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Rett syndrome (RTT) is a severe genetic disorder resulting from mutations in the X-linked methyl-CpG-binding protein 2 ( MECP2 ) gene. Recently, a zebrafish carrying a mecp2 -null mutation has been developed with the resulting phenotypes exhibiting defective sensory and thigmotactic responses, and abnormal motor behavior reminiscent of the human disease. Here, we performed a proteomic analysis to examine protein expression changes in mecp2 -null vs. wild-type larvae and adult zebrafish. We found a total of 20 proteins differentially expressed between wild-type and mutant zebrafish, suggesting skeletal and cardiac muscle functional defects, a stunted glycolysis and depleted energy availability. This molecular evidence is directly linked to the mecp2 -null zebrafish observed phenotype. In addition, we identified changes in expression of proteins critical for a proper redox balance, suggesting an enhanced oxidative stress, a phenomenon also documented in human patients and RTT murine models. The molecular alterations observed in the mecp2- null zebrafish expand our knowledge on the molecular cascade of events that lead to the RTT phenotype. Biological significance We performed a proteomic study of a non-mammalian vertebrate model (zebrafish, Danio rerio ) for Rett syndrome (RTT) at larval and adult stages of development. Our results reveal major protein expression changes pointing out to defects in energy metabolism, redox status imbalance, and muscle function, both skeletal and cardiac. Our molecular analysis grants the mecp2 -null zebrafish as a valuable RTT model, triggering new research approaches for a better understanding of the RTT pathogenesis and phenotype expression. This non-mammalian vertebrate model of RTT strongly suggests a broad impact of Mecp2 dysfunction.

Rett Syndrome (RTT) is a rare X-linked neurodevelopmental disorder which affects about 1: 10000 live births. In >95% of subjects RTT is caused by a mutation in Methyl-CpG binding protein-2 (MECP2) gene, which encodes for a transcription regulator with pleiotropic genetic/epigenetic activities. The molecular mechanisms underscoring the phenotypic alteration of RTT are largely unknown and this has impaired the development of therapeutic approaches to alleviate signs and symptoms during disease progression. A defective proteasome biogenesis into two skin primary fibroblasts isolated from RTT subjects harbouring non-sense (early-truncating) MeCP2 mutations (i.e., R190fs and R255X) is herewith reported. Proteasome is the proteolytic machinery of Ubiquitin Proteasome System (UPS), a pathway of overwhelming relevance for post-mitotic cells metabolism. Molecular, transcription and proteomic analyses indicate that MeCP2 mutations down-regulate the expression of one proteasome subunit, ?7, and of two chaperones, PAC1 and PAC2, which bind each other in the earliest step of proteasome biogenesis. Furthermore, this molecular alteration recapitulates in neuron-like SH-SY5Y cells upon silencing of MeCP2 expression, envisaging a general significance of this transcription regulator in proteasome biogenesis.

International audience; Rett syndrome (RTT) is a leading cause of severe intellectual disability in females, caused by de novo loss-of function mutations in the X-linked methyl-CpG binding protein 2 (MECP2). To better investigate RTT disease progression/pathogenesis animal models of Mecp2 deficiency have been developed. Here, Mecp2 mouse models are employed to investigate the role of protein patterns in RTT. A proteome analysis was carried out in brain tissue from i) Mecp2 deficient mice at the pre-symptomatic and symptomatic stages and, ii) mice in which the disease phenotype was reversed by Mecp2 reactivation. Several proteins were shown to be differentially expressed in the pre-symptomatic (n = 18) and symptomatic (n = 20) mice. Mecp2 brain reactivated mice showed wild-type comparable levels of expression for twelve proteins, mainly related to proteostasis (n = 4) and energy metabolic pathways (n = 4). The remaining ones were found to be involved in redox homeostasis (n = 2), nitric oxide regulation (n = 1), neurodevelopment (n = 1). Ten out of twelve proteins were newly linked to Mecp2 deficiency. Our study sheds light on the relevance of the protein-regulation of main physiological process in the complex mechanisms leading from Mecp2 mutation to the RTT clinical phenotype.SignificanceWe performed a proteomic study of a Mecp2stop/y mouse model for Rett syndrome (RTT) at the pre-symptomatic and symptomatic Mecp2 deficient mice stage and for the brain specific reactivated Mecp2 model. Our results reveal major protein expression changes pointing out to defects in proteostasis or energy metabolic pathways other than, to a lesser extent, in redox homeostasis, nitric oxide regulation or neurodevelopment. The Mecp2 mouse rescued model provides the possibility to select target proteins more susceptible to the Mecp2 gene mutation, potential and promising therapeutical targets.

