performed more than 5 decades ago with brain lesioning and stimulation experiments. On the basis of these studies the "satiety center" was located in the ventromedial hypothalamic nucleus, since lesions of this region caused overfeeding and excessive weight gain, while its electrical stimulation suppressed eating. On the contrary, lesioning or stimulation of the lateral hypothalamus elicited the opposite set of responses, thus leading to the conclusion that this area represented the "feeding center". The subsequent expansion of our knowledge of specific neuronal subpopulations involved in energy homeostasis has replaced the notion of specific "centers" controlling energy balance with that of discrete neuronal pathways fully integrated in a more complex neuronal network. The advancement of our knowledge on the anatomical structure and the function of the hypothalamic regions reveals the great complexity of this system. Given the aim of this review, we will focus on the major structures involved in the control of energy balance.

Abstract: OBJECTIVE: To investigate whether intima-media thickness (IMT) of the common carotid artery (CCA), an early marker of asymptomatic atherosclerosis, is significantly and independently associated with plasma concentrations of leptin, an adipose tissue hormone that has recently been proposed as a cardiovascular risk factor in obese patients. DESIGN: Cross-sectional sample of normal-weight and obese men and women. SUBJECTS: One-hundred and twenty healthy subjects (52 men and 68 women), aged 18-45 y and with a wide range of BMI, were recruited for the study. MEASUREMENTS: Fasting plasma leptin concentrations and the IMT of the CCA were measured in all subjects. Leptin concentrations were measured by radioimmunoassay and the IMT of the CCA was quantified by high resolution B-mode ultrasound imaging. Central fat (measured by waist circumference), smoking habits, blood pressure, insulin sensitivity (measured by the insulin tolerance test), and fasting plasma glucose, insulin and lipid pattern (cholesterol, HDL-cholesterol, triglycerides, LDL-cholesterol) were also measured. RESULTS: IMT of the CCA was positively correlated with log leptin concentrations (P<0.005 in men and P<0.001 in women), body mass index (P<0.001 in men and women), waist circumference (P<0.001 in men and women), age (P<0.001 in men and P<0.05 in women), and negatively associated with insulin sensitivity in both sexes (P<0.05). IMT was also directly correlated with cholesterol (P<0.05), LDL-cholesterol (P<0.01) and systolic blood pressure in men (P<0.05), and with diastolic blood pressure levels in women (P<0.05). When a multiple linear regression model was used without body mass index (BMI), the correlation between leptin and IMT was maintained in both men (P<0.01) and women (P<0.005), independent of age, insulin sensitivity, smoking habits, systolic blood pressure, fasting glucose, triglycerides, cholesterol, LDL-cholesterol and HDL-cholesterol. By contrast, BMI-adjusted leptin concentrations were not significantly associated with IMT (Pc (partial correlation): 0.41 in men and 0.15 in women). Moreover, when BMI was entered into a multiple linear regression model without leptin, the correlation between BMI and IMT was maintained in both men (P<0.005) and women (P<0.01), independent of the same parameters. CONCLUSION: Plasma leptin concentrations are independently associated with the IMT of the CCA, suggesting that the increase of adipose tissue mass (or leptin per se) may have an unfavourable influence on the development of atherosclerosis. However, the association between IMT and leptin seems to be dependent and/or confounded by the relationship between IMT and obesity.

Abstract: There is currently a lively debate involving scholars from diverse cultural background on the subject of evidence-based medicine. In order to set out the grounds of discussion this paper deals with the problem of "evidence", both the concept and meaning of this term. After a brief history of the idea of "evidence" from antiquity to the present day, it will be underlined how modern day thinking recognizes the existence of two types of "evidence": rational evidence supported first by René Descartes, and perceptible evidence, defended by Francis Bacon. In contemporary epistemology the idea of "evidence" has been conceived above all as perceptible and is closely linked to the idea of fact. The concept of "scientific fact" will therefore be analyzed and will reveal how in science, contrary to prevailing opinion, "pure facts" practically do not exist and how the "facts" which scientists talk about in their research are always "facts depending on theory". Subsequently the capacity of "facts" to prove scientific hypothesis will be examined. In the light of more recent epistemological reflection the thesis will be maintained according to which facts are not capable of definitively proving the truth of a theory. Such reflection leads to the conclusion that in medicine "evidence" is always dependent on the theoretical and practical context in which researchers work. In the final part of the paper the epistemological presumptions and ambitions of evidence-based medicine will be examined. This analysis concludes that the epistemology on which evidence-based medicine relies upon does not grasp the true complexity of the scientific methodology and can weaken in doctors the beneficial stimulus that is doubt.

Abstract: Neuropeptide Y (NPY) participates in the regulation of reproduction and food intake. The adipose-secreted hormone, leptin, has also been involved in these processes, and has been shown to exert its effects in part by controlling NPY synthesis and release at the hypothalamic level. In the present study, we utilized the SH-SY5Y human neuroblastoma cell line, to study the leptin-NPY interrelationships. SH-SY5Y cells were found to express leptin receptors (RT-PCR and Western blot analyses). A 24-h treatment with leptin at different concentrations did not affect NPY gene expression, but resulted in a stimulation of NPY release. This stimulated secretion was blocked by the combined treatment with leptin and the muscarinic agonist carbachol or the phorbol ester TPA. Leptin and carbachol also caused an increased intracellular content of NPY. In conclusion, the SH-SY5Y human neuroblastoma cell line appears to be a suitable in vitro model for studying the pharmacological effects of leptin on the biosynthesis and secretion of NPY.

Abstract: The preferential channeling of different fuels to fat and changes in the transcription profile of adipose tissue and skeletal muscle are poorly understood processes involved in the pathogenesis of obesity and insulin resistance. Carbohydrate and lipid metabolism may play relevant roles in this context. Freely moving lean Zucker rats received 3- and 24-h infusions of Intralipid (Pharmacia and Upjohn, Milan, Italy) plus heparin, or saline plus heparin, to evaluate how an increase in free fatty acids (nonesterified fatty acid [NEFA]) modulates fat tissue and skeletal muscle gene expression and thus influences fuel partitioning. Glucose uptake was determined in various tissues at the end of the infusion period by means of the 2-deoxy-[1-3H]-D-glucose technique after a euglycemic-hyperinsulinemic clamp: high NEFA levels markedly decreased insulin-mediated glucose uptake in red fiber-type muscles but enhanced glucose utilization in visceral fat. Using reverse transcriptase-polymerase chain reaction and Northern blotting analyses, the mRNA expression of fatty acid translocase (FAT)/CD36, GLUT4, tumor necrosis factor (TNF)-alpha, peroxisome proliferator-activated receptor (PPAR)-gamma, leptin, uncoupling protein (UCP)-2, and UCP-3 was investigated in different fat depots and skeletal muscles before and after the study infusions. GLUT4 mRNA levels significantly decreased (by approximately 25%) in red fiber-type muscle (soleus) and increased (by approximately 45%) in visceral adipose tissue. Furthermore, there were marked increases in FAT/CD36, TNF-alpha, PPAR-gamma, leptin, UCP2, and UCP3 mRNA levels in the visceral fat and muscle of the treated animals in comparison with those measured in the saline-treated animals. These data suggest that the in vivo gene expression of FAT/CD36, GLUT4, TNF-alpha, PPAR-gamma, leptin, UCP2, and UCP3 in visceral fat and red fiber-type muscle are differently regulated by circulating lipids and that selective insulin resistance seems to favor, at least in part, a prevention of fat accumulation in tissues not primarily destined for fat storage, thus contributing to increased adiposity and the development of a prediabetic syndrome.

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Abstract: The methodological aspects of making a differential diagnosis have so far received little attention. The aim of the present study is therefore to analyze the logical structure of this phase of clinical procedure. It is commonly believed that making a differential diagnosis involves an inferential process through which the clinician first makes diagnostic hypotheses, rules out all these prospective hypotheses except for one, and then concludes that the one remaining hypothesis identifies the disease affecting the patient. Some authoritative scientific methodologists sustain that the logical mechanism leading to the rejection of a diagnostic hypothesis is the negation of the consequent, and is thus the falsification proposed by Popper as the fundamental inference in experimental science. In the present study it is therefore suggested that in making a differential diagnosis, clinicians utilize disjoined syllogism rather than a negation of the consequent. A description is given of the structures of the two types of disjoined syllogism, which are inclusive and exclusive. An in-depth analysis is made of the entire diagnostic-differential reasoning process, and the authors suggest that, on a logical plane, it consists of two distinct sequential inferences, one being an eliminative induction and the other an inclusive disjoined syllogism. Lastly, it is pointed out that, as in the clinic, perfect pathognomonic signs are rarely encountered, each inference leading to a rejection of a diagnostic hypothesis can never be apodictic, and can only lead to a probable conclusion. The confutation of diagnostic hypotheses thus appears to be based on the rule of the denial of inductive logic.

