Visualization of transcription sites at the electron microscope

In order to localize at EM level the sites of transcription of both pre-mRNA and pre-rRNA, we have detected the DNA/RNA hybrid molecules and m3Gcapped structures by means of specific antibodies after short bromo-uridine (BrU) incorporation. In addition, the sections have been stained by a selective RNA stain, terbium citrate. Our data indicate that perichromatin fibrils incorporate BrU and are labeled by the anti-hybrid probe; this supports the idea that they are the pre-mRNA transcription sites. On the contrary, interchromatin granules do not incorporate BrU after short pulses and are not labeled by the anti-hybrid probe. Concerning the nucleolus, anti-hybrid and anti-BrdU antibodies colocalize only on the dense fibrillar component, suggesting that this is the site of rRNA transcription. Interestingly, the dense fibrillar component and the granular component, after specific RNA staining, show remarkable structural similarities, both containing fibrogranular RNA structures

Locally increased P-glycoprotein function in major depression: a PET study with [C-11]verapamil as a probe for P-glycoprotein function in the blood-brain barrier

The aetiology of depressive disorder remains unknown, although genetic susceptibility and exposure to neurotoxins are currently being discussed as possible contributors to this disorder. In normal circumstances, the brain is protected against bloodborne toxic influences by the blood-brain barrier, which includes the molecular efflux pump P-glycoprotein (P-gp) in the vessel wall of brain capillaries. We hypothesized that P-gp function in the blood-brain barrier is changed in patients with major depression. Positron emission tomography Was used to measure brain uptake of [C-11]verapamil, which is normally expelled from the brain by P-gp. Cerebral Volume of distribution (V-T) of [C-11]verapamil was used as a measure of P-gp function. Both region-of-interest (ROI) analysis and voxel analysis using statistical parametric mapping (SPM2) were performed to assess regional brain P-gp function. We found that patients with a major depressive episode, using antidepressants, compared to health), controls showed a significant decrease of [C-11]verapamil uptake in different areas throughout the brain, in particular in frontal and temporal regions. The decreased [C-11]verapamil uptake correlates with an increased function of the P-gp protein and may be related to chronic use of psychotropic drugs, Our results may explain why treatment-resistant depression can develop

Long-Term Effects of a Classic Ketogenic Diet on Ghrelin and Leptin Concentration : a 12-Month Prospective Study in a Cohort of Italian Children and Adults with GLUT1-Deficiency Syndrome and Drug Resistant Epilepsy

The classical ketogenic diet (cKD) is an isocaloric, high fat, very low-carbohydrate diet that induces ketosis, strongly influencing leptin and ghrelin regulation. However, not enough is known about the impact of a long-term cKD. This study evaluated the effects of a 12-month cKD on ghrelin and leptin concentrations in children, adolescents and adults affected by the GLUT1-Deficiency Syndrome or drug resistant epilepsy (DRE). We also investigated the relationship between the nutritional status, body composition and ghrelin and leptin variations. We carried out a longitudinal study on 30 patients: Twenty-five children and adolescents (15 females, 8 \ub1 4 years), and five adults (two females, 34 \ub1 16 years). After 12-monoths cKD, there were no significant changes in ghrelin and leptin, or in the nutritional status, body fat, glucose and lipid profiles. However, a slight height z-score reduction (from -0.603 \ub1 1.178 to -0.953 \ub1 1.354, p 64 0.001) and a drop in fasting insulin occurred. We found no correlations between ghrelin changes and nutritional status and body composition, whereas leptin changes correlated positively with variations in the weight z-score and body fat (\u3c1 = 0.4534, p = 0.0341; \u3c1 = 0.5901, p = 0.0135; respectively). These results suggest that a long-term cKD does not change ghrelin and leptin concentrations independently of age and neurological condition

To treat or not to treat: comparison of different criteria used to determine whether weight loss is to be recommended

Background: Excess body fat is a major risk factor for disease primarily due to its endocrine activity. In recent years several criteria have been introduced to evaluate this factor. Nevertheless, treatment need is currently assessed only on the basis of an individual's Body Mass Index (BMI), calculated as body weight (in kg) divided by height in m2. The aim of our study was to determine whether application of the BMI, compared to adiposity-based criteria, results in underestimation of the number of subjects needing lifestyle intervention. Methods: We compared treatment need based on BMI classification with four adiposity-based criteria: percentage body fat (%BF), considered both alone and in relation to metabolic syndrome risk (MS), waist circumference (WC), as an index of abdominal fat, and Body Fat Mass Index (BFMI, calculated as fat mass in kg divided by height in m2) in 63 volunteers (23 men and 40 women, aged 20 – 65 years). Results: According to the classification based on BMI, 6.3% of subjects were underweight, 52.4% were normal weight, 30.2% were overweight, and 11.1% were obese. Agreement between the BMI categories and the other classification criteria categories varied; the most notable discrepancy emerged in the underweight and overweight categories. BMI compared to almost all of the other adiposity-based criteria, identified a lower percentage of subjects for whom treatment would be recommended. In particular, the proportion of subjects for whom clinicians would strongly recommend weight loss on the basis of their BMI (11.1%) was significantly lower than those identified according to WC (25.4%, p = 0.004), %BF (28.6%, p = 0.003), and MS (33.9%, p = 0.002). Conclusion: The use of the BMI alone, as opposed to an assessment based on body composition, to identify individuals needing lifestyle intervention may lead to unfortunate misclassifications. Population-specific data on the relationships between body composition, morbidity, and mortality are needed to improve the diagnosis and treatment of at-risk individual

