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Feeding disorders (FDs) are a large group of behaviours characterized by selectivity, inadequate intake and refusal of food. These disorders are frequent in children (10-15% < 3 years old) and their causes are multiple and variably combined. FDs have a significant impact in everyday life for the child, their family and caregivers inside or outside the school environment. In the last few years in the Department of Paediatric Dysphagia in Bologna (Italy) an increase in the number of healthy children with insufficient oral intake or extremely selective food behaviour, due to selecting in type, texture, smell or appearance of food, has been observed. These children tend to develop dysfunctional or oppositional behaviours during mealtime. This growing need requires the implementation of a network with local paediatricians that focalize the feeding disorder and refer the family to a multidisciplinary assessment. The multidisciplinary team identifies behavioural strategies, structural changes to the mealtime and at the same time introduces intervention for the sensorial normalization. The primary goal is the caregivers’ training: parents and school staff are constantly supported as they offer the child new, different, pleasant and adequate food proposals, avoiding to stop at first refusal. Parent training plays a significant role during such intervention, as it reduces mealtime stress and improves child-parent relationship. The introduction of new strategies during mealtime in a playful, positive and cheerful setting encourages the child’s willingness to taste new foods or new textures. Moreover, their peer and parent modelling improves the outcome.

Purpose: In search of novel strategies to improve the outcome of advanced prostate cancer, we considered that prostate cancer cells rearrange iron homeostasis, favoring iron uptake and proliferation. We exploited this adaptation by exposing prostate cancer preclinical models to high-dose iron to induce toxicity and disrupt adaptation to androgen starvation. Experimental Design: We analyzed markers of cell viability and mechanisms underlying iron toxicity in androgen receptor–positive VCaP and LNCaP, castration-resistant DU-145 and PC-3, and murine TRAMP-C2 cells treated with iron and/or the antiandrogen bicalutamide. We validated the results in vivo in VCaP and PC-3 xenografts and in TRAMP-C2 injected mice treated with iron and/or bicalutamide. Results: Iron was toxic for all prostate cancer cells. In particular, VCaP, LNCaP, and TRAMP-C2 were highly iron sensitive. Toxicity was mediated by oxidative stress, which primarily affected lipids, promoting ferroptosis. In highly sensitive cells, iron additionally caused protein damage. High-basal iron content and oxidative status defined high iron sensitivity. Bicalutamide–iron combination exacerbated oxidative damage and cell death, triggering protein oxidation also in poorly iron-sensitive DU-145 and PC-3 cells. In vivo, iron reduced tumor growth in TRAMP-C2 and VCaP mice. In PC-3 xenografts, bicalutamide–iron combination caused protein oxidation and successfully impaired tumor expansion while single compounds were ineffective. Macrophages influenced body iron distribution but did not limit the iron effect on tumor expansion. Conclusions: Our models allow us to dissect the direct iron effect on cancer cells. We demonstrate the proof of principle that iron toxicity inhibits prostate cancer cell proliferation, proposing a novel tool to strengthen antiandrogen treatment efficacy.

Adaptation to import iron for proliferation makes cancer cells potentially sensitive to iron toxicity. Iron loading impairs multiple myeloma (MM) cell proliferation and increases the efficacy of the proteasome inhibitor bortezomib. Here, we defined the mechanisms of iron toxicity in MM.1S, U266, H929, and OPM-2 MM cell lines, and validated this strategy in preclinical studies using Vk*MYC mice as MM model. High-dose ferric ammonium citrate triggered cell death in all cell lines tested, increasing malondialdehyde levels, the by-product of lipid peroxidation and index of ferroptosis. In addition, iron exposure caused dose-dependent accumulation of polyubiquitinated proteins in highly iron-sensitive MM.1S and H929 cells, suggesting that proteasome workload contributes to iron sensitivity. Accordingly, high iron concentrations inhibited the proteasomal chymotrypsin-like activity of 26S particles and of MM cellular extracts in vitro. In all MM cells, bortezomib-iron combination induced persistent lipid damage, exacerbated bortezomib-induced polyubiquitinated proteins accumulation, and triggered cell death more efficiently than individual treatments. In Vk*MYC mice, addition of iron dextran or ferric carboxymaltose to the bortezomib-melphalan-prednisone (VMP) regimen increased the therapeutic response and prolonged remission without causing evident toxicity. We conclude that iron loading interferes both with redox and protein homeostasis, a property that can be exploited to design novel combination strategies including iron supplementation, to increase the efficacy of current MM therapies.

IntroductionSome hemodialysis patients are reluctant to COVID-19 for the development of adverse events (AEs). The aim of this study was to verify the safety of mRNA-1273 vaccine in hemodialysis patients.MethodsWe conducted a retrospective analysis of in-center hemodialysis patients who underwent mRNA-1273 vaccine from March 1st to April 30th, 2021. All AEs occurring after the first and the second doses were collected and classified as local or systemic.ResultsOverall, 126 patients on chronic maintenance dialysis without a prior COVID-19 diagnosis were vaccinated with two doses of mRNA-1273 vaccine. Mean age was 68 (IQR, 54,7-76) years and 53.6% of patients were aged ≥ 65 years. During the observational period of 68 (IQR, 66-70) days, AEs occurred in 57.9% and 61.9% of patients after the first dose and second dose, respectively. The most common AEs were: injection-site pain (61.9%), erythema (4.8%), itching (4.8%), swelling (16.7%), axillary swelling/tenderness (2.4%), fever (17.5%) headache (7.9%), fatigue (23.8%), myalgia (17.5%), arthralgia (12.7%), dyspnoea (2.4%), nausea/vomiting (7.1%), diarrhoea (5.6%), shivers (4%) and vertigo (1.6%).The rates of local AEs were similar after the first and second doses (P=0.8), whereas systemic AEs occurred more frequently after the second dose (P=0.001). Fever (P=0.03), fatigue (P=0.02) and nausea/vomiting (P=0.03) were significantly more frequent after the second dose of the vaccine. There were no age-related differences in the rate of AEs. Overall, vaccine-related AEs in hemodialysis patients seem to be lower than in the general population.ConclusionRNA-1273 vaccine was associated with the development of transient AEs after the first (57.9%) and second doses (61.9%) in patients on chronic maintenance hemodialysis. Systemic AEs were more common after the second dose. Overall, all AEs lasted for a few days, without any apparent sequelae.