Abstract: As PAI-1, a cardiovascular risk factor linked to insulin-resistance, may be influenced by a 4G/5G gene polymorphism in disease states, we studied both PAI-1 plasma concentration (PAI-1:Ag) and 4G/5G polymorphism, and their relationship with anthropometric and endocrinemetabolic parameters in 93 obese patients and 79 lean normal subjects. In obese patients PAI-1:Ag levels were significantly increased, namely in males and in those with central obesity, and tightly related to the insulin-resistance parameters. In obese patients the 4G/5G polymorphism was a determinant of PAI-1:Ag levels, which were highest in 4G/4G, intermediate in 4G/5G and lowest in 5G/5G genotype carriers. PAI-1:Ag levels were significantly associated with most of anthropometric and endocrine-metabolic parameters only in 4G allele obese carriers. Moreover, only in patients with central obesity was the relationship between genotype and PAI-1 concentration maintained, with the highest levels in the 4G/4G patients. In each genotype subset of patients with central, but not peripheral, obesity PAI-1:Ag levels were significantly increased compared to their lean counterparts. In conclusion, the 4G/5G polymorphism may influence PAI-1 expression in obesity, with a crucial role in central but not peripheral adiposity. Since subjects with central obesity are at high risk for cardiovascular disease, the effects of the 4G/5G polymorphism on PAI-1 concentration may further enhance this risk.

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Abstract: Little is known about the mechanisms involved in the preferential channeling of different fuels to fat and how the target tissue participates in this process. Dietary fatty acids have been shown to act as signaling molecules that bind and activate a new class of nuclear receptors, the peroxisome proliferator-activated receptors (PPARs). PPAR-gamma is particularly interesting because it may have the potential to link particular fatty acids with a program of gene expression involved in lipid storage and metabolism. We investigated whether a nutrient-sensing pathway is activated by an increased availability of lipid fuels in nine normal weight male volunteers. Using reverse transcriptase-polymerase chain reaction analysis, the mRNA expression of fatty acid translocase (FAT)/CD36, PPAR-gamma2, leptin, uncoupling protein (UCP)-2 and UCP-3, and tumor necrosis factor (TNF)-alpha was investigated in gluteal subcutaneous fat biopsies before and after 5 h infusions of saline or Intralipid (Pharmacia and Upjohn, Milan, Italy) plus heparin, which does not modify insulinemia. Marked increases in FAT/CD36 (724+/-18%; P < 0.05), PPAR-gamma2 (200+/-8%; P < 0.05), leptin (110+/-13%; P < 0.05), UCP-2 (120+/-7%; P < 0.05), UCP-3 (80+/-5%; P < 0.05), and TNF-alpha mRNA (130+/-12%; P < 0.05) were observed in comparison with pretreatment levels, whereas there was no change after saline infusion. These data suggest that the in vivo gene expression of FAT/CD36, PPAR-gamma2, leptin, UCP-2, UCP-3, and TNF-alpha in subcutaneous adipose tissue is regulated by circulating lipids independent of insulin and that prolonged hyperlipidemia may therefore contribute to increased fat metabolism and storage as a result of the increased expression of these proteins.

Abstract: The authors examine the problem of defining alternative medicine, and after a brief analysis conclude that a satisfactory unifying definition of the different practices is not possible. Scientific knowledge is a function of scientific method. In turn the principle of falsifiability proposed by Karl Popper is used as a demarcation line between science and pseudoscience. They assert that the various alternative modalities do not represent authentic scientific disciplines, as they lack many of the minimum requirements of scientific discourse and, above all, because they violate the principle of falsifiability. Until they overcome these methodological shortcomings, alternative medical practices cannot become authentic scientific disciplines.

Abstract: Increased basal plasma FFA and lactate concentrations are often present in obesity and may deeply affect insulin action. The inhibition of glucose transport or phosphorylation is thought to be involved in this phenomenon, but the molecular mechanisms on the basis are still unknown. In our laboratory we observed that a chronic infusion of Intralipid plus heparin in rats significantly decreased the insulin dependent-glucose uptake, as well as GLUT4 gene expression in muscular tissue. On the other hand it has been shown that an enhanced plasma lactate concentration may increase insulin secretion and hepatic insulin clearance. Moreover we observed that chronic hyperlactatemia in rats is able to decrease glucose uptake in muscles, while reducing GLUT4 mRNA and protein in the same tissues. In obesity, lactate and FFA overproduction from visceral fat may therefore play a synergic role in reducing insulin sensitivity.

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Abstract: Leptin is secreted by adipocytes and regulates food intake and energy balance through the activation of specific receptors (OB-R). Recent evidence suggests that it is also involved in the control of reproductive processes, by possibly acting on central and peripheral targets. In particular, it has been shown that leptin may indirectly stimulate GnRH release from hypothalamic fragments by acting on interneurons impinging on GnRH-secreting neurons. The possibility that leptin might additionally modulate the activity of GnRH-secreting neurons in a direct way has been addressed in the present study, by using the immortalized GnRH-secreting cell line GT1-7. The presence of OB-R messenger RNA (mRNA) (long form) was detected by RT-PCR analysis of total RNA from GT1-7 cells. An OB-R protein is also expressed in these cells, as shown by immunocytochemistry and by Western blot analysis. The latter has revealed the presence of a single immunoreactive OB-R with an approximate size of 130 kDa. To study the functionality of these receptors, the effect of leptin treatment on GnRH secretion and gene expression in GT1-7 cells were evaluated. Under static conditions, GnRH release was stimulated by exposure to low concentrations of leptin (10(-12) M after 30 min; 10(-10) M after 60 min). The 10(-12) M dose was selected for studying the effect of leptin on GnRH secretion under dynamic conditions. To this purpose, GT1-7 cells were placed in a perifusion system; treatment with leptin (10(-12) M) for 60 min stimulated GnRH release with no changes of pulse frequency. On the contrary, exposure to leptin (10(-12)-10(-10) M) for 1, 3, 6, and 24 h did not affect GnRH gene expression in GT1-7 cells. The present results indicate that GT1-7 cells possess OB-Rs and that leptin may directly affect their function. Taken together with the available reports, these findings suggest that leptin might participate in the regulation of reproductive processes by acting at multiple levels, both centrally and peripherally.

Abstract: After elucidating the controversy that accompanied the birth of laboratory medicine, the Authors define and examine the concepts underlying clinical methodology. The mental operations of the clinician are analyzed, a distinction being made between the processes of: a) "categorization" of the patient's disease, and b) "explanation" for pathologic phenomena. The diagnostic procedure is usually based above all on categorization. In the first phase of this procedure the clinician searches for data that are of the greatest possible "informative value", and then goes works out a certain number of "syndromic complexes". In the second phase, on the basis of these "groups of signs", the physician must formulate a certain number of diagnostic hypotheses and evaluate the probability of the presence of a particular disease, in view of the presence of particular signs. Finally, two fundamental arguments of clinician are dealt with: "the confirmatory argument" and "the falsifying argument", and the value of these two inferences in conferring certainty or a certain grade of reliability on clinical judgement is analyzed and discussed.

Abstract: Mechanisms of regulation of plasma leptin in lean and genetically obese animals are not completely understood. In particular a relation has been proposed between energy metabolism and leptin. However, it is not clear how energy expenditure and leptin are related under exercise in lean and obese animals. To clarify these aspects we investigated lean and genetically obese (fa/fa) Zucker rats undergoing a single bout (30 min) of swimming and measured several biochemical and hormonal parameters of energy metabolism and leptin changes throughout the study. Moreover ob-gene expression in adipose tissue was also measured. Our results showed that plasma leptin is decreased by 30% at the end of exercise in lean animals while resulting unaffected in obese animals. Leptin changes in lean rats are concomitant with the peak of NEFA and glycerol release from adipose tissue rather than with the reduction of plasma insulin. Ob-gene expression in adipose tissue was markedly increased in fa/fa compared to lean rats, but was not modified by exercise both in lean and obese animals. In conclusion our data show that leptin changes during exercise are related to lipolytic events in adipose tissue and support a link between leptin and energy expenditure.

