Circulating myomiRNAs as biomarkers in patients with Cushing's syndrome.

PURPOSE: Impairment of skeletal muscle mass and strength affects 40-70% of patients with active Cushing's syndrome (CS). Glucocorticoid excess sustains muscle atrophy and weakness, while muscle-specific microRNAs (myomiRs) level changes were associated with muscle organization and function perturbation. The aim of the current study is to explore changes in circulating myomiRs in CS patients compared to healthy controls and their involvement in IGFI/PI3K/Akt/mTOR pathway regulation in skeletal muscle. METHODS: C2C12, mouse myocytes, were exposed to hydrocortisone (HC), and atrophy-related gene expression was investigated by RT-qPCR, WB and IF to assess HC-mediated atrophic signalling. miRNAs were evaluated in HC-treated C2C12 by PCR Arrays. MyomiRs significantly overexpressed in C2C12 were investigated in 37 CS patients and 24 healthy controls serum by RT-qPCR. The anti-anabolic role of circulating miRNAs significantly upregulated in CS patients was explored in C2C12 by investigating the IGFI/PI3K/Akt/mTOR pathway regulation. RESULTS: HC induced higher expression of atrophy-related genes, miR-133a-3p, miR-122-5p and miR-200b-3p in C2C12 compared to untreated cells. Conversely, the anabolic IGFI/PI3K/Akt/mTOR signalling was reduced and this effect was mediated by miR-133a-3p. In CS patients miR-133a-3p and miR-200b-3p revealed higher circulating levels (p < 0.0001, respectively) compared to controls. ROC curves for miR-133a-3p (AUC 0.823, p < 0.0001) and miR-200b-3p (AUC 0.850, p < 0.0001) demonstrated that both myomiRs represent potential biomarkers to discriminate between CS and healthy subjects. Pearson's correlation analysis revealed that circulating levels of miR-133a-3p are directly correlated with 24 h urinary-free cortisol level (r = 0.468, p = 0.004) in CS patients. CONCLUSIONS: HC induces atrophic signals by miR-133a-3p overexpression in mouse myocytes and humans. Circulating miR-133a-3p is promising biomarkers of hypercortisolism.

The dopaminergic control of Cushing's syndrome.

Cushing's Syndrome (CS), or chronic endogenous hypercortisolism, is a rare and serious disease due to corticotroph pituitary (Cushing's disease, CD) and extra-pituitary (ectopic CS) tumours overproducing ACTH, or cortisol-secreting adrenal tumours or lesions (adrenal CS). The first-line treatment for CS is represented by the surgical removal of the responsible tumour, but surgery might be unfeasible or ineffective and medical treatment can be required in a relevant percentage of patients with CS, especially CD and ectopic CS. Corticotroph pituitary and extra-pituitary tumours, as well as adrenal tumours and lesions responsible for CS express dopamine receptors (DRs), which have been found to mediate inhibition of hormone secretion and/or cell proliferation in experimental setting, suggesting that dopaminergic system, particularly DRs, might represent a target for the treatment of CS. Dopamine agonists (DAs), particularly cabergoline (CAB), are currently used as off-label treatment for CD, the most common form of CS, demonstrating efficacy in controlling hormone secretion and tumour growth in a relevant number of cases, with the improvement of clinical picture, and displaying good safety profile. Therefore, CAB may be considered a reasonable alternative treatment for persistent or recurrent CD after pituitary surgery failure, but occasionally also before pituitary surgery, as adjuvant treatment, or even instead of pituitary surgery as first-line treatment in case of surgery contraindications or refusal. A certain beneficial effect of CAB has been also reported in ectopic CS. However, the role of DAs in the clinical management of the different types of CS requires further evaluations.

Long-term high-dose L-arginine supplementation in patients with vasculogenic erectile dysfunction: a multicentre, double-blind, randomized, placebo-controlled clinical trial.