BackgroundKidney transplant recipients with COVID-19 are at high risk of poor outcome because of comorbidities and immunosuppression. The effects of immunosuppressive therapy reduction are unclear in patients with COVID-19.MethodsWe conducted a retrospective study on 45 consecutive kidney transplant recipients followed at the University Hospital of Modena who tested positive for COVID-19 by RT-PCR analysis.ResultsThe median age of patients was 56.1 (interquartile range, [IQR] 47.3-61.1) years with a predominance of male (64.4%). Kidney transplantation vintage was 10.1 (2.7-16) years, and more than half of patients (55.6%) was on triple immunosuppressive therapy. Early reduction of immunosuppression occurred in 62.8% of patients and included antimetabolite (88.8%) and calcineurin inhibitor withdrawal (22.2%).Of the 45 patients, 88.9% became symptomatic and 40% required hospitalization. Overall mortality was 17.8%. There were no differences in outcomes between full- and reduced-dose immunosuppressive therapy at the end of follow-up. One hospitalized patient experienced irreversible graft failure. There were no differences in serum creatinine level and proteinuria in non-hospitalized patients with COVID-19. Admitted patients had better kidney function after dismission (P=0.019). Risk factors for death were age (odds ratio [OR]: 1.19; 95%CI: 1.01-1.39), and duration of kidney transplant (OR: 1.17; 95%CI: 1.01-1.35). One kidney transplant recipient experienced symptomatic COVID-19 reinfection after primary infection and anti-SARS-CoV-2 mRNA vaccine.ConclusionsDespite the reduction of immunosuppression, COVID-19 affected survival of kidney transplant recipients with COVID-19. Age and duration of kidney transplant were independent predictors of death in COVID-19. Early kidney function was favorable in most survivors after COVID-19.

Research has shown emotions affect decision-making in ways that do not simply undermine rationality. Instead, in recent decades researchers have recognized that emotions also motivate and focus individuals and moderate how they make decisions. Initial research into emotions divided these simply into positive and negative, but this perspective has largely been displaced in political psychology by an emphasis on the impact of distinct emotions; among these, anxiety has received the most scholarly attention, rivaled only by anger. The causes of anxiety, also termed fear and unease, are diverse, but research highlights certain attributes of situational evaluation such as low self-control, low certainty, and low external agency. Once present, anxiety has important consequences for decision-making. First, anxiety increases how much information individuals seek out, a pattern of behavior meant to reduce uncertainty. Second, anxiety decreases heuristic processing and weakens the reliance of underlying convictions in determining decisions. Instead, anxious individuals are more likely to think systematically about choices they face. Importantly, anxiety can affect choices and decisions even if they are not directly related to what caused anxiety to emerge, that is, if anxiety is incidental rather than integral. In addition to influencing how people make decisions, anxiety may also directly influence the decisions individuals make. Thus, anxiety increases risk aversion, leading individuals to choose safer paths of action. Anxiety also makes individuals less likely to take action at all, with the most common response being withdrawal and passivity. Applied to political decision-making, anxiety may have the important consequence of decreasing political participation. Research into the role of anxiety in decision-making is fast moving and vibrant, but to become fully established it needs to ensure rigor in measurement and research design; this will require considerable methodological research. Substantively, future research should focus on the effects of elite messages on anxiety as well as on how anxiety influences citizen attitudes and evaluations.

Complex ‘Big Data’ questions that involve machine learning require large datasets for training. This is particularly problematic for Deep Learning methods in the biomedical imaging domain and specifically Digital Pathology. Transfer Learning has been shown to be a promising method for training classifiers on smaller sized datasets. In this work we investigate the effectiveness of aggregated Transfer Learning using VGG19 trained on ImageNet and then fine-tuning parameters with tissue histopathological patches from breast cancer metastatic tissue patches to classify soft tissue sarcoma patches. We compare results with and without transfer learning, and fine tuning applied to different layers. From the results, it is apparent that fine-tuning earlier VGG19 convolutional blocks with breast cancer patches and applying bottleneck feature extraction to soft tissue sarcoma can have an adverse effect on accuracy and other performance measures. Nevertheless, the aggregated approach is a promising method for digital pathology and requires much more investigation.

Ulceration of the plantar aspect of the first metatarsophalangeal joint is a common localization in the diabetic foot. Conservative treatment of this lesion is a challenging problem, performed through the soft tissues and osseous debridement. The present study included a cohort of 28 patients affected by diabetes mellitus and a first ray lesion penetrating the bone. After surgical debridement with removal of the infected bone, we positioned antibiotic-loaded bone cement and stabilized the treated area with an external fixator. All patients with critical limb ischemia had their vascular disease treated before the procedure. The mean follow-up was 12.2 ± 6.9 months. Four patients developed a relapse of the ulceration after the procedure. In the postoperative period, 1 patient (3.57%) developed dehiscence of the surgical site and underwent a second procedure. In the follow-up period, 2 patients (7.14%) experienced bone cement dislocation. In 1 of these patients, a new ulceration was observed dorsally to the surgical site. The approach was surgical revision with bone cement replacement and stabilization with a new external fixator. In the other patient, given the absence of ulcerations, the cement was removed, and arthrodesis with internal stabilization using 2 cannulated screws was performed. One patient (3.57%), who had developed a relapse of ulceration after recurrent critical ischemia, underwent a percutaneous revascularization procedure and transmetatarsal amputation. During the follow-up period, no ulceration recurrences, transfer ulcerations, shoe fit problems, or gait abnormalities were detected in the other 24 patients. Our study presents the results of a technique requiring a 1-stage surgical approach to a relatively common problem, which is often difficult to solve.