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Abstract: Leptin, the satiety hormone expressed almost exclusively in adipose tissue, is a marker of body fat accumulation in humans. Recent studies have shown that plasminogen activator inhibitor-1 (PAI-1), a prothrombotic factor associated with atherosclerosis complications, is also produced in adipose tissue. The objective of the present study was to determine whether PAI-1 antigen plasma concentrations are associated with leptin plasma levels or the body fat mass (FM) independently of the variables known to influence PAI-1 production. Sixty-one nondiabetic women aged 18 to 45 years with a wide range of values for the body mass index ([BMI] 18.1 to 37.7 kg/m2) were evaluated for (1) body FM and fasting plasma levels of (2) PAI-1 antigen, (3) PAI-1 activity, (4) leptin, (5) insulin, (6) blood glucose, and (7) lipids (cholesterol, high-density lipoprotein [HDL]-cholesterol, and triglycerides [TG]). Body FM and fat-free mass (FFM) were estimated during fasting conditions by the bioimpedance analysis (BIA) method using a tetrapolar device. Body fat distribution was evaluated by the waist circumference and the waist to hip ratio (WHR). FM was directly associated with both PAI-1 antigen (r = .585, P < .001) and PAI-1 activity (r = .339, P < .001). Seemingly, leptin was positively related to both PAI-1 antigen (r = .630, P < .001) and PAI-1 activity (r = .497, P < .001). Moreover, both PAI-I antigen and PAI-1 activity were directly correlated with FFM (r = .285, P < .05, and r = .336, P < .01, respectively), BMI (r = .594, P < .001, and r = .458, P < .001, respectively), and WHR (r = .510, P < .001, and r = .391, P < .005, respectively). Insulin was directly related to PAI-1 antigen (r = .540, P < .001), PAI-1 activity (r = .259, P < .05), leptin (r = .447, P < .001), and FM (r = .435, P < .001). The association between PAI-1 antigen (dependent variable) and leptin or FM was tested by a stepwise regression model simultaneously including leptin, FM, BMI, WHR, age, FFM, and fasting insulin, blood glucose, TG, cholesterol, and HDL-cholesterol as independent variables. PAI-1 antigen maintained a significant positive independent relationship only with leptin (t = 2.923, P < .01), insulin (t = 3.489, P < .001), and fasting blood glucose (t = 2.092, P < .05), and a negative independent relationship with HDL-cholesterol (t = -2.634, P < .05). In conclusion, the strong relationship between PAI-1 antigen and leptin irrespective of other variables known to influence these factors seems to indicate that leptin per se may potentially increase PAI-1 plasma concentrations in obese subjects.

Abstract: The paper discusses the epistemological question of pain and the way this concept forms part of scientific knowledge. The first part gives a rapid overview of the main aspects of modern scientific knowledge. In earlier times Bacon, Galileo and Descartes felt that the new form of knowledge could head to a knowledge of truth and certainty. Today, these ideals have proved unattainable and science only takes the form of well founded, rigorous and objective knowledge. Moreover, the objectivity of science is based on intersubjectivity. The second part of the paper examines pain and underlines that it is a subjective phenomenon which cannot be ascertained by the doctor in the same way as commonplace anatomic and physiological phenomena. Our scientific knowledge of pain is based on operations for which it is possible to achieve an intersubjective knowledge: these operations constitute the protocol criteria on which it is possible to base a scientific knowledge of pain. In the last part of this paper, having stressed that pain is a concept belonging to many disciplines--scientific, psychological, philosophical, religious--the emphasis is placed on pain itself as opposed to suffering. While the former is a concept that belongs to scientific medicine, the latter belongs to that area of medicine which lies beyond the boundaries of science and concerns the activity of a man who helps a fellow human who is suffering.

Abstract: An increased basal plasma lactate concentration is present in many physiological and pathological conditions, including obesity and diabetes. We previously demonstrated that acute lactate infusion in rats produced a decrease in overall glucose uptake. The present study was carried out to further investigate the effect of lactate on glucose transport and utilization in skeletal muscle. In chronically catheterized rats, a 24-h sodium lactate or bicarbonate infusion was performed. To study glucose uptake in muscle, a bolus of 2-deoxy-[3H]glucose was injected in basal condition and during euglycemic-hyperinsulinemic clamp. Our results show that hyperlactatemia decreased glucose uptake in muscles (i.e., red quadriceps; P < 0.05). Moreover in red muscles, both GLUT-4 mRNA (-30% in red quadriceps and -60% in soleus; P < 0.025) and protein (-40% in red quadriceps; P < 0.05) were decreased, whereas the (E1alpha)pyruvate dehydrogenase (PDH) mRNA was increased (+40% in red quadriceps; P < 0.001) in lactate-infused animals. PDH protein was also increased (4-fold in red gastrocnemius and 2-fold in red quadriceps). These results indicate that chronic hyperlactatemia reduces glucose uptake by affecting the expression of genes involved in glucose metabolism in muscle, suggesting a role for lactate in the development of insulin resistance.

Abstract: AIMS:To verify if plasma leptin concentrations of newborns at birth differ significantly between sexes; and to investigate the potential interactions between plasma leptin and growth-regulating hormones at birth. SUBJECTS: 98 healthy newborns (48 male, 50 female) were studied. Leptin, insulin, cortisol, insulin-like growth factor-1 (IGF-1), testosterone, and sex hormone binding globulin (SHBG) concentrations were measured from venous blood collected from the umbilical cord vein immediately after birth. RESULTS: The serum leptin concentration of newborns averaged 8.05(0.5) ng/ml. Females had significantly (P<0.005) higher serum leptin values than males [9. 6(0.8) vs 6.0(0.6) ng/ml]. IGF-1 was significantly (P<0.05) higher in females than in males [87(4) vs 74(5) microg/l], whereas SHBG was slightly lower [29(1) vs 33(2) nmol/l]. Insulin, cortisol, and testosterone serum concentrations were not statistically different between the sexes. Among the variables examined, birth weight (expressed as Z-score of weight) and insulin showed the highest degree of relationship with serum leptin in newborns (r=0.48 and r=0.31 respectively, P<0.001). Multiple regression analysis showed that Z-score of birth weight, gender and cortisol were able to account for approximately 44% of inter-individual variability of serum leptin concentrations in newborns. CONCLUSIONS: Female newborns have significantly higher serum leptin concentrations than males. Insulin, IGF-1, testosterone, and SHBG did not independently affect leptin inter-individual variability when gender, Z-score of body weight, and cortisol were taken into account. Other factors may be involved in the differences in circulating leptin concentrations between the sexes in newborns.

Abstract: The aim of this study is to explain homeopathy, and make a critical evaluation of the doctrine and its clinical practice. The discipline of homeopathy, a medical doctrine advanced by Samuel Hahnemann in the late eighteenth and the early nineteenth centuries, is based on a physiological theory according to which a "vital force dominates the human body in an unopposed and dynamic way"; disease processes are considered the consequence of a disturbance of this vital force. According to this doctrine, there are only three diseases: psora, sycosis and lues. Drugs are believed to act according to the principle of similitude, their efficacy increasing with their progressive dilution. Several schools of homeopathy have emerged since Hahnemann's death. Although they differ from each other, they share two of the fundamental principles advanced by the founder. Homeopathic clinical teaching aims at identifying the greatest number of signs present in the patient and then, on the basis of these signs, and irrespective of any diagnosis, the homeopathic specialist decides what drug(s) should be administered. Homeopathy is a doctrine that can be rationally criticized from three standpoints. First, its content contrasts radically with current scientific knowledge of chemistry, pharmacology, and pathology. Second, despite the fact that homeopathic specialists claim many therapeutic successes, the small number of rigorous studies conducted have not as yet provided convincing evidence that homeopathic treatment is effective against particular disease processes. Third, from a methodological standpoint, homeopathy has a number of serious flaws: above all, it violates both the principle of falsifiability enunciated by Karl Popper as a criterion for the demarcation between science and pseudo-science, and the principle of operative definition. Homeopathy cannot therefore be considered a scientific discipline.

Abstract: Abnormalities of the hypothalamus-pituitary-adrenal axis and hypersensitivity to corticosteroids have been suggested as major determinants of the development of visceral obesity. Since at the cellular level most effects of corticosteroids are mediated by specific receptors, we evaluated the number of type I and type II corticosteroid receptors in mononuclear leucocytes of 26 obese and 13 control subjects. We also studied the relationship between corticosteroid receptors, measured by radioreceptor assay, and abdominal visceral fat, evaluated by computed tomography scan, plasma and urine corticosteroid hormone concentrations and overall glucose metabolism, assessed by euglycaemic-hyperinsulinaemic clamp. We observed a decrease in type II receptors in the obese subjects (1746 +/- 160 vs 2829 +/- 201 per cell; P < 0.0001), with no change in type I receptors. Type II receptors decreased in relation to body mass index (r = -0.53; P < 0.005) and total glucose disposal (r = 0.51; P < 0.01). Abdominal visceral fat did not correlate with type II receptor number, but did correlate with total glucose disposal (r = -0.35; P < 0.05); the rate of glucose disposal was lower in obese subjects (3.3 +/- 0.3 vs 7.4 +/- 0.4 mg/kg per min; P < 0.001). Plasma and urine cortisol did not differ between the two groups. However, a direct correlation between type II receptor number and both plasma (r = 0.43; P < 0.02) and urine cortisol concentrations (r = 0.60; P < 0.05) was observed. In conclusion, the number of type II corticosteroid receptors in mononuclear leucocytes was found to be lower in obese subjects. This abnormality appears to be related to the degree of adiposity and to the main endocrine-metabolic features of the obesity syndrome, further supporting the hypothesis of involvement of hypothalamus-pituitary-adrenal axis hyperactivity in the pathophysiology of obesity.