Evaluation of nutritional status in children with refractory epilepsy

BACKGROUND: children affected by refractory epilepsy could be at risk of malnutrition because of feeding difficulties (anorexia, chewing, swallowing difficulties or vomiting) and chronic use of anticonvulsants, which may affect food intake and energy metabolism. Moreover, their energy requirement may be changed as their disabilities would impede normal daily activities. The aim of the present study was to evaluate nutritional status, energy metabolism and food intake in children with refractory epilepsy. METHODS: 17 children with refractory epilepsy (13 boys and 4 girls; mean age 9 ± 3,2 years; Body Mass Index 15,7 ± 3,6) underwent an anthropometric assessment, body composition evaluation by dual-energy X-ray absorptiometry, detailed dietetic survey and measurement of resting energy expenditure by indirect calorimetry. Weight-for-age, height-for-age (stunting) and weight-for-height (wasting) were estimated compared to those of a reference population of the same age. RESULTS: 40% of children were malnourished and 24% were wasted. The nutritional status was worse in the more disabled children. Dietary intake resulted unbalanced (18%, 39%, 43% of total daily energy intake derived respectively from protein, lipid and carbohydrate). Adequacy index [nutrient daily intake/recommended allowance (RDA) × 100] was < 60% for calcium iron and zinc. CONCLUSION: many children with refractory epilepsy would benefit from individual nutritional assessment and management as part of their overall care

Selective Survival and Maturation of Adult-Born Dentate Granule Cells Expressing the Immediate Early Gene Arc/Arg3.1

Progenitor cells in the adult dentate gyrus provide a constant supply of neuronal precursors, yet only a small fraction of these cells survive and develop into mature dentate granule cells (DGCs). A major challenge of current research is thus to understand the stringent selection process that governs the maturation and functional integration of adult-born DGCs. In mature DGCs, high-frequency stimulation (HFS) of the perforant path input elicits robust expression of the immediate early gene Arc/Arg3.1, trafficking of its mRNA to dendrites, and local synthesis of the protein necessary for consolidation of long-term potentiation (LTP). Given the synaptic commitment inherent in LTP consolidation, we considered that HFS-evoked expression of Arc could be used to timemap the functional integration of newborn DGCs. Dividing cells were birthmarked by BrdU-labeling at 1, 7, 14, 21, or 28 days prior to induction of LTP and expression of Arc was examined by confocal microscopy. Contrary to expectation, LTP did not induce Arc expression in newborn cells at any age, suggesting they might be refractory to synaptically-evoked Arc expression for at least one month. Importantly, however, spontaneous expression of Arc was detected in BrdU-labeled cells and strongly associated with the survival and maturation of NeuN-positive DGCs. Moreover, Arc expression at the earliest ages (1 and 7 days), clearly precedes the formation of glutamatergic synapses on new neurons. These results suggest an unexpected early role for Arc in adult-born DGCs, distinct from its functions in LTP, LTD, and homeostatic synaptic plasticity

Stress, ageing and their influence on functional, cellular and molecular aspects of the immune system

The immune response is essential for keeping an organism healthy and for defending it from different types of pathogens. It is a complex system that consists of a large number of components performing different functions. The adequate and controlled interaction between these components is necessary for a robust and strong immune response. There are, however, many factors that interfere with the way the immune response functions. Stress and ageing now consistently appear in the literature as factors that act upon the immune system in the way that is often damaging. This review focuses on the role of stress and ageing in altering the robustness of the immune response first separately, and then simultaneously, discussing the effects that emerge from their interplay. The special focus is on the psychological stress and the impact that it has at different levels, from the whole system to the individual molecules, resulting in consequences for physical health

Visualization of transcription sites at the electron microscope

6 p.-5 fig.In order to localize at EM level the sites of transcription of both pre-mRNA and pre-rRNA, we have detected the DNA/RNA hybrid molecules and m3Gcapped structures by means of specific antibodies after short bromo-uridine (BrU) incorporation. In addition, the sections have been stained by a selective RNA stain, terbium citrate. Our data indicate that perichromatin fibrils incorporate BrU and are labeled by the anti-hybrid probe; this supports the idea that they are the pre-mRNA transcription sites. On the contrary, interchromatin granules do not incorporate BrU after short pulses and are not labeled by the anti-hybrid probe. Concerning the nucleolus, anti-hybrid and anti-BrdU antibodies colocalize only on the dense fibrillar component, suggesting that this is the site of rRNA transcription. Interestingly, the dense fibrillar component and the granular component, after specific RNA staining, show remarkable structural similarities, both containing fibrogranular RNA structures.This work was supported by grants from Italian M.U.R.S.T. (Cofin 2002) and Fondo di Ateneo per la Ricerca.Peer reviewe