The inflammatory microenvironment induces tumours to acquire an aggressive and immunosuppressive behaviour. Since acid sphingomyelinase (A-SMase) downregulation in melanoma was shown to determine a malignant phenotype, we aimed here to elucidate the role of A-SMase in the regulation of tumour immunogenic microenvironment usingin vivomelanoma models in which A-SMase was either downregulated or maintained at constitutively high levels. We found high levels of inflammatory factors in low A-SMase expressing tumours, which also displayed an immunosuppressive/protumoural microenvironment: high levels of myeloid-derived suppressor cells (MDSCs) and regulatory T lymphocytes (Tregs), as well as low levels of dendritic cells (DCs). In contrast, the restoration of A-SMase in melanoma cells not only reduced tumour growth and immunosuppression, but also induced a high recruitment at tumour site of effector immune cells with an antitumoural function. Indeed, we observed a poor homing of MDSCs and Tregs and the increased recruitment of CD8+and CD4+T lymphocytes as well as the infiltration of DCs and CD8+/CD44highT lymphocytes. This study demonstrates that change of A-SMase expression in cancer cells is sufficientper seto tunein vivomelanoma growth and that A-SMase levels modulate immune cells at tumour site. This may be taken into consideration in the setting of therapeutic strategies.

VLCKD is becoming an elective choice to promote weight loss, especially in the case of severe obesity and its metabolic complications. Some authors have assumed that VLCKD can better pre­serve lean body mass during weight loss, saving from diet-induced sarcopenia. BIA evaluation is a simple method to estimate body compostion. At the Maggiore Hospital in Bologna in 2017-2018 we conducted a dietary intervention with a VLCKD ranging from 700 to 800 kcal-die for 1 month in 52 female patients (mean age 48 years), to estimate the effectiveness on body composition. Biochemical indices exploring metabolic and nutritional condition and BIA evaluation was performed before and after VLCKD Before and after VLCKD the average of weight (108,6 vs 104,6), BMI (41,3 vs 39,6) and FM (50,5 vs 47,7 kg) was significantly different, meanwhile no signficant differences was found in Phase angle (6,5 vs 6,35), Body cell mass (BCM: 31,4 vs 30,5 kg) and Muscle Mass (MM: 38,6 vs 37,6). Significantly different were instead the value of basal insulin (12.9 vs 9.1) and the HOMA index (3,2 vs 2), but no differences was found for basal glycemia (94 vs 89), creatinine (0,7 vs 0,7) and serum electrolytes. Our study showed that a VLCKD was highly effective in terms of body weight reduction without to induce body cell mass loss, preventing the risk of sarcopenia. It was also able to reduce the basal insulin level without modify glycemia level and not to change kydney function and electrolytes level

Various nutritional evaluation tools have been proposed over the years. A good tool should involve simple and easy-to-detect measures and should be applicable to a large number of patients. In the home setting it is even more difficult to collect useful data for nutritional evaluation. Our unit carries out the evaluations and prescriptions for the HEN in patients discharged from the same presidium and for patients discharged from other structures but afferent to the territory of the AUSL. During the 1st evaluation, the anthropometric parameters (height weight or, where not available, arm and calf circumference plus ulna lenght) and nutritional indices are required. A clinical-laboratory follow-up is provided at fixed intervals. From January 2018 to August 2018, 50 patients initiated HEN (19 M and 31 F, average age 76 ± 22 years). 20 patients were carriers of PEG and 30 of SNG. At the 1st evaluation the average weight was 59 kg and the average BMI was 22.5 ± 4.2, not indicative of malnutrition, but the mean values of Albumin (2.6 ± 0.6 g/L) Prealbumin (12 ± 8 mg/dl) Total Protein (5.6 ± 1.1 g/dl) Transferrin (141 ± 75 mg/dl) and Lymphocytes (1312 ± 640) were all indicative of moderate malnutrition. Follow-up data collection is in progress. Our results show that weight and BMI in HEN setting are not really indicative of the nutritional status of patients, but it is necessary to use more specific tools such as the dosage of biochemical nutritional indices.

We investigated the significance of microangiopathy in the development of foot ulcer, which is still disputed. We assessed microangiopathy by histological analysis of the capillary ultrastructure using transmission electron microscopy and capillary density and arteriolar morphology in paraffin-embedded sections from the skin of type 2 diabetic patients: 30 neuroischaemic patients (Isc) revascularised with peripheral angioplasty and 30 neuropathic patients (Neu) with foot ulcer, compared with ten non-diabetic volunteers. In the diabetic patients, capillaries in the dermal papillary layer were fewer (−22.2%, 159 ± 43 vs 205 ± 52 mm2 in non-diabetic volunteers, p

In recent years the Italian party system has experienced major changes. The crisis of party government caused by the collapse of the ‘First Republic’ and its main political parties might make Italy...

Delayed Enhancement Magnetic Resonance Imaging can be used to non-invasively differentiate viable from non-viable myocardium within the Left Ventricle in patients suffering from myocardial diseases. Automated segmentation of scarified tissue can be used to accurately quantify the percentage of myocardium affected. This paper presents a method for cardiac scar detection and segmentation based on supervised learning and level set segmentation. First, a model of the appearance of scar tissue is trained using a Support Vector Machines classifier on image-derived descriptors. Based on the areas detected by the classifier, an accurate segmentation is performed using a segmentation method based on level sets.