Abstract: Leptin is a hormone produced in the adipose tissue and its concentrations in peripheral blood are significantly correlated with the amount of body fat. Whether other factors, including the pattern of body fat distribution and several hormones (such as insulin, sex steroids, and glucocorticoids), may be involved in the regulation of circulating blood leptin levels is controversial. Women with the polycystic ovary syndrome (PCOS) are hyperandrogenic and most of them are characterized by hyperinsulinemia, insulin resistance, and obesity, particularly the visceral phenotype. To assess the potential contribution of anthropometric factors, androgens, and insulin in determining leptin levels, we examined their relationship with body-mass index (BMI), visceral (VAT) and subcutaneous (SAT) adipose tissue areas, basal androgen levels, and fasting and glucose-stimulated (AUC) insulin in different groups of obese women with PCOS (n = 23) and of age-matched obese (n = 16) and non-obese (n = 10) otherwise healthy controls. The VAT/SAT ratio was measured as a parameter of body fat distribution. Serum leptin levels were significantly higher in obese PCOS women than in obese and normal-weight healthy controls and, within the controls, in the obese than in the non-obese group. In all women considered together, and in each group separately, leptin concentrations were highly significantly correlated with BMI. In addition, after adjusting for BMI, both VAT and the VAT/SAT ratio were positively and significantly correlated with leptin. Partial correlations with the VAT/SAT ratio remained significant in both the obese PCOS group and in controls considered separately, whereas the correlation with the SAT value was significant only in the control group. After adjusting for BMI, no correlation between leptin, androgens and fasting or stimulated (like AUC) insulin was found. These findings indicate that leptin levels in obese women with PCOS are higher than those observed in obese and non-obese controls. Moreover, they suggest that, other than BMI, the pattern of body fat distribution may be an independent factor related to circulating leptin levels, which, on the contrary, do not appear to be related to either androgen or insulin concentrations.

Abstract: Intracerebroventricular administration of neuropeptide Y to normal rats induces a syndrome characterised by obesity, hyperinsulinaemia, insulin resistance and over expression of the adipose tissue ob gene. Little is known about the effect of circulating neuropeptide Y on glucose metabolism, insulin secretion and leptin. We therefore aimed to evaluate the effect of an intravenous infusion of neuropeptide Y on glucose disposal, endogenous glucose production, whole body glycolytic flux, and glucose storage as assessed during euglycaemic hyperinsulinaemic clamp. In addition, the insulin-stimulated glucose utilisation index in individual tissues was measured by the 2-deoxy-[1-3H]-glucose technique. The effect of neuropeptide Y on insulin secretion was evaluated by hyperglycaemic clamp. Infusion did not induce any change in endogenous glucose production during basal conditions or at the end of the clamp. Glucose disposal was significantly increased in the rats given neuropeptide Y compared with controls (27.8 +/- 1.3 vs 24.3 +/- 1.6 mg x min(-1) x kg(-1); p < 0.05) as was the glycolytic flux (18.9 +/- 1.6 vs 14.4 +/- 0.8 mg x min(-1) x kg(-1); p < 0.05), while glucose storage was comparable in the two groups. In skeletal muscle, the glucose utilisation index was increased significantly in rats given neuropeptide Y. The glucose utilisation index in subcutaneous and epididimal adipose tissue was not significantly different between the two groups. Plasma leptin was significantly increased by hyperinsulinaemia, but was not affected by neuropeptide Y infusion. Both the early and late phase of the insulin response to hyperglycaemia were significantly reduced by neuropeptide Y. In conclusion neuropeptide Y infusion may increase insulin-induced glucose disposal in normal rats, accelerating its utilisation through the glycolytic pathway. Neuropeptide Y reduces both phases of the insulin response to hyperglycaemia.

Abstract: Plasma lactate is elevated in many physiological and pathological conditions, such as physical exercise, obesity, and diabetes, in which a reduction of insulin sensitivity is also present. Furthermore, an increased production of lactate from muscle and adipose tissue together with increased gluconeogenic substrate flux to the liver plays a primary role in enhancing hepatic glucose production (HGP) in diabetes. It has been shown that lactate may interfere with the utilization and oxidation of other substrates such as free fatty acids (FFAs). The aim of this study was to investigate if lactate infusion affects peripheral glucose utilization in rats. Animals were acutely infused with lactate to achieve a final lactate concentration of 4 mmol/L. They were then submitted to a euglycemic-hyperinsulinemic clamp to study HGP and overall glucose metabolism (rate of disappearance [Rd]). At the end of the clamp, a bolus of 2-deoxy-[1-3H]-glucose was injected to study insulin-dependent glucose uptake in different tissues. The results show that lactate infusion did not affect HGP either in the basal state or at the end of clamp, whereas glucose utilization significantly decreased in lactate-infused rats (26.6 +/- 1.1 v 19.5 +/- 1.4 mg.kg-1.min-1, P < .01). A reduction in the tissue glucose utilization index was noted in heart (18.01 +/- 4.44 v 46.21 +/- 6.51 ng.mg-1.min-1, P < .01), diaphragm (5.56 +/- 0.74 v 9.01 +/- 0.93 ng.mg-1.min-1, P < .01), soleus (13.62 +/- 2.29 v 34.05 +/- 6.08 ng.mg-1.min-1, P < .01), and red quadricep (4.43 +/- 0.73 v 5.88 +/- 0.32 ng.mg-1.min-1, P < .05) muscle in lactate-infused animals, whereas no alterations were observed in other muscles or in adipose tissue. Therefore, we suggest that acute lactate infusion induces insulin resistance in the heart and some muscles, thus supporting a role for lactate in the regulation of peripheral glucose metabolism.

Abstract: In a previous study performed in adult obese and normal-weight male subjects, we found that suppression of insulin levels by diazoxide reduced testosterone and increased sex hormone-binding globulin (SHBG) blood concentrations. These and other data suggested that insulin may have a regulatory capacity in testosterone secretion and/or metabolism in men, similar to what has already been demonstrated in women. In this study, we investigated the effects of acute hyperinsulinemia on major androgen levels, including testosterone, in two groups of normal-weight in = 11) and obese (n = 9) men. Acute hyperinsulinemia was obtained by the euglycemic-hyperinsulinemic clamp technique. Relationships between the degree of insulin resistance (ie, total glucose disposal [M value]) and testosterone levels were also evaluated. Basal testosterone levels in obese subjects (10.40 +/- 3.02 nmol/L) were significantly lower than in normal-weight controls (15.50 +/- 4.65 nmol/L, P < .01), whereas no difference was present in androstenedione and dehydroepiandrosterone sulfate (DHEA-S) concentrations. During the clamp study, testosterone was significantly increased in the obese group (11.79 +/- 3.64 nmol/L, P < .05) but not in the control group (15.81 +/- 4.54 nmol/L, P = NS). The other two androgens did not significantly change in either the obese or control group. There was a highly significant correlation between baseline testosterone concentrations, with M values suggesting a relationship between impaired peripheral insulin sensitivity and reduced plasma testosterone concentrations. It should be pointed out that there was a certain discrepancy in the testosterone variations, particularly in the control group, in which two thirds of the subjects had no change or some decrease in testosterone levels, whereas in the remainder testosterone increased over the values of the assay variation coefficient. These findings are consistent with the hypothesis that insulin may regulate testosterone blood levels also in male subjects. Whether these effects are primarily due to increased hormone secretion or reduced clearance needs to be investigated.

Abstract: OBJECTIVES: To evaluate the plasma leptin concentration in lean and genetically obese fa/fa rats and to assess the response to 2 h hyperinsulinaemia. BACKGROUND: The recently discovered peptide leptin is a putative link between the size of the adipose mass and the hypothalamic centres controlling feeding behaviour. Several genetic models of animal obesity have been characterized as carriers of mutations of either the ob gene or leptin receptor. EXPERIMENTAL DESIGN: Lean (+/?) and obese (fa/fa) Zucker rats were studied under pentobarbital anaesthesia and underwent a 2 h euglycaemic hyperinsulinaemic clamp. Plasma leptin was measured in basal condition and at the end of the clamp study. Glucose rate of disappearance was evaluated by means of the isotope dilution technique using 3-3H-glucose as tracer. RESULTS: fa/fa rats showed a 40 fold higher leptin concentration compared to lean littermates (0.47 +/- 0.10 vs 19.55 +/- 1.50 ng/ml, P < 0.0001). Euglycaemic hyperinsulinaemia increased plasma leptin in lean but not in genetically obese rats. CONCLUSIONS: Our results suggest that insulin may be a regulator of in vivo leptin secretion by adipose tissue of lean rates whereas it is ineffective in increasing plasma leptin in obese Zucker rats.

Abstract: It has been suggested that the extrinsic coagulation system plays a crucial role in the initiation of blood coagulation in atherosclerotic disease and that TFPI, the inhibitor of the factor VIIa/tissue factor complex, bound to the endothelial cells, could prevent in vivo blood clotting. Because obesity has a role in the pathogenesis of atherosclerotic cardiovascular disease, we measured TFPI antigen plasma levels (ng/mL) by ELISA at baseline and 5 minutes after an IV bolus of 20 IU/kg body weight of unfractionated commercial mucous heparin in 12 obese patients with a mean body mass index (BMI) of 41.4 +/- 1.4 kg/m2 and 14 normal-weight control subjects (BMI 23.1 +/- 1.3 kg/m2). All subjects were submitted to an OGTT. The obese patients displayed a normal glucose tolerance. However, they had a different glucose-induced hyperinsulinemia (14.9 +/- 2.0 versus 7.8 +/- 0.8 mU/L, p < 0.01). Total serum cholesterol did not differ between controls and obese patients, whereas plasma triglycerides were higher in the latter group. Basal TFPI antigen plasma levels were similar in obese and controls (83.8 +/- 5.0 versus 77.7 +/- 3.5 ng/mL, p = N.S.). In contrast, after heparin a significantly lower rise in TFPI antigen plasma levels was observed in obese patients (511.2 +/- 43.4 ng/mL) (p < 0.003). Moreover, a significant inverse correlation was found between the heparin-stimulated TFPI antigen plasma levels and both BMI and basal plasma insulin concentrations. Thus, the link between insulin level and endothelial cell-associated TFPI could at least partially explain why obese patients are more prone to develop cardiovascular disorders.