PURPOSE: The current randomized, double-blind, placebo-controlled clinical trial addressed the effects on penile erectile function of relatively high daily oral doses (6 g/day) of L-ARG for 3 months (N = 51) compared to placebo (N = 47), in patients with vasculogenic ED, with comparison between mild-moderate and severe vasculogenic ED. METHODS: The outcome measures included IIEF-6 score and cavernous arteries peak systolic flow velocity (PSV) at dynamic penile duplex ultrasonography (PDU). RESULTS: L-ARG supplementation for 3 months significantly increased IIEF-6 score in the overall cohort (p < 0.0001) and in subgroups of patients with mild-moderate (p < 0.0001) and severe (p = 0.007) vasculogenic ED; PSV was significantly increased in the overall cohort (p < 0.0001) and in patients with mild-moderate (p < 0.0001), but not severe vasculogenic ED. At study completion, 74% of patients improved ED degree category, although only 24% of patients, mainly belonging to the baseline category of mild ED, reached IIEF-6 scores compatible with absence of ED; moreover, 20% of patients, exclusively belonging to the baseline category of mild-moderate vasculogenic ED, reached PSV values compatible with absence of ED. CONCLUSION: The results of the current study demonstrated that supplementation with relatively high doses of L-ARG as a single compound for 3 months significantly improved penile erectile function, assessed by both IIEF-6 score and PSV at dynamic PDU in patients with mild-moderate, and improved IIEF-6 score, but not PSV, in patients with severe vasculogenic ED, therefore suggesting that L-ARG might be an alternative treatment in mild-moderate vasculogenic ED patients experiencing adverse effects or with contraindications for chronic treatment with PDE5i compounds.

The treatment with pasireotide in Cushing's disease: effect of long-term treatment on clinical picture and metabolic profile and management of adverse events in the experience of a single center.

PURPOSES: Pasireotide is the first medical therapy officially approved for adult patients with Cushing's disease (CD) experiencing failure of pituitary surgery or not candidates for surgery. The current study aimed at investigating pasireotide effects on clinical picture and metabolic profile in patients enrolled in the phase III CSOM230B2305 trial at Naples center. In addition, the current study focused on safety issues encountered during the study, detailing the management of the different adverse events associated with the treatment with pasireotide in Naples center. METHODS: Fourteen patients entered the study; eight patients, receiving pasireotide for at least 6 months, were considered for the efficacy analysis, whereas the entire cohort of 14 patients was considered for the safety analysis. RESULTS: Full or partial disease control was obtained in 85.7% of patients, according to a "per-protocol" methodology analysis, and in 42.9% of patients, according to an "intention-to-treat" methodology analysis, after 12 months of treatment. A relevant improvement in clinical signs and symptoms, mainly in facial rubor, supraclavicular fat pad, bruising, hirsutism, and muscle strength was observed; body weight, body mass index, and waist circumference significantly reduced, and a slight non-significant reduction was observed in the prevalence of visceral obesity, hypercholesterolemia, and hypertriglyceridemia. Deterioration of glucose metabolism represented the most common adverse event, occurring in 71.4% of patients, and requiring a dietary regimen as first step, metformin therapy and/or long-acting insulin as second step, and short-acting insulin, as third step; no patients discontinued treatment for hyperglycaemia. Additional adverse events of interest were nausea (21.4%), and vomiting (14.3%), spontaneously resolved in few weeks or some months, except in one patient unsuccessfully treated with metoclopramide and ondansetron, and diarrhoea (14.3%), improved with loperamide treatment. Millimetric gallstones and biliary sludge (7.1%) were managed with ursodeoxycholic acid, inducing lithiasis and biliary sludge resolution, whereas hypocortisolism-related adverse events (7.1%) were resolved with a reduction in the pasireotide dose. CONCLUSIONS: The current study on a limited series of patients contributes to confirm that pasireotide may be considered a valid option for treatment of patients with CD, although it requires an appropriate management of adverse events, especially hyperglycaemia.

Growth hormone/IGF-1 axis longitudinal evaluation in clinically isolated syndrome patients on interferon ß-1b therapy: stimulation tests and correlations with clinical and radiological conversion to multiple sclerosis.