Pax proteins are transcriptional regulators that control a variety of developmental decisions in vertebrates. During development, the paired-box gene 8 (PAX8) is expressed in the thyroid, kidney, and several areas of the central nervous system. It is also expressed in the adult thyroid gland, in which it mediates TSH-induced modulation of the expression of important genes, such as those encoding thyroglobulin, thyroperoxidase, and the sodium/iodide symporter (NIS). Thus far, placental expression of PAX8 has been described only in mice. In the present study, we show that PAX8 is also expressed in the human placenta at term. In an in vitro model of placental cancer, the JAR choriocarcinoma cell line, human chorionic gonadotropin (hCG) increased levels of PAX8 mRNA and protein, and gel retardation assays indicated that the up-regulation of PAX8 protein expression is associated with an increase in its DNA-binding activity. The effects of hCG were mimicked by forskolin, indicating that they are cAMP dependent. Levels of mRNA for the Wilms' tumor 1 (WT1) and NIS genes were increased in JAR cells by hCG treatment, whereas overexpression of PAX8 increased only levels of WT1 mRNA. In cells transfected with PAX8-specific small interfering RNA, the stimulatory effects of hCG on WT1 mRNA levels were abolished, but hormonal enhancement of NIS mRNA levels was unchanged. These findings indicate that, in JAR cells, hCG activates a cAMP-dependent pathway that can up-regulate WT1 expression through PAX8.

Background: Cross-reacting material 197 is a nontoxic mutant of diphtheria toxin that cross-reacts immunologically with the native counterpart. The molecule was recently tested as a systemic anticancer agent with promising results but its exact mechanism of action has remained unclear. Objective: The main goal of this study was to investigate this mechanism through the possible histologic changes induced by direct inoculation of the molecule into a tumor lesion. A secondary objective was to evaluate toxic, hematologic and antitumor effects of the procedure. Methods: Cross-reacting material 197 was administered by injection into accessible lesions of 12 patients with advanced cancer refractory to standard therapies. Two different doses were used according to the maximal diameter of the selected lesions. Lesions up to 2.5 cm were injected with 0.017 mg/day contained in 0.33 ml of a dilution buffer. Lesions up to 5.0 cm were injected with 0.051 mg/day contained in 1.0 ml of dilution buffer. The treatment schedule consisted of six injections administered on alternate days. A biopsy was taken from each selected lesion before the start of treatment and again 15 days later. Results: Toxicities and effects of cross-reacting material 197 on the injected lesion varied according to the patients’ degree of reactivity to diphtheria toxin. Patients with poor reactivity experienced negligible toxicity and no change or minimal change in the number of circulating neutrophils, in the post-treatment histology of their lesions and in the final lesion size. Patients with strong reactivity experienced mild-to-moderate flu-like syndrome, a prominent increase in the number of circulating neutrophils and a dense neutrophilic infiltration of the injected lesion leading to inflammation and, sometimes, to suppuration. Shrinkage of the index lesion ranging from 60 to 100% was observed in three of the patients. Conclusion: Neutrophils seemed to play a crucial role in achieving these results. One patient was available for post-treatment biopsy only at day 30 from the start of treatment. Examination of these specimens evidenced a striking granulomatous reaction surrounding a core of dead and viable tumor cells.

Fournier's gangrene is a life-threatening acute necrotizing fasciitis of perianal,genitourinary and perineal areas. Nowadays, is well known that Fournier gangrene is almost never an idiopathic disease. In this article we report a case of a 70-year-old patient that initially was not treated properly. The gold standard therapy of the Fournier's gangrene remains today a complete, early and extended surgical debridement.

Blood cadmium (B-Cd) levels were determined in non-occupationally exposed men aged 55–75 years living in the Rome area (Italy) who participated in an epidemiological survey for coronary heart disease (New Risk Factors Project). Among the subjects considered in this study (n= 1223), the geometric mean and the geometric standard deviation for B-Cd were 4.58 and 1.84 nmol/liter, respectively. In never smoked and current smokers, the geometric means (geometric standard deviation) for B-Cd were 2.98 nmol/liter (1.61 nmol/liter) and 7.83 nmol/liter (1.68 nmol/liter), respectively. In linear regression analysis B-Cd levels were significantly and positively associated with daily cigarette consumption (r= 0.5501,P< 0.0001), alcohol consumption (g/day;r= 0.1033,P= 0.0003), non–high-density-lipoprotein cholesterol levels (mmol/liter;r= 0.0848,P= 0.0030), triglyceride levels (mmol/liter, after log-normal transformation;r= 0.0695,P= 0.0151), and blood lead levels (μmol/liter, after log-normal transformation;r= 0.2506,P< 0.0001). B-Cd levels were inversely related to skinfold thicknesses (mm;r= −0.1094,P< 0.0001), body mass index (kg/m2;r= −0.0482,P= 0.0922), and diastolic blood pressure (mm Hg;r= −0.0638,P= 0.0257). No linear correlation was found between B-Cd and systolic blood pressure or mean blood pressure. In multiple regression analysis daily cigarette consumption and blood lead levels were significant predictors of B-Cd level, explaining 30.26 and 2.86% of total variance, respectively. B-Cd was not a significant predictor of diastolic, systolic, or mean blood pressure in multiple regression analyses.

This paper aims to identify approaches that generate appropriate synthetic data (computer generated) for cardiac phase-resolved blood-oxygen-level-dependent (CP-BOLD) MRI. CP-BOLD MRI is a new contrast agent-and stress-free approach for examining changes in myocardial oxygenation in response to coronary artery disease. However, since signal intensity changes are subtle, rapid visualization is not possible with the naked eye. Quantifying and visualizing the extent of disease relies on myocardial segmentation and registration to isolate the myocardium and establish temporal correspondences and ischemia detection algorithms to identify temporal differences in BOLD signal intensity patterns. If transmurality of the defect is of interest pixel-level analysis is necessary and thus a higher precision in registration is required. Such precision is currently not available affecting the design and performance of the ischemia detection algorithms. In this work, to enable algorithmic developments of ischemia detection irrespective to registration accuracy, we propose an approach that generates synthetic pixel-level myocardial time series. We do this by 1) modeling the temporal changes in BOLD signal intensity based on sparse multi-component dictionary learning, whereby segmentally derived myocardial time series are extracted from canine experimental data to learn the model; and 2) demonstrating the resemblance between real and synthetic time series for validation purposes. We envision that the proposed approach has the capacity to accelerate development of tools for ischemia detection while markedly reducing experimental costs so that cardiac BOLD MRI can be rapidly translated into the clinical arena for the noninvasive assessment of ischemic heart disease.