International audience; Krabbe disease (KD) is a rare and devastating pediatric leukodystrophy caused by mutations in the galactocerebrosidase (GALC) gene. The disease leads to impaired myelin formation and extensive myelin damage in the brain. Oxidative stress is implicated in the pathogenesis of KD but insofar few information is available. The gray and white matter of the brain are rich in docosahexaenoic acid and adrenic acid respectively and under non-enzymatic oxidative stress, release isoprostanoids, i.e. F4-neuroprostanes (F4-NeuroPs) and F2-dihomo-isoprostanes (F2-dihomo-IsoPs). In this study, the formation of isoprostanoids in brain tissue was investigated in a well-established KD mouse model (twitcher) that recapitulates the human pathology. According to the genotype determinations, three groups of mice were selected: wild-type control mice (n = 13), heterozygotes mice (carriers of GALC mutations, n = 14) and homozygous twitcher mice (n = 13). Measurement of F2-dihomo-IsoP and F4-NeuroP levels were performed on whole brain tissue obtained at day 15 and day 35 of the life cycle. Brain isoprostanoid levels were significantly higher in the twitcher mice compared to the heterozygous and wild-type control mice. However, F2-dihomo-IsoP and F4-NeuroP levels did not differ in brain of day 15 compared to day 35 of the heterozygote mice. Interestingly, isoprostanoid levels were proportionally enhanced with disease severity (F2-dihomo-IsoPs, rho = 0.54; F4-NeuroPs, rho = 0.581; P values ≤ 0.05; n = 13). Our findings are the first to show the key role of polyunsaturated fatty acid oxidative damage to brain grey and white matter in the pathogenesis and progression of KD. This shed new insights on the biochemical indexes of KD progression, and potentially provide information for novel therapeutic targets.

Neuroprostanes, a family of non-enzymatic metabolites of the docosahexaenoic acid, have been suggested as potential biomarkers for neurological diseases. Objective biological markers are strongly needed in Rett syndrome (RTT), which is a progressive X-linked neurodevelopmental disorder that is mainly caused by mutations in the methyl-CpG binding protein 2 (MECP2) gene with a predominant multisystemic phenotype. The aim of the study is to assess a possible association between MECP2 mutations or RTT disease progression and plasma levels of 4(RS)-4-F4t-neuroprostane (4-F4t-NeuroP) and 10(RS)-10-F4t-neuroprostane (10-F4t-NeuroP) in typical RTT patients with proven MECP2 gene mutation. Clinical severity and disease progression were assessed using the Rett clinical severity scale (RCSS) in n = 77 RTT patients. The 4-F4t-NeuroP and 10-F4t-NeuroP molecules were totally synthesized and used to identify the contents of the plasma of the patients. Neuroprostane levels were related to MECP2 mutation category (i.e., early truncating, gene deletion, late truncating, and missense), specific hotspot mutations (i.e., R106W, R133C, R168X, R255X, R270X, R294X, R306C, and T158M), and disease stage (II through IV). Circulating 4-F4t-NeuroP and 10-F4t-NeuroP were significantly related to (i) the type of MECP2 mutations where higher levels were associated to gene deletions (p ≤ 0.001), (ii) severity of common hotspot MECP2 mutation (large deletions, R168X, R255X, and R270X), (iii) disease stage, where higher concentrations were observed at stage II (p ≤ 0.002), and (iv) deficiency in walking (p ≤ 0.0003). This study indicates the biological significance of 4-F4t-NeuroP and 10-F4t-NeuroP as promising molecules to mark the disease progression and potentially gauge genotype–phenotype associations in RTT.