BACKGROUND AND PURPOSE: Growth hormone (GH)/insulin-like growth factor 1 (IGF-1) axis abnormalities in multiple sclerosis (MS) suggest their role in its pathogenesis. Interferon ß (IFN-ß) efficacy could be mediated also by an increase of IGF-1 levels. A 2-year longitudinal study was performed to estimate the prevalence of GH and/or IGF-1 deficiency in clinically isolated syndrome (CIS) patients and their correlation with conversion to MS in IFN treated patients. METHODS: Clinical and demographic features of CIS patients were collected before the start of IFN-ß-1b. IGF-1 levels and GH response after arginine and GH releasing hormone + arginine stimulation tests were assessed. Clinical and magnetic resonance imaging evaluations were performed at baseline, 1 year and 2 years. RESULTS: Thirty CIS patients (24 female) were enrolled. At baseline, four patients (13%) showed a hypothalamic GH deficiency (GHD), whilst no one had a pituitary GHD. Baseline demographic, clinical and radiological data were not related to GHD, whilst IGF-1 levels were inversely related to age (P < 0.001) and GH levels (P = 0.03). GH and IGF-1 serum mean levels were not significantly modified after 1 and 2 years of treatment in the whole group, although 3/4 GHD patients experienced a normalization of GH levels, whilst one dropped out. After 2 years of treatment 13/28 (46%) patients converted to MS. The presence of GHD and GH and IGF-1 levels were not predictive of relapses, new T2 lesions or conversion occurrence. CONCLUSIONS: Growth hormone/IGF-1 axis function was found to be frequently altered in CIS patients, but this was not related to MS conversion. Patients experienced an improvement of GHD during IFN therapy. Longer follow-up is necessary to assess its impact on disease progression.

Insulin-like growth factor-1 predicts cognitive functions at 2-year follow-up in early, drug-naïve Parkinson's disease.

BACKGROUND AND PURPOSE: Cognitive impairment is common in Parkinson's disease (PD), even in the early stages. We aimed to assess the relationship between insulin-like growth factor-1 (IGF-1) and cognitive functions in early, drug-naïve patients with PD. METHODS: Serum IGF-1 was measured in 65 early, drug-naïve patients with PD that underwent a complete neuropsychological battery at baseline and after 2 years. Linear regression analysis was used to evaluate the relationships between neuropsychological scores and IGF-1. Repeated-measures anova was applied to assess changes in neuropsychological variables over time. RESULTS: At baseline, IGF-1 levels were related to phonological fluency. At follow-up, IGF-1 levels were associated with the Rey auditory verbal learning test (RAVLT) - immediate and delayed recall, Frontal Assessment Battery, verbal span and Benton judgement of the line orientation test. Patients with low IGF-1 levels at baseline showed a significantly faster decline of performances than patients with high IGF-1 levels on immediate and delayed recall of the RAVLT and interference task of the Stroop test. CONCLUSIONS: Low serum IGF-1 levels are related to poor performance on executive tasks in early, drug-naïve patients with PD, and may predict poor performance on attention/executive and verbal memory tasks after a 2-year follow-up.

Assessment and clinical implications of RANK/RANKL/OPG pathway as markers of bone tumor progression in patients with NET harboring bone metastases.

INTRODUCTION: The impact on the survival of bone metastases (BM) in patients with neuroendocrine tumor (NET) is a matter of debate. BM have a key role in causing symptoms and in decreasing patients' quality of life. Although the mechanisms of the development of BM are not completely clear, it is now well understood that the Receptor Activator of Nuclear factor Kappa-B-/Ligand (RANK/RANKL)/osteoprotegerin (OPG) pathway plays a relevant role. AIM: To characterize the RANK/RANKL/OPG pathway in patients affected with NET. PATIENTS AND METHODS: Two cohorts of 15 patients each were enrolled in the study; one cohort was affected with NET without BM and the second cohort was affected with NET with BM. The serum RANK/RANKL/OPG pathway was assessed in both the groups. RESULTS: Serum OPG levels and RANKL/OPG ratio were lower and higher, respectively, in NET patients harboring BM than in those without BM. During the ROC analysis, a cut-off value of 1071 pg/ml for OPG and 0.62 for RANKL/OPG ratio were able to significantly distinguish between the two groups. CONCLUSIONS: This study indicates that RANK/RANKL/OPG pathway is imbalanced in patients with NET harboring BM. Specific alterations of this pathway could predict an early development of BM.

Insulin-like growth factor (IGF)-I and IGF-binding protein-3 serum levels in relapsing-remitting and secondary progressive multiple sclerosis patients.