Abstract: In obesity, a central pattern of fat distribution is mostly associated with hyperinsulinemia, insulin resistance, and hyperlipemia, thus promoting the development of non-insulin-dependent diabetes mellitus and cardiovascular disease. In addition, in obesity, changes in energy expenditure are hypothesized to be involved in the development or maintenance of excessive body fat storage. In this study, abdominal fat distribution by computed tomographic (CT) scan was used to study the relation between the visceral fat depot, insulin secretion, and insulin sensitivity in a group of obese subjects with normal glucose tolerance (n = 26; body mass index [BMI], 39 +/- 1 kg/m2) and a group of normal-weight control subjects (n = 9; BMI, 23 +/- 1 kg/m2). The minimal model method was used to assess insulin sensitivity, S(I), and first-phase (phi1) and second-phase (phi2) beta-cell sensitivity from plasma glucose, insulin, and C-peptide concentrations measured during an intravenous glucose tolerance test ([IVGTT] 0.33 g/kg body weight). Moreover, we evaluated the relationships between these parameters and the resting metabolic rate (RMR) and glucose-induced thermogenesis (GIT) measured by indirect calorimetry. The data show the following: (1) in obese subjects, phi1 is greater but not statistically different from the value in control subjects (252 +/- 41 v 157 +/- 25 dimensionless 10(9)); (2) phi2 is significantly higher in obese subjects (27 +/- 4 v 14 +/- 2 min(-1) x 10(9), P < .05), with a positive correlation between the amount of visceral adipose tissue (VAT) and phi2 (r = .49, P < .05); (3) S(I) is decreased in the obese group (2.8 +/- 0.3 v 9.7 +/- 1.6 10(-4) x min(-1)/microU x mL(-1)), P < .0001), with a negative correlation of S(I) with the adiposity index BMI (r = -.67, P < .0001) and VAT (r = .56, P < .05); (4) RMR, expressed in absolute terms, was significantly increased in obese versus lean subjects (5.9 +/- 0.2 v 4.6 +/- 0.3 kJ/min, P < .01), whereas when RMR was adjusted for fat-free mass (FFM), the difference between the two groups disappeared (0.09 +/- 0.003 v 0.09 +/- 0.002 kJ/min x kg FFM). We did not observe any difference in GIT between lean and obese subjects. Moreover, GIT was significantly correlated with FFM (r = .69, P < .005), but not with BMI. The amount of VAT did not correlate with RMR or GIT. In conclusion, these results suggest that in obese subjects with normal glucose tolerance, insulin sensitivity is impaired and the beta-cell hyperresponse to glucose is mainly due to an enhanced second-phase beta-cell secretion. The degree of visceral fat deposition seems to affect insulin secretion and worsens insulin sensitivity, but does not influence energy expenditure.

Abstract: OBJECTIVE: Synovial fluid effusions and/or arthritis may be found in patients with chronic lymphocytic thyroiditis (CLT), even in the absence of hypothyroidism. Sometimes these arthropathies can be attributed to the rheumatic diseases frequently associated with CLT, but in some instances the arthritis seems to be independent of any of the diseases known to cause arthritis, and therefore remains unclassified. This study was carried out in an attempt to characterize the type and outcome of arthritis associated with CLT. METHODS: We performed a prospective study with a follow-up of 6.42 years (range 4-13) on 33 patients affected with CLT and presenting with arthritis. All conditions known to cause arthritis were previously excluded. Investigations included HLA typing, x-ray of the affected joints and, when possible, synovial fluid (SF) analysis with an interleukin (IL)-1 beta determination. Patients were divided based on their clinical presentation into two groups: those with polyarthritis and those with oligoarthritis. RESULTS: During the follow-up, 8 out of 16 patients with polyarthritis developed severe rheumatoid arthritis, characterized by bone erosions, high levels of SF IL-1 beta and an increased frequency of HLA DR4. The other 8 patients had polyarthritis in a mild, non-erosive form, which responded well to symptomatic drugs. Oligoarthritis, found in 17 patients, also showed a mild evolution, with frequent spontaneous remissions. The non-rheumatoid polyarthritis and oligoarthritis patients were characterised by the absence of bone erosions, low levels of SF IL-1 beta and an increased frequency of HLA DR3. CONCLUSIONS: We conclude that it is possible to find in association with CLT a type of inflammatory arthritis characterized by a mild, non-erosive evolution, low SF levels of IL-1 beta and an increased frequency of HLA-DR3. This arthritis seems to be independent of thyroid dysfunction and shows a clinical pattern similar to the arthritis usually found in connective tissue diseases.

Abstract: Testosterone levels are decreased in obese men but increased in obese women. The interplay between gonadal steroids and leptin is, at present, far from being elucidated. This study was carried out to investigate the relationship between serum leptin, plasma insulin, insulin sensitivity and free testosterone in 46 men (29 obese and 17 lean) and 65 premenopausal women (42 obese and 23 lean). In all subjects, anthropometric parameters and serum levels of insulin, leptin, free testosterone (T), dehydroepiandrosterone sulphate and sex hormone-binding globulin were measured. An oral glucose tolerance test (OGTT) and an insulin tolerance test were also performed to determine the insulin sensitivity index. Our results show a significant difference in serum leptin between lean and obese men (3.19 +/- 0.71 vs. 20.28 +/- 0.26 ng mL-1; P < 0.0005) as well as between lean and obese women (10.78 +/- 2.14 vs. 34.79 +/- 2.26 ng mL-1; P < 0.00001). Basal T concentration in the obese men was significantly lower than in the control group (18.6 +/- 1.3 vs. 23.3 +/- 1.4 ng L-1; P < 0.01), whereas in the obese women it was significantly higher than in the control group (2.0 +/- 0.2 vs. 1.3 +/- 0.1 ng L-1; P < 0.05). When multiple linear regression was performed without body mass index (BMI) in the statistical model, leptin was correlated with basal insulin (P < 0.0001), insulin sensitivity (P < 0.0001) and T (P < 0.0001) in both men and women. When BMI was included in the model as an independent variable, leptin was significantly correlated only with BMI (P < 0.0001), the degree of insulin resistance (P < 0.05) and T (only in men, P < 0.05). This study confirms that serum leptin is strongly correlated with the degree of obesity and female sex. The negative correlation between leptin and T in men, independent of BMI, is consistent with the hypothesis that T may possess an inhibitory effect on adipocyte ob gene transcription.

Abstract: Both hyperglycemia and hyperinsulinemia stimulate whole body and muscle glucose disposal. To define the impact of increased lactate concentration (4-5 mM) on muscle glucose disposal during hyperglycemia, we studied anesthetized normal rats infused with either sodium lactate or sodium bicarbonate as control. Animals were studied under hyperglycemic clamp (13 mM) using [3-3H]glucose (study 1) and 2-deoxy-[1-3H]glucose (study 2) to assess glucose rate of disappearance (Rd), glycolytic flux (GF), glycogen synthesis, and glucose utilization index by different tissues. Moreover, in study 3, the effect of lactate on the pattern of plasma insulin response to hyperglycemia was evaluated. In study 1, lactate infusion resulted in an increased Rd (38.7 +/- 1.7 vs. 32.3 +/- 1.3 mg.min-1.kg-1; P < 0.01), which was explained by an enhanced rate of glycogen synthesis (23.0 +/- 1.7 vs. 14.7 +/- 1.2 mg.min-1.kg-1; P < 0.001), whereas GF was unchanged. In study 2, lactate-infused animals showed an increased 2-deoxy-glucose disposal and a stimulated glycogen synthase activity as well as an increased glycogen accumulation at the end of the study in several skeletal muscles. In study 3, lactate did not induce any change in either early or late insulin response to hyperglycemia. In conclusion, our results show that muscle glycogen deposition may be enhanced by elevated lactate levels under hyperglycemic conditions and support a role for lactate in the regulation of glucose homeostasis.