One of biggest problems when assessing radical surgery of prostatic cancer is progression of the locally advanced disease when lymph nodes are positive. The following questions should be asked: 1) if there are patterns of lymph node involvement 2) if retroperitoneal lymphadenectomy is indispensable in pre-operative clinical removal 3) if the prognosis, in relation to lymph node invasion, worsens with the increase in number of positive lymph nodes or if the lesion becomes bilateral. The authors’ experience is reported.

Late enhancement cardiac magnetic resonance images (MRI) has the ability to precisely delineate myocardial scars. We present a semi-automated method for detecting scars in cardiac MRI. This model has the potential to improve routine clinical practice since quantification is not currently offered due to time constraints. A first segmentation step was developed for extracting the target regions for potential scar and determining pre-candidate objects. Pattern recognition methods are then applied to the segmented images in order to detect the position of the myocardial scar. The database of late gadolinium enhancement (LE) cardiac MR images consists of 111 blocks of images acquired from 63 patients at the University Hospital Southampton NHS Foundation Trust (UK). At least one scar was present for each patient, and all the scars were manually annotated by an expert. A group of images (around one third of the entire set) was used for training the system which was subsequently tested on all the remaining images. Four different classifiers were trained (Support Vector Machine (SVM), k-nearest neighbor (KNN), Bayesian and feed-forward neural network) and their performance was evaluated by using Free response Receiver Operating Characteristic (FROC) analysis. Feature selection was implemented for analyzing the importance of the various features. The segmentation method proposed allowed the region affected by the scar to be extracted correctly in 96% of the blocks of images. The SVM was shown to be the best classifier for our task, and our system reached an overall sensitivity of 80% with less than 7 false positives per patient. The method we present provides an effective tool for detection of scars on cardiac MRI. This may be of value in clinical practice by permitting routine reporting of scar quantification.

The procedures adopted in the Italian external quality assessment scheme (EQAS) for blood Pb (B-Pb) and Cd (B-Cd) determination, including the preparation of control materials, are described. Each scheme involves the use of internal quality control materials and the participation in periodical external quality assessment trials. All control materials are prepared at the Laboratory of Clinical Biochemistry, Italian National Institute of Health (Istituto Superiore di Sanita, ISS), as the reference centre. Computerized procedures have been adopted whenever possible, i.e. sample randomization, data transfer and treatment. The analytical performance of the participating laboratories, evaluated from the results obtained in ten years of activity, is reported. A total of eight phases has been carried out between 1983 and 1993. Over the course of these ten years, a positive trend was observed for B-Pb towards a smaller bias and a lower percentage of laboratories with poor performance. In the eighth phase, taking into account the results from all participants, the mean absolute inaccuracy for B-Pb was 32 μg/l (mean Pb concentration in control samples, 165 μg/l). The percentage of laboratories providing results within established acceptability limits for at least 80% of the examined samples (good performers) increased with time from 28% in the first phase to 48% in the last phase. The laboratories providing less than 50% of acceptable results (poor performers) decreased from 35% in the first phase to 22% in the last phase. As regards B-Cd, mean absolute inaccuracy decreased from 0.96 μg/l to 0.55 μg/l between the first phase and the sixth phase. The percentage of good performers increased from 45% to 62% and that of poor performers decreased from 31% to 5%. From the seventh phase a large group of new laboratories agreed to participate in the EQAS for B-Cd and the inaccuracy rose to 0.75 μg/l (mean concentration: 2.7 μg/l) and the percentage of poor performers rose to 24%, whereas the percentage of good performers remained almost unchanged.

Blood lead (B-Pb) levels were determined in 1978-1979, 1980-1981, and 1985-1986 in subjects living in various regions in Italy according to the regulations contained in European Community Directive 77/312 and in Presidential Decree DPR 496/82. From the data obtained, a lowering time trend in median B-Pb levels was evident for subjects from most of the considered Italian regions. In the total samples of male subjects, the median B-Pb levels were 200, 179, and 153 μg/liter in 1978-1979, 1980-1981, and 1985-1986, respectively. In female subjects median B-Pb levels were 140 μg/liter in 1978-1979, 122 μg/liter in 1980-1981, and 100 μg/liter in 1985-1986. A lowering time trend in B-Pb levels has been also observed in the period between 1980 and 1992 in subjects living in the province of Rome. Median B-Pb levels decreased from 190 to 66 μg/liter in males and from 150 to 42 μg/liter in females. During the period of the monitoring program, all over the national territory, the maximum lead content in gasoline has been progressively reduced from 0.635 g/liter in 1980 to 0.15 g/liter in 1991. These data demonstrate how an appropriate measure may succeed in reducing environmental lead contamination, finally resulting in an actual reduction in the level of human exposure. Within the framework of the monitoring program, we studied the associations of blood lead to selected social, environmental, constitutional, and biochemical factors and to clinical parameters. B-Pb was directly related to alcohol consumption, cigarette smoking, and car-driving habits. A positive association was also found between B-Pb and both HDL-cholesterol and γ-glutamyl-transpeptidase, two biochemical indexes of alcohol consumption. Finally, B-Pb was directly and significantly related to both diastolic and systolic blood pressure.