BACKGROUND: Insulin-like growth factor (IGF)-I has a role in remyelination, and insulin-like growth factor-binding protein-3 (IGFBP-3) might reduce its bioavailability. A role of IGFBP-3 in multiple sclerosis (MS) progression was hypothesized in patients with primary progressive (PP) MS. OBJECTIVE: To evaluate serum levels of IGF-I and IGFBP-3 in patients with relapsing-remitting (RR) and secondary progressive (SP) MS and their correlations with disease activity and progression. METHODS: Sixty-three (41 RR and 22 SP) 'naive' MS patients and 60 age-matched healthy controls were enrolled. Patients were assessed through clinical [Expanded Disability Status Scale (EDSS), Multiple Sclerosis Severity Scale (MSSS), number of relapses] and laboratory investigations. IGF-I and IGFBP-3 were measured by ELISA. RESULTS: Levels of IGF-I and IGFBP-3 were similar in the two MS groups. IGFBP-3 levels were higher in patients with MS than in controls (P < 0.001), with a reduction in IGF-I/BP3 ratio (P < 0.001). Patients showing IGFBP-3 levels higher than 2SD of the normal population had a higher EDSS (mean EDSS 3.7 vs. 2.8, P = 0.021). MSSS was not related to IGF-I or IGFBP-3 serum levels. CONCLUSIONS: Our patients showed high IGFBP-3 serum levels respect to controls and higher serum levels were associated with a higher EDSS, despite of comparable disease duration. Therefore, MS and higher disability seem to be associated with a reduction in bioavailability of IGF-I. MSSS score was not related to IGFBP-3 levels, suggesting that IGFBP-3 might not have the pathogenetic role previously suggested for PP MS, in the mechanism of progression in the SP form of disease.

The GH-IGF system in amyotrophic lateral sclerosis: correlations between pituitary GH secretion capacity, insulin-like growth factors and clinical features.

BACKGROUND AND PURPOSE: The growth hormone (GH) and insulin-like growth factor (IGF) system may be involved in neurodegenerative processes, and some abnormalities have been reported in amyotrophic lateral sclerosis (ALS). Our aim was to investigate the GH-IGF axis in patients with ALS and evaluate correlations between this endocrine system and clinical features. METHODS: Serum levels of GH, IGF-I, IGF-II, insulin, IGF-binding protein 1 (IGF-BP1), and IGF-binding protein 3 (IGF-BP3) were measured in 25 patients with ALS and 25 age-, gender-, and BMI-matched healthy controls. A GHRH plus arginine test was performed in patients and controls. Clinical status of patients was evaluated with the ALS Functional Rating Scale - Revised (ALSFRS-R) and upper motor neuron (UMN) score. RESULTS: GHRH plus arginine test showed GH deficiency (GHD) in 13 (52%) patients with ALS; severe GHD was found in 6 (24%) and partial GHD in 7 (28%) patients. IGF-I levels were significantly higher in patients with ALS than in healthy controls (182.9 +/- 90.8 vs. 139.4 +/- 58.1 ng/ml; P = 0.015). IGF-I levels were higher in patients with ALS with UMN score >10 than those with UMN score <10 (217.8 +/- 100.8 vs. 155.5 +/- 74.6 ng/ml, P = 0.05). IGF-II levels were significantly lower in patients with ALS than in healthy controls (720.9 +/- 215 vs. 1001.9 +/- 475.4 ng/ml; P = 0.03). CONCLUSIONS: The results demonstrate an impairment of the GH-IGFs system in ALS. The degenerative process in ALS might lead to a compensatory increase in IGF-I in an attempt to provide additional support to motor neurons or degenerating muscle fibers. The decrease in IGF-II levels may also be of pathological significance.

[Effects of different chicken breeding on microbiological characteristics of fresh eggs]. / Effetti di diversi sistemi di allevamento sulle caratteristiche microbiologiche delle uova fresche.

The influence of three different systems of chicken breeding (battery, outdoor and free-range type) on contamination of the shell, yolk and albumen on category A eggs was evaluated. The eggs collected from free-range type did not resulted in conforming according to Italian legislation (5%) that foresees total absence of Salmonella spp. both on shells that yolk and albumen. Moreover the handled eggs can transmit microorganisms present on the shell because non-hygienic operator's hands favour horizontal bacterial transmission. In conclusion, this survey revealed that the breeding type as well as storing time and handling quality influence the product quality.