Abstract: OBJECTIVE: Women with visceral obesity may have hyperactivity of the hypothalamic-pituitary-adrenal (HPA) axis. Since glucocorticoids are involved in the expression of the ob gene, this study was carried out to investigate the relationship between serum leptin and the activity of the HPA axis in women with different obesity phenotypes. DESIGN: Cross sectional clinical study. SUBJECTS: Fifteen obese (Body Mass Index BM128 kg/m2) women and ten normal weight control women (BM126 kg/m2) were included in the study. MEASUREMENTS: Body fat distribution was defined by CT scan at the L4-L5 level. Baseline blood samples were obtained for hormone concentrations. The activity of the HPA axis was evaluated by measuring ACTH and cortisol blood levels after combined iv administration of corticotropin releasing factor (100 microg) + arginine-vasopressin (0.3 IU). RESULTS: Baseline cortisol, ACTH, and androgen levels were similar in all groups, whereas leptin levels were significantly higher in the obese groups than in normal weight controls, without any significant difference between women with different obesity phenotypes. Incremental areas of ACTH and cortisol were significantly higher in women with visceral obesity than in those with subcutaneous obesity and controls. No significant correlation was found between the activity of the HPA axis and leptin concentrations. Leptin showed a highly significant correlation with BMI and subcutaneous fat and a weak but significant correlation with visceral fat and the visceral-to-subcutaneous fat ratio. CONCLUSION: Women with different obesity phenotypes had similar serum leptin concentrations but different HPA axis activity, and there was no correlation between them.

Abstract: A reduction of postprandial thermogenesis has been described in obesity; insulin resistance and/or decreased sympathetic nervous system activity seem to play the major role in its pathogenesis. On the other hand, a normal energy expenditure during exercise has been reported. At present, the response and the role of catecholamines in energy metabolism during exercise in obesity have not been well clarified yet. The aim of this work was to study the metabolic and hormonal changes caused by intense exercise in obesity. Nine obese subjects and ten normal weight controls were submitted to exhaustive exercise on a cycloergometer. Blood glucose, free fatty acids (FFA), glycerol, lactate, beta-OH-butyrate, insulin, glucagon, plasma growth hormone (HGH), catecholamine plasma levels were assayed before and at the end of exercise, and after a recovery period. The energy cost of exercise was evaluated by indirect calorimetry. In our experiment muscular exercise did not provoke any change in blood glucose and FFA plasma levels in either of our groups. In the obese subjects the insulin plasma levels were higher than in the controls. Glucagon plasma levels did not change. The exercise responses of norepinephrine (NE) (4.28 +/- 0.74 vs 8.81 +/- 1.35 nmol/l; P < 0.01), epinephrine (E) (234.21 +/- 64.18 vs 560.51 +/- 83.38 pmol/l; P < 0.01) and plasma growth hormone (HGH) (134.84 +/- 58.97 vs 825.92 +/- 195.25 pmol/l; P < 0.01) were significantly lower in obese subjects. At the end of exercise, the thermic effect of exercise did not differ between obese and control subjects (0.335 +/- 0.038 vs 0.425 +/- 0.040 kJ/min x kg fat-free mass. Our findings indicate that an impaired counterregulatory hormone response to exercise exists in obese subjects. The thermic effect of exercise does not seem to be affected by either the reduced catecholamine response nor insulin resistance.

Abstract: The aim of our study was to investigate whether sodium lactate has any effect on hepatic insulin dynamics in perfused rat liver. Rat livers were perfused in situ with saline or increasing concentrations of sodium lactate, and hepatic insulin extraction was calculated from the difference in insulin concentration between the portal and the suprahepatic vein. Our results show that hepatic insulin extraction at the three lactate concentrations added was higher than in the control experiments (lactate 1 mmol/l: 263 +/- 51 vs. 765 +/- 114, P < 0.005; lactate 5 mmol/l: 341 +/- 80 vs. 906 +/- 109; P < 0.005; lactate 15 mmol/l: 438 +/- 21 vs. 981 +/- 66 microIU.g-1.30 min-1, P < 0.005). No significant differences were observed in net glucose balance across the liver during perfusion with lactate. Moreover perfused liver displayed a net lactate production during infusion with saline or lactate added at the lower concentrations (1 and 5 mM), whereas at the highest (15 mM), a net lactate uptake by the liver was observed (P < 0.05). Our results suggest that, in perfused rat liver, lactate may increase hepatic insulin clearance. Thus energy fuels such as lactate and nonesterified fatty acid have opposite effects on hepatic insulin clearance and may therefore contribute to the regulation of posthepatic insulin delivery.

Abstract: Hyperinsulinaemia and insulin resistance have been hypothesized to be the common pathophysiological factor of hypertension, NIDDM and obesity. To evaluate the possible role of hyperinsulinaemia and insulin resistance on hypertension, we studied a group of 37 patients with insulinoma who were admitted to our department in the period from 1966 to 1990. We recorded blood pressure and assayed blood glucose, plasma insulin, plasma triglycerides and serum uric acid levels, before and after surgery, in these patients and in a 37-subject control group. No significant increase in blood pressure and triglyceride plasma levels was recorded in the chronic hyperinsulinaemic hypoglycaemic patients, suggesting the lack of a direct role of hyperinsulinaemia on hypertension.

Abstract: For many years a series of studies has been carried out to evaluate the role of endogenous opioid peptides on glucose metabolism. In this work we studied the influence of endogenous opioid peptides on insulin response to OGTT and glucose-induced thermogenesis before and after a prolonged oral treatment with Naltrexone (50 mg/daily for 6 days), an opioid receptor antagonist, in a group of 9 obese subjects. Moreover in obese patients we evaluated the effect of this anti-opioid drug on insulin secretion and insulin sensitivity during an IVGTT using the minimal model approach. We compared the pre-treatment results with data coming from a group of 5 normal-weight subjects. We measured blood glucose, plasma insulin and C-peptide concentrations and evaluated the following parameters: first (phi 1) and second (phi 2) phase of beta-cell sensitivity, insulin sensitivity and glucose effectiveness. Obese subjects displayed an increased insulin response to oral and i.v. glucose load, due to an increased first phase of insulin secretion (phi 1), a reduced insulin sensitivity (Si) and glucose effectiveness (Sg) in respect to normal-weight subjects. They showed no difference in glucose and insulin area during oral load and in their profiles during i.v. glucose load after naltrexone treatment. Similarly no significant variation in insulin sensitivity and glucose effectiveness was observed. The glucose-induced thermogenesis, measured by indirect calorimetry, was not modified by naltrexone. Therefore our study demonstrates that endogenous opioids do not play any role in the impairment of peripheral insulin sensitivity and energy expenditure in human obesity.

Abstract: The link between hyperinsulinemia and hypertension is imperfectly understood. Recently, a renin gene (the mouse DBA/REN-2d gene) has been transfected into rats, leading to high blood pressure in transgene-positive animals, TGR(mREN-2)27 rats. We tested whether heterozygous hypertensive TGR(mREN-2)27 rats presented evidence of insulin resistance in comparison with the parent strain of Sprague-Dawley rats. Despite their higher blood pressure (203 +/- 8 vs. 112 +/- 6 mmHg, P < 0.001), transgenic rats had normal fasting levels of plasma glucose, insulin, free fatty acids, and triglycerides and had normal fasting rates of hepatic glucose production (by [14C]glucose infusion). During a euglycemic hyperinsulinemic clamp (3 mU/min), stimulation of whole body glucose utilization was equivalent in transgenic and control animals (12.6 +/- 0.6 vs. 10.9 +/- 1.0 mg.min-1.kg-1, respectively). Likewise, suppression of hepatic glucose output by insulin was complete in both groups. The glucose utilization index (as measured by the 2-deoxy-D-[3H]glucose technique) was similar between transgenic and control animals in several skeletal muscles (soleus, extensor digitorum longus, tibialis, diaphragm, white and red quadriceps, and white and red gastrocnemius), in white adipose tissue (periovarian and inguinal), and in brown adipose tissue. We conclude that single gene hypertension does not alter whole body and individual tissue insulin sensitivity.

Abstract: Given that several genetically obese rodents characterized by hyperphagia, hyperinsulinemia, and insulin resistance have increased hypothalamic neuropeptide Y (NPY) mRNA and peptide content, the impact of NPY administered intracerebroventricularly (i.c.v.) for 7 days to normal, awake rats was investigated. NPY produced marked hyperphagia, increased body weight gain, increased basal insulinemia, and, more importantly, a much greater insulin response to meal feeding than that of saline-infused controls. NPY administration also resulted in a pronounced increase in the in vivo insulin-stimulated glucose uptake by adipose tissue but in a marked decrease in uptake by eight different muscle types. Increased insulin responsiveness of the glucose transport process by adipose tissue was accompanied by increases in both GLUT4 mRNA and protein levels. In contrast, the decreased insulin responsiveness of glucose uptake in muscles from NPY-administered rats was not related to GLUT4 expression. We conclude that i.c.v. NPY administration to normal rats produces a hormonal-metabolic situation that is similar to that reported in the dynamic phase of the genetic obesity of the fa/fa strain. Thus, NPY could be of primary importance in the establishment of obesity syndromes with incipient insulin resistance.