The association of blood lead (B-Pb) concentration to blood pressure was investigated in men aged 55 to 75 years living in the Rome area, who had no history of exposure to lead in the workplace and who participated between 1989 and 1990 in an epidemiologic survey for coronary heart disease (New Risk Factor Project). Of the 1856 individuals eligible for the study, 59 were excluded from analyses because not all relevant data were available; and 478 were excluded because they were treated for hypertension. In the remaining subjects (n = 1319) the median B-Pb concentration was 113 micrograms/l (range: 40-442 micrograms/l). Systolic blood pressure (SBP) averaged 140 +/- 18 (standard deviation) mm Hg (range 98-220) and diastolic blood pressure (DBP) 84 +/- 9 mm Hg (range 56-118). Median B-Pb values increased significantly from 111 micrograms/l in subjects with normal blood pressure (n = 668) to 113.5 micrograms/l in subjects with borderline high blood pressure (n = 373) and to 120 micrograms/l in subjects with increased blood pressure (n = 278). After log-normal conversion of B-Pb, the linear correlation coefficient between In[B-Pb(ug/l)] and both SBP and DBP was statistically significant (r = 0.1332, p < 0.001 and r = 0.0737, p = 0.007, respectively). The linear regression coefficient was 6.8 mm Hg/In(micrograms/l) for SBP and 1.8 mm Hg/In(microgram/l) for DBP. Multiple regression analyses revealed that, after correction for body mass index (BMI), age, heart rate, skinfold thickness, serum lipids, and glucose levels; blood lead was still a significant predictor of increased SBP and DBP.(ABSTRACT TRUNCATED AT 250 WORDS)

Summary Serum ascorbic acid (SAA) levels were determined in 500 subjects aged 55–75 years participating in the New Risk Factors (NRF) Survey between March and December 1990. In the total sample mean concentration of SAA was 8.75 mg/1 (95 % confidence interval: 8.41–9.20). SAA levels of 2 mg/l or less (severe hypovitaminosis C) were found in 19 subjects (3.8 %). A further 9.6% of subjects (n=48) has SAA levels between 2 and 4 mg/l (marginal hypovitaminosis C). Subjects with hypovitaminosis C reported a significantly higher daily cigarette consumption and had higher blood lead (B-Pb) and blood cadmium (B-Cd) levels. An Inverse relationship was found between SAA levels and smoking status, body mass index (BMI) and B-Pb. Among current smokers aged 55–65 years the prevalence of hypovitaminosis C was not significantly higher than in non smokers (x 2 was 2.4 for severe and 4.7 for marginal hypovitaminosis C, respectively). On the contrary, in subjects aged 66–75 years the occurrence of hypovitaminosis C was significantly higher in smokers than in non-smokers (x 2 was 6.6 for severe and 10.3 for marginal hypovitaminosis C. respectively).

Context: Notch genes encode receptors for a signaling pathway that regulates cell growth and differentiation in various contexts, but the role of Notch signaling in thyroid follicular cells has never been fully published. Objective: The objective of the study was to characterize the expression of Notch pathway components in thyroid follicular cells and Notch signaling activities in normal and transformed thyrocytes. Design/Setting and Patients: Expression of Notch pathway components and key markers of thyrocyte differentiation was analyzed in murine and human thyroid tissues (normal and tumoral) by quantitative RT-PCR and immunohistochemistry. The effects of Notch overexpression in human thyroid cancer cells and FTRL-5 cells were explored with analysis of gene expression, proliferation assays, and experiments involving transfection of a luciferase reporter construct containing human NIS promoter regions. Results: Notch receptors are expressed during the development of murine thyrocytes, and their expression levels parallel those of thyroid differentiation markers. Notch signaling characterized also normal adult thyrocytes and is regulated by TSH. Notch pathway components are variably expressed in human normal thyroid tissue and thyroid tumors, but expression levels are clearly reduced in undifferentiated tumors. Overexpression of Notch-1 in thyroid cancer cells restores differentiation, reduces cell growth rates, and stimulates NIS expression via a direct action on the NIS promoter. Conclusion: Notch signaling is involved in the determination of thyroid cell fate and is a direct regulator of thyroid-specific gene expression. Its deregulation may contribute to the loss of differentiation associated with thyroid tumorigenesis.

Context: BRAF mutations are common in papillary thyroid carcinomas (PTCs). By affecting the expression of genes critically related to the development and differentiation of thyroid cancer, they may influence the prognosis of these tumors. Objective: Our objective was to characterize the expression of thyroid-specific genes associated with BRAF mutation in PTCs. Design/Setting and Patients: We examined the expression of key markers of thyrocyte differentiation in 56 PTCs with BRAF mutations (BRAF-mut) and 37 with wild-type BRAF (BRAF-wt). Eight samples of normal thyroid tissue were analyzed as controls. Quantitative PCR was used to measure mRNA levels for the sodium/iodide symporter (NIS), apical iodide transporter (AIT-B), thyroglobulin (Tg), thyroperoxidase (TPO), TSH receptor (TSH-R), the transcription factor PAX8, and glucose transporter type 1 (Glut1). NIS protein expression and localization was also analyzed by immunohistochemistry. Results: mRNA levels for all thyroid-specific genes were reduced in all PTCs vs. normal thyroid tissues. NIS, AIT-B, Tg, and TPO expression was significantly lower in BRAF-mut tumors than in the BRAF-wt group. Glut-1 transcript levels were increased in all PTCs, and additional increases were noted in BRAF-mut tumors. In both tumor subsets, the NIS protein that was expressed was abnormally retained in the cytoplasm. Conclusion: BRAF V600E mutation in PTCs is associated with reduced expression of key genes involved in iodine metabolism. This effect may alter the effectiveness of diagnostic and/or therapeutic use of radioiodine in BRAF-mut PTCs.

Nucleotide receptors signaling affects cell proliferation, with possible implications on tumorigenic processes. However, molecular targets and action mechanisms of the extracellular nucleotides are still poorly elucidated. We have previously shown in ARO cells that APE1/Ref-1, a transcriptional coactivator responsible for the maintenance of the cellular proliferative rate, is functionally controlled by P2-mediated signaling. Here, we demonstrate that extracellular ATP has a mitogenic effect on ARO cells, increasing ERK phosphorylation, AP1 activation, and cyclin D1 expression. Using the ATP/ADPase apyrase and the P2 receptor antagonist suramin, we show that the extracellular ATP, physiologically released by ARO cells, exerts mitogenic effects. A differential proteomic approach was used to identify molecular events associated with the ATP-induced cell proliferation. Among other proteins, Hsp90 was found upregulated upon ATP stimulation. Pretreatment with suramin completely blocked the ATP-induced Hsp90 activation, confirming the involvement of cell-surface P2 nucleotide receptors in the ATP-mediated activation of ARO cells. Treatment of proliferating ARO cells with suramin and apyrase significantly reduced the intracellular levels of Hsp90, suggesting an autocrine/paracrine mechanism of control on Hsp90 expression by extracellular ATP. The influence of Hsp90 on ATP-induced cell proliferation was also demonstrated by its specific inhibition with 17-AAG. The molecular pathway by which ATP stimulates cell proliferation was further investigated by siRNA strategies showing that Hsp90 is a target of APE1/Ref-1 functional activation. Stimulation of ARO cells with specific nucleotide receptors agonists evidenced a major involvement of P2Y1 and P2Y2 receptors in controlling the Hsp90 activation. Accordingly, these two receptors resulted significantly upregulated in sample biopsies from different thyroid tumors.