Purpose A role for inflammation and oxidative stress is reported in autism spectrum disorders (ASDs). Here, we tested possible changes in expression and/or oxidative status for plasma proteins in subjects with ASDs. Experimental design To evaluate protein expression and protein adducts of lipid peroxidation-derived aldehyde, analysis of plasma proteins was performed in 30 subjects with ASDs and compared with 30 healthy controls with typical development, using a proteomic approach. Results Significant changes were evidenced for a total of 12 proteins. Of these, ten were identified as proteins involved in the acute inflammatory response including alpha-2-macroglobulin, alpha-1-antitrypsin, haptoglobin, fibrinogen, serum transferrin, prealbumin, apolipoprotein A-I apolipoprotein A-IV, apolipoprotein J, and serum albumin. In addition, significant changes occurred for two immunoglobulins alpha and gamma chains. Conclusions and clinical relevance Our present data indicate that an inflammatory response, coupled with increased lipid peroxidation, is present in subjects with ASDs. This information can provide new insight into the identification of potential plasma protein biomarkers in autism.

Neurological disorders can be associated with protein glycosylation abnormalities. Rett syndrome is a devastating genetic brain disorder, mainly caused by de novo loss-of-function mutations in the methylCpG binding protein 2 (MECP2) gene. Although its pathogenesis appears to be closely associated with a redox imbalance, no information on glycosylation is available. Glycoprotein detection strategies (i.e., lectin-blotting) were applied to identify target glycosylation changes in the whole brain of Mecp2 mutant murine models of the disease. Remarkable glycosylation pattern changes for a peculiar 50 kDa protein, i.e., the N-linked brain nucleotide pyrophosphatase-5 were evidenced, with decreased N-glycosylation in the presymptomatic and symptomatic mutant mice. Glycosylation changes were rescued by selected brain Mecp2 reactivation. Our findings indicate that there is a causal link between the amount of Mecp2 and the N-glycosylation of NPP-5. (C) 2015 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved.

International audience; Isoprostanoids are a large family of compounds derived from non-enzymatic oxidation of polyunsaturated fatty acids (PUFAs). Unlike other oxidative stress biomarkers, they provide unique information on the precursor of the targeted PUFA. Although they were discovered about a quarter of century ago, the knowledge on the role of key isoprostanoids in the pathogenesis of experimental and human disease models remains limited. This is mainly due to the limited availability of highly purified molecules to be used as a reference standard in the identification of biological samples. The accurate knowledge on their biological relevance is the critical step that could be translated from some mere technical/industrial advances into a reliable biological disease marker which is helpful in deciphering the oxidative stress puzzle related to neurological disorders. Recent research indicates the value of isoprostanoids in predicting the clinical presentation and evolution of the neurological diseases. This review focuses on the relevance of isoprostanoids as mediators and potential biomarkers in neurological diseases, a heterogeneous family ranging from rare brain diseases to major health conditions that could have worldwide socioeconomic impact in the health sector. The current challenge is to identify the preferential biochemical pathways that actually follow the oxidative reactions in the neurological diseases and the consequence of the specific isoprostanes in the underlying pathogenic mechanisms.