Abstract: Intracerebroventricular neuropeptide Y (NPY) administration to normal rats for 7 days produced a sustained, threefold increase in food intake, resulting in a body weight gain of more than 40 g. Basal plasma insulin and triglyceride levels were increased in NPY-treated compared to vehicle-infused rats by about four- and two-fold, respectively. The glucose utilization index of white adipose tissue, measured by the labelled 2-deoxy-D-glucose technique was four times higher in NPY-treated rats compared to controls. This change was accompanied by an increase in the insulin responsive glucose transporter protein (GLUT 4). In marked contrast, muscle glucose utilization was decreased in NPY-treated compared to vehicle-infused animals. This change was accompanied by an increase in triglyceride content. When NPY-treated rats were prevented from overeating, there was no decrease in muscle glucose uptake, nor was there an increase in muscle triglyceride content. This suggests that muscle insulin resistance of ad libitum-fed NPY-treated rats is due to a glucose-fatty acid (Randle) cycle. When intracerebroventricular NPY administration was stopped and rats kept without any treatment for 7 additional days, all the abnormalities brought about by the neuropeptide were normalized. A tonic central effect of NPY is therefore needed to elicit and maintain most of the hormonal and metabolic abnormalities observed in the present study. Such abnormalities are analogous to those seen in the dynamic phase of obesity syndromes in which high hypothalamic NPY levels have been reported.

Abstract: Recently, a role of beta-Endorphin on peripheral tissue metabolism has been suggested. A lipolytic effect of beta-Endorphin has been observed both in vivo and in vitro in animals but, at present, there is no evidence for a similar effect in humans. In this study, we investigated the lipolytic effect of beta-Endorphin in isolated human adipocytes. beta-Endorphin induced a significant increase in glycerol release in isolated human fat cells. Naloxone was able to inhibit the beta-Endorphin-induced lipolysis. The opioid antagonist alone had no effect on basal lipolysis and on Epinephrine-stimulated lipolysis when administered together with this hormone. Our results suggest that beta-Endorphin may play a role on lipolysis also in human fat cells and that this effect may be mediated by a specific opiate receptor.

Abstract: Chronic intracerebroventricular (icv) administration of neuropeptide-Y (NPY; 10 micrograms/day) was performed in normal female rats to investigate its hormonal and metabolic consequences. Intracerebroventricular NPY produced hyperphagia, increased basal insulinemia, as well as liver and adipose tissue lipogenic activity. It also increased basal morning corticosteronemia. When NPY-induced hyperphagia was prevented by pair-feeding, the icv NPY treatment resulted in the same increases in basal insulinemia and corticosteronemia, and liver and white adipose tissue lipogenesis was still higher than that in respective controls. Under the ad libitum and pair-feeding conditions, icv NPY stimulated glucose uptake as well as total lipoprotein lipase activity in white adipose tissue; it resulted in an increase total activity of hepatic and white adipose tissue acetyl coenzyme-A-carboxylase. As all hormonal and metabolic changes elicited by icv NPY remained present (at the same or to a lesser extent depending upon the parameter considered) when hyperphagia was prevented by pair-feeding, it was, thus, shown that icv NPY per se induces peripheral hormonal and metabolic alterations via efferent routes, which remain to be determined. The effects of icv NPY reported in this study are similar to the defects observed in the early phase of genetic obesity in rodents, the hypothalamus of which has increased NPY levels. NPY could, thus, be of relevance in the occurrence of genetically induced obesity.

Abstract: Anomalies in prostaglandin (PG) synthesis have been suggested in both experimental and human diabetes mellitus; increased levels of plasma and tissue eicosanoids has been recently reported by several investigators. One step in prostaglandin synthesis is the enzymatic hydrolysis of membrane phospholipids by Phospholipase A2 (PLA2). Nevertheless the alternative pathway involving Phospholipase C must be considered. An evaluation of PLA2 activity is therefore a useful method for studying prostaglandin synthesis in the peripheral target tissues of insulin activity. We studied PLA2 activity in normal and diabetic rat muscle. Streptozotocin-induced diabetic rats showed significantly higher muscular PLA2 activity when compared with controls (3.04 x 10(-2) +/- 0.50 x 10(-2) versus 1.34 x 10(-2) +/- 0.35 x 10(-2) arachidonic acid pMol.mg protein-1.min-1 (p less than 0.01). This effect was not observed in diabetic animals successfully treated with insulin (1.78 x 10(-2) +/- 0.5 x 10(-2) versus 1.34 x 10(-2) +/- 0.35 x 10(-2) arachidonic acid pMol.mg protein-1.min-1), and a significant correlation was found between blood glucose and muscular PLA2 activity (r = 0.42; p less than 0.05). Our results clearly show that in streptozotocin-induced diabetic rats muscular PLA2 activity is significantly higher. The relationship between blood glucose levels and muscular PLA2 activity and the decrease of PLA2 activity after insulin treatment suggest that these changes may be related to a defect in insulin effect.

Abstract: In six patients with pheochromocytoma oral glucose tolerance test (OGTT) and arginine test were carried out. Blood insulin and glucagon response were investigated. In subjects with adrenal tumor glycemic curve pattern was typical: a rapid and exaggerated increase of glycemia followed by an abrupt drop. Absolute insulinemic response to oral glucose was normal, but inappropriate to glycemic stimulus. Arginine infusion provoked a slightly above normal increase in blood glucose and a normal increase in blood glucagon. In three of the patients studied postoperatively, reduced glycemic response to glucose was observed, whereas there were no evident variations in blood insulin and glucagon response. These data suggest that in pheochromocytoma impaired glucose tolerance is partly due to the reduced insulin response to oral glucose load.

Abstract: Somatostatin analogue (Sandostatin; SMS 201-995) is utilized as a therapy in acromegaly because of its efficiency in inhibiting GH secretion; it induces some clinical improvements, such as headache remission in acromegalic patient, which seem to be unrelated to Gh normalization. We have examined 8 acromegalic patients, suffering from headache, after injection of saline solution and subsequently of SMS 201-995 (100 y), in order to study the mechanism of analgesic effect induced by Sandostatin administration. Headache, by autovaluation test, heart rate frequency, PAO, sistolic and diastolic blood velocity in medial cerebral artery, by utilizing Transcranial Doppler Sonography (SDSV), have been measured before and after saline and after SMS 201-995. GH and beta-endorphin have been also assayed in plasma samples. All patients have shown a rapid and complete improvement in headache after Sandostatin administration. At the same time we have observed an increase in SDSV and a parallel slight increase in PAO values, more evident in the diastolic phase. Plasma beta-endorphin assay has shown rather conflicting results after SMS 201-995 administration. Our results confirm an important and rapid analgesis effect of Sandostatin on acromegaly headache unrelated to GH normalization. The cerebral emodinamic changes suggest their involvement in Sandostatin induced analgesia.

Abstract: In order to check whether reduced postprandial thermogenesis, as found in obese subjects depends on insulin resistance, the study tested whether the thermogenetic response to glucose in a group of obese subjects and a group of normal weight subjects differed from that obtained using an insulin-independent monosaccharide such as fructose. Nine obese subjects and 6 control subjects were included in the study. An oral glucose tolerance and fructose tolerance test (75 g) was performed in all subjects on different days. Energy expenditure was calculated both in basal conditions and during the test (resting metabolic rate: RMR) using indirect calorimetry expressed per kg of lean weight, as assessed using bioimpedance measurement techniques. Blood samples were collected to assay glycemia and insulinemia. Results show that increased RMR induced by glucose was significantly reduced in the group of obese subjects compared to controls. In the same group of obese subjects, RMR was found to be significantly higher following fructose in comparison to the glucose response but did not differ from that in controls. Data confirm the existence of reduced thermogenesis in obese subjects induced by glucose. The fact that this phenomenon was not recorded in the same subjects following the fructose tolerance test, whose metabolism is insulin-independent, supports the hypothesis that reduced glucose-induced thermogenesis in obese subjects may depend on insulin resistance.

Abstract: Since increased opiate production in obesity has been reported, the effects of naloxone in obese subjects were studied in order to ascertain whether endogenous opioid peptides play a role in the abundant insulin secretion of obesity. The results obtained showed that intravenous administration of naloxone considerably reduced insulin of obese subjects to a mixed meal, whereas it did not modify the blood insulin response to arginine or glucose infusion. Glucagon secretion to ingestion of a mixed meal and to arginine infusion was not modified by the opioid receptor blocking agent. This study seems to indicate that hyperproduction of endogenous opioid peptides in obesity increases insulin secretion stimulated by food intake, whereas it does not appreciably affect insulin production stimulated by circulating glucose or aminoacids.

Abstract: The possibility that dietary-induced thermogenesis may be decreased in obesity has been proposed in recent years. However, the results of human studies so far obtained are conflicting. The present research was undertaken in order to clarify this question. We studied postprandial thermogenesis induced by ingestion of a mixed meal and of a carbohydrate mixture in 15 normal and 12 obese subjects. Blood glucose and plasma insulin levels were measured at the same time. The data obtained have shown that the mean resting metabolic rate (RMR) expressed as a function of body weight3/4, is almost the same in obese as in normal-weight subjects (0.115 +/- 0.018 vs 0.133 +/- 0.021 kj/min/kg3/4, respectively). Moreover, the increment of mixed-meal induced thermogenesis (MM-IT) was 48 +/- 22% in normal and -0.8 +/- 12% in obese subjects, respectively (p less than 0.01). Carbohydrate induced thermogenesis (CHO-IT) appeared slightly higher in normal-weight than in obese subjects (159 +/- 66 vs 98 +/- 46). After carbohydrate ingestion we observed a higher glycemic and insulinemic response in obesity. These results indicate that thermogenesis induced by ingestion of food is reduced in obese subjects; they are also compatible with the idea that insulin resistance could play a role in this phenomenon.