A resolutive therapy for Duchene muscular dystrophy, a severe degenerative disease of the skeletal muscle, is still lacking. Because autophagy has been shown to be crucial in clearing dysfunctional organelles and in preventing tissue damage, we investigated its pathogenic role and its suitability as a target for new therapeutic interventions in Duchenne muscular dystrophy (DMD). Here we demonstrate that autophagy is severely impaired in muscles from patients affected by DMD and mdx mice, a model of the disease, with accumulation of damaged organelles. The defect in autophagy was accompanied by persistent activation via phosphorylation of Akt, mammalian target of rapamycin (mTOR) and of the autophagy-inhibiting pathways dependent on them, including the translation-initiation factor 4E-binding protein 1 and the ribosomal protein S6, and downregulation of the autophagy-inducing genes LC3, Atg12, Gabarapl1 and Bnip3. The defective autophagy was rescued in mdx mice by long-term exposure to a low-protein diet. The treatment led to normalisation of Akt and mTOR signalling; it also reduced significantly muscle inflammation, fibrosis and myofibre damage, leading to recovery of muscle function. This study highlights novel pathogenic aspects of DMD and suggests autophagy as a new effective therapeutic target. The treatment we propose can be safely applied and immediately tested for efficacy in humans.

Context: Evidence from in vitro studies or animal models has shown that TSH affects thyrocytes by thyroid-specific expression modulation.Objective: The objective of our study was to analyze the role of TSH in human thyroid gene expression in vivo.Design/Setting: Thirty-nine normal thyroid tissues were collected at the same center.Study Subjects: Patients were divided into two groups based on serum TSH levels: 17 with normal TSH levels (1–4 mU/liter; group 1) and 22 with TSH levels below 0.5 mU/liter (group 2).Intervention: Group 2 underwent thyroidectomy after suppressive l-T4 therapy.Main Outcome Measures: mRNA levels of thyroid genes such as sodium/iodide symporter (NIS), apical iodide transporter, pendrin, thyroglobulin, thyroperoxidase, TSH receptor, paired box transcription factor 8, and thyroid transcription factor-1 were evaluated by quantitative PCR.Results: The reduction of TSH stimulation causes decreases in NIS and apical iodide transporter gene expression in normal tissues and more limited reductions in thyroglobulin, thyroperoxidase, and paired box transcription factor 8, but it has no significant effect on TSH receptor, pendrin, or thyroid transcription factor-1. Comparison of NIS levels in normal and nodular tissues from the same patient confirmed that it is differentially expressed in nodules only in the presence of normal TSH (P < 0.01). In patients with suppressed TSH, nodular NIS levels were similar to those in normal tissues.Conclusions: Our data represent the first demonstration in human thyroid tissues that TSH contributes to the regulation of thyrocyte differentiation by modulating thyroid gene levels. It exerts a particularly important effect on the transcription of NIS, which becomes very low after prolonged TSH suppression.

Purpose: Many years ago, diphtheria toxin (DT) showed antitumor activity in mice and in humans, but it was unclear whether this depended on the toxicity of the molecule only or on its strong inflammatory-immunological property as well. To deal with this open question, we planned to treat a group of cancer patients with cross-reacting material 197 (CRM197). CRM197 is a nontoxic mutant of DT that shares the immunological properties of the native molecule and its ability to bind to heparin-binding epidermal growth factor (HB-EGF), the specific cell-membrane receptor for DT that is often overexpressed in cancer. Methods: 25 outpatients with various advanced tumors who were refractory to standard therapies (23 subjects) or had refused, in whole or in part, conventional therapies (2 subjects) were treated with CRM197 injected subcutaneously in the abdominal wall, on alternate days, for 6 days. Three different dosages (1.7, 2.6, or 3.5 mg/day) were used according to the patient’s degree of immunological reactivity to DT/CRM197 (none, moderate, or high). Results: After the first administration of CRM197, a significant increase in the number of circulating neutrophils and in the serum level of TNF-α was detected. Toxicities were minimal. Only patients with delayed-type hypersensitivity to DT/CRM197 had irritating skin reactions in the injection sites and a flu-like syndrome with fever. Pharmacokinetics showed a mean peak concentration (12.7 ng/ml) 12 h after the first injection and a mean half-life of 18.1 h. There were two complete and one partial responses (metastatic breast carcinoma, neuroblastoma, and metastatic breast carcinoma) lasting 4, 45+, and 15 months, respectively. Six cases of stable disease, lasting from 1 to 15 months, were also recorded. Conclusions: CRM197 injected subcutaneously elicited an inflammatory-immunological reaction, caused tolerable toxicities, was absorbed to a good extent into the circulatory system, and exerted some degree of biological antitumor activity. A possible role of neutrophils and TNF-α in the mode of action of the molecule is hypothesized.

Expression of Concern to: Cell Death and Disease (2012) 3, e418; doi:10.1038/cddis.2012.159; published online 15 November 2012 Cell Death and Disease is publishing an Expression of Concern regarding the above-mentioned publication (PMID 23152054). This paper was first corrected on 7 March 2013 (CellDeath and Disease (2013) 4, e529) after the authors noticed that lanes displaying the actin bands in Figures 1, 5b and Supplementary Figure 1a were not the appropriate actin loading controls.