Abstract:

Abstract: BK virus (BKV) DNA was detected by Southern blot hybridization in 19 out of 74 (25.6%) human brain tumors and in 4 out of 9 (44.4%) human tumors of pancreatic islets. BKV DNA was free, in an episomal state and generally in a low copy number (0.2 to 2 genome equivalents per cell). Only occasional tumors contained 10 to 20 genome copies per cell. Viral DNA sequences integrated into cellular DNA were not detected. A number of tumors expressed BKV-specific RNA and T antigen. By transfection of total tumor DNA into human embryonic fibroblasts, viruses with the biological and antigenic properties of BKV were rescued from 6 brain tumors and from 2 tumors of pancreatic islets. Restriction endonuclease mapping of the genomes of the rescued viruses showed that they differ from wild-type BKV. They are all similar to each other and to BKV-IR, a virus previously rescued from a human tumor of pancreatic islets, suggesting the possible association of a BKV variant with specific types of human neoplasms. The significance of the relationship of these BKV variants to human tumors and their possible etiologic role in human oncogenesis are discussed.

Abstract:

Abstract: Prolonged muscular exercise stimulates glucose uptake by the working muscles themselves. The mechanism of this phenomenon is at present unclear. It has been proposed that the kallikrein-kinin-prostaglandin system plays a role in the physiological regulation of muscular glucose metabolism during exercise. Since bradykinin can stimulate phospholipase A2, a key enzymatic step in prostaglandin synthesis, phospholipase A2 activity was assayed in rats at rest and in rats compelled to swim for 60 minutes. The physiological significance of an increase in muscular phospholipase A2 activity is not clear. Since bradykinin can stimulate both muscular glucose uptake and phospholipase A2 activity, it is possible that the increased activity of this enzyme is involved in the exercise-induced increase of muscular glucose uptake. Phospholipase A2 activity was strongly increased in the exercising rat muscles. A small but significant increase in phospholipase A2 activity was observed in the heart, whereas no variation in activity was demonstrated in either the kidney or the liver of exercising rats. These findings strongly indicate that prolonged exercise increases muscular phospholipase A2 activity only in the muscle and heart. This phenomenon appears to be strongly related to muscular contraction, since other stress situations such as cold exposure did not modify phospholipase A2 activity. Our data are in agreement with the hypothesis of a possible involvement of prostaglandins in the priming action of insulin on glucose uptake during muscular work.

Abstract: Many experimental observations show that prolonged physical exercise produces an increase of muscular glucose uptake. Recent findings suggest that the kallikrein-kinin-prostaglandin system may be related to this phenomenon, but so far, no direct evidence of quantitative alteration in this system has been observed during exercise. We measured plasma kallikrein and muscular phospholipase A2 activity, respectively the first and the last steps of reactions leading to prostaglandin synthesis. We demonstrated, for the first time, that during exercise plasma kallikrein activity increases in rats. We also observed an increase of muscular phospholipase A2 activity after exercise and a positive correlation between these parameters. Our findings demonstrate, under physiological conditions of enhanced muscular glucose uptake, a concomitant significant increase of plasma kallikrein and muscular phospholipase A2 activity, supporting the hypothesis that activation of the kallikrein-kinin-prostaglandin system may play some part in the enhanced muscular glucose uptake during physical activity.

Abstract:

Abstract: Type 1 diabetes mellitus is thought to derive from organ-specific autoimmune reactions, probably triggered by environmental factors. In view of the possible involvement of mumps virus and reoviruses in the pathogenesis of autoimmune endocrine disease, serum antibody levels to these viruses were measured in newly-diagnosed diabetic patients aged 5 to 25 years and in matched control subjects. Diabetic patients showed a significantly lower prevalence and reduced titers of antibodies to mumps and reoviruses. By contrast, the antibody response to measles virus (a non-diabetogenic agent) was remarkably similar in the two groups. It is suggested that individuals with an impaired humoral response to some viral agents are at increased risk of developing diabetes when exposed to pancreotropic viruses.

Abstract: Antithrombin III (AT III) levels have been reported to be low, normal, and high in diabetes mellitus. Furthermore, a discrepancy between AT III activity and antigen concentration was reported. We have evaluated the behaviour of AT III activity and antigen level both in type 1 and type 2 diabetes, either in uncontrolled or in well controlled patients. AT III activity and antigen levels showed values similar to normal. No difference was seen between type 1 and type 2 diabetes. Similar results were observed in the group of well controlled diabetic patients. AT III activity and antigen did not correlate with blood glucose and glycosylated haemoglobin (HbA1). No difference was observed between AT III activity and antigen levels in any group. Therefore the hypercoagulable state found in diabetes mellitus does not depend on AT III modifications. A discrepancy between AT III activity and antigen was not confirmed. A dysantithrombinaemia, explained on the basis of an inactivation of protein glycosylation in diabetes mellitus has not been confirmed.

Abstract: It has recently been observed that administration of bradykinin to diabetic patients improves peripheral glucose utilization. To verify whether there is an alteration of the kallikrein-kinin system in human diabetes, plasma kallikrein activity was measured in 47 diabetic patients and in 20 control subjects. In diabetics plasma kallikrein activity was significantly higher than in controls: 1.04 +/- 0.04 U/ml (p less than 0.001). Although they do not refute the hypothesis that there is an alteration of the kallikrein-kinin system in diabetes mellitus, these findings do not support such a hypothesis either. Increased synthesis of plasma kallikrein activity may be due to increased synthesis of carbohydrate-protein compounds in diabetes mellitus.

Abstract:

Abstract: The first phase of the intrinsic coagulation system was studied in twenty-six diabetic outpatients. The parameters tested were kallicrein, prekallicrein, factor XI and factor XII activities. Kallicrein and factor XI showed significantly elevated activities as compared with a normal control group (P less than 0.05 and less than 0.001, respectively). Factor XI and factor XII levels were higher in non-insulin dependent than in insulin dependent diabetics. We found that the kallicrein-prekallicrein system was activated in our diabetic patients and that such activation could probably be secondary to lesions of the vascular endothelium present in long-term diabetics. No correlation was found between glycaemia and activity levels of this system. Our data represent another demonstration of the existence of a hypercoagulable state in diabetes mellitus.

Abstract: In vitro calcium plays a fundamental role in regulating insulin secretion. On the other hand, the influence of calcium excess on insulin release in vivo is not clearly defined. Recently, calcium infusion has been proposed as a provocative test for the diagnosis of insulin-secreting tumors. A 2-h infusion of calcium gluconate was performed (4 mg/kg . h) in six patients with islet cell adenoma. As a result, mean calcium plasma levels increased from 9.6 +/- 0.4 to 11.6 +/- 0.8 mg/100 ml. During calcium infusion, blood glucose and plasma insulin concentrations remained unchanged. These observations suggest that calcium fails to stimulate basal insulin secretion even in cases of organic hyperinsulinism. They show that calcium infusion is not helpful as a provocative test in the diagnosis of insulinoma.

Abstract: L-carnitine is essential for the transport of long-chain fatty acids into mitochondria and their oxidation. Recently, a relationship between plasma free fatty acids (FFA) and L-carnitine metabolism has been observed. Plasma free L-carnitine (FC), FFA, triglycerides, cholesterol, blood glucose concentration and daily excretion of FC were determined in 20 diabetic patients as well as in 18 control subjects. Both in male diabetics and in male controls, plasma FC was significantly higher than in females. Mean plasma FC was found to be significantly reduced in diabetics (21 +/- 2 vs 35 +/- 2 mumol/1 in non-diabetic subjects; p less than 0.005). Daily urinary excretion of FC was clearly lower in diabetic patients than in controls (172 +/- 34 vs 403 +/- 38 mumol/24 h; p less than 0.001). The reduced plasma FC in diabetes mellitus may be due to redistribution between circulating free and esterified carnitine and to increased utilization of FC for synthesis of acylcarnitine in tissues.

Abstract: Although it is well known that growth hormone may influence protein metabolism, few investigations have been undertaken on serum protein levels in acromegaly; recently hypoglobulinemia has been observed in this endocrine disorder. In 28 acromegalic patients and 56 control subjects sera have been analyzed for total protein determination and for electrophoretic protein separation. The results have shown that in acromegalic there is a slight but statistically highly significant decrease of serum total protein (mean +/- SE 6.66 +/- 0.07 g/100 ml vs 7.10 +/- 0.07 g/100 ml), albumin, alpha 1 and alpha 2-globulins. Since growth hormone stimulates protein synthesis, the pathophysiological significance of hypoproteinemia of acromegaly is at present obscure.