The association of serum selenium (S-Se) with selected cardiovascular risk factors has been studied in 3404 (1520 men, 1884 women) of 4699 subjects aged 20-73 years, who underwent a comprehensive examination between April 1986 and December 1987 within the framework of the Di. S.Co. Research Project of the National Research Council. Mean S-Se concentrations were 1.163 (SD 0.170) μmol/liter, 1.156 (0.163) μmol/liter, and 1.171 (0.179) μmol/liter in the total group, and female and male subjects, respectively. The difference by sex was statistically significant (F = 6.97, P = 0.0083). In male subjects S-Se levels were inversely associated to age (simple correlation coefficient, r = − 0.2135, P < 0.001), alcohol consumption (Alcohol, r = − 0.0688, P < 0.01), smoking habit (Smoke, r = − 0.0663, P < 0.01), body mass index (BMI, r = − 0.0596, P < 0.02), and lognormal transformation of triglycerides (Ln-Trig, r = − 0.0767, P < 0.005). In multiple regression analysis Age, Smoke, Ln-Trig, and Alcohol remained significantly and inversely related to S-Se; a significant direct association of S-Se to high-density lipoprotein cholesterol (HDL) and nonHDL cholesterol (non-HDL) was also pointed out. In female subjects, S-Se was directly related to HDL (r = 0.1436, P < 0.001) and non-HDL (r = 0.0967, P < 0.001). In multiple regression analysis S-Se was directly related to both HDL and non-HDL and an inverse significant association of S-Se to Age and Alcohol was evidenced. In multivariate regression models analyzing systolic (SBP), diastolic (DBP), and mean blood pressure (MBP) as dependent variables, S-Se was a weak significant positive predictor in male but not in female subjects

An occupational group living in Rome and composed of 3,007 men aged 46 to 65 years who were free from previous major coronary events was screened for a number of coronary risk factors and then followed up for 10 years. In all, 107 coronary deaths occurred in 10 years. There was a positive relation between coronary death rate and increasing levels of triglycerides, but the difference between the extreme quintile classes was not any more significant after adjustment for cholesterol levels. A cross-classification involving low-density lipoprotein (LDL) cholesterol, high-density lipoprotein (HDL) cholesterol and triglyceride levels showed a higher coronary artery disease (CAD) mortality in subjects with a higher LDL cholesterol/HDL cholesterol ratio and higher triglycerides. However, the excess risk in this subgroup was largely explained by the mean levels of total cholesterol. The multivariate proportional-hazards Cox model with coronary deaths as the end point, and age and triglycerides as predictors, produced a significant coefficient for triglycerides that became nonsignificant when other lipids alone or in combination (total, HDL and LDL cholesterol and some of their ratio) were fed into the model as further covariates. It is concluded that there is no evidence of an independent role of triglycerides in the prediction of coronary deaths.

Blood lead (B-Pb) levels were determined in 1802 out of 1856 non-occupationally exposed men aged 55-75 years living in the Rome area who participated, between 1989 and 1990, in an epidemiological survey for coronary heart disease (New Risk Factors Project). The median B-Pb level was 113 micrograms/l (10th-90th centiles: 74-180 micrograms/l) and only 0.7 per cent (n = 14) of the subjects had B-Pb values higher than 300 micrograms/l. B-Pb levels were significantly and positively associated to alcohol consumption. Moderate and heavy drinkers had median B-Pb level of 143 micrograms/l (10th-90th centiles: 92-233) and 165 micrograms/l (10th-90th centiles: 102-285) respectively, whereas non-drinkers had a median B-Pb level of 96 micrograms/l (10th-90th centiles: 66-143). The influence of smoking habits was less relevant. Subjects who never smoked and subjects smoking more than 20 cigarettes daily had median B-Pb levels of 103 and 133 micrograms/l, respectively. Individuals classified as habitual car-drivers had slightly higher Pb levels than non-drivers. Subjects classified as manual workers had higher B-Pb levels in comparison with non-manual workers and retired subjects. B-Pb levels were directly related to HDL-cholesterol (HDL-C, r = 0.2252) and gamma-glutamyltransferase (gamma-GT, r = 0.2207) serum levels. The alleged alcohol consumption was more related to B-Pb level (r = 0.3848) than to serum level of HDL-C (r = 0.2474) or gamma-GT (r = 0.2469). A significant correlation (r = 0.2409) also existed between B-Pb and blood cadmium levels (B-Cd). Subjects with a low Gaensler ratio, an index of respiratory function, had higher B-Pb levels. In multiple regression analyses alcohol intake was the most important predictor of B-Pb level, explaining more (14.27%) of the total variance than did B-Cd (4.98%), HDL-C (1.89%), driving habits (1.46%), gamma-GT (1.09%), skinfold thickness (0.96%), and Gaensler index (0.38%). The risk ratio of having B-Pb level higher than 180 micrograms/l (90th centile of B-Pb distribution in our subjects) was 5.3 (95% CI: 2.7-10.4) for drinkers versus non-drinkers and 1.9 (95% CI: 1.2-3.1) for current smokers versus subjects who had never smoked. B-Pb was, at least in our subjects, a more specific and sensitive objective index of alcohol consumption than gamma-GT and HDL-C.(ABSTRACT TRUNCATED AT 400 WORDS)

The aim of this study was to determine, in a population of Italian adolescents, the association of serum selenium levels with precursors of biochemical and anthropometric variables known as being among the major risk factors for cardiovascular diseases in the adult population. The following measurements were taken in a school sample of 627 adolescents (aged 12-13 years); serum selenium, total cholesterol, high density lipoprotein cholesterol, non-HDL cholesterol, height, weight, body mass index, systolic blood pressure and diastolic blood pressure. The serum selenium levels were slightly higher in males (83.1 +/- 10.1 micrograms/l) than in females (81.7 +/- 11.0 micrograms/l), but the difference was not statistically significant. Serum selenium was positively correlated with total cholesterol, diastolic blood pressure and HDL cholesterol in both sexes; moreover it was positively correlated with non-HDL cholesterol and negatively correlated with height in males only.