F4-neuroprostanes (F4-NeuroPs) are non-enzymatic oxidized products derived from docosahexaenoic acid (DHA) and are suggested to be oxidative damage biomarkers of neurological diseases. However, 128 isomers can be formed from DHA oxidation and among them, 4(RS)−4-F4t-NeuroP (4-F4t-NeuroP) and 10(RS)−10-F4t-NeuroP (10-F4t-NeuroP) are the most studied. Here, we report the identification and the clinical relevance of 4-F4t-NeuroP and 10-F4t-NeuroP in plasma of four different neurological diseases, including multiple sclerosis (MS), autism spectrum disorders (ASD), Rett syndrome (RTT), and Down syndrome (DS). The identification and the optimization of the method were carried out by gas chromatography/negative-ion chemical ionization tandem mass spectrometry (GC/NICI-MS/MS) using chemically synthesized 4-F4t-NeuroP and 10-F4t-NeuroP standards and in oxidized DHA liposome. Both 4-F4t-NeuroP and 10-F4t-NeuroP were detectable in all plasma samples from MS (n = 16), DS (n = 16), ASD (n = 9) and RTT (n = 20) patients. While plasma 10-F4t-NeuroP content was significantly higher in patients of all diseases as compared to age and gender matched healthy control subjects (n = 61), 4-F4t-NeuroP levels were significantly higher in MS and RTT as compared to healthy controls. Significant positive relationships were observed between relative disease severity and 4-F4t-NeuroP levels (r = 0.469, P

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Herd immunity towards measles, one of the 20 most lethal diseases in human history, has been recently challenged on a global scale. Despite a missing causal relationship, vaccine fear has triggered a global anti vaccine movement. We investigated i) the extent of the vaccination-autism false belief in a selected Italian population from two geographical areas with and without an ongoing epidemics for a potentially vaccination-preventable infectious disease (Neisseria meningitidis, groups C and B); ii) the corresponding information source; and iii) the belief in a possible global conspiracy. Four different population sub-categories (I-general population; II-parents of autistic children; III-paramedics; IV-physicians, biologists and pharmacists; n=424) were administered anonymous questionnaires. A total of 30.1% of the general population and the 54.5% of autism parents participants believed in a vaccine-autism relationship (P

Rett syndrome (RTT) is a rare neurodevelopmental disorder usually caused by mutations in the X-linked gene methyl-CpG-binding protein 2 (MECP2). Several Mecp2 mutant mouse lines have been developed recapitulating part of the clinical features. In particular, Mecp2-308 female heterozygous mice, bearing a truncating mutation, are a validated model of the disease. While recent data suggest a role for inflammation in RTT, little information on the inflammatory status in murine models of the disease is available. Here, we investigated the inflammatory status by proteomic 2-DE/MALDI-ToF/ToF analyses in symptomatic Mecp2-308 female mice. Ten differentially expressed proteins were evidenced in the Mecp2-308 mutated plasma proteome. In particular, 5 positive acute-phase response (APR) proteins increased (i.e., kininogen-1, alpha-fetoprotein, mannose-binding protein C, alpha-1-antitrypsin, and alpha-2-macroglobulin), and 3 negative APR reactants were decreased (i.e., serotransferrin, albumin, and apolipoprotein A1). CD5 antigen-like and vitamin D-binding protein, two proteins strictly related to inflammation, were also changed. These results indicate for the first time a persistent unresolved inflammation of unknown origin in the Mecp2-308 mouse model.

A consistent fraction of published data on scientific journals is not reproducible mainly due to insufficient knowledge of statistical methods. Here, we discuss on the use of proper statistical tools in biomedical research and statistical pitfalls potentially undermining the scientific validity of published data. Apart from unaware errors, a growing concern exists regarding data fabrication and scientific misconduct. Indeed, the social impact of false scientific data can be largely unpredictable and devastating, as shown by the worldwide dramatic effects on vaccinations coverage following a retracted paper published on a highly authoritative medical journal. Unfortunately, statistics shows a quite limited power in detecting false science, although a few statistical tools, such as the Benford’s law, are known. Taken together, statistics in biomedical sciences i) is a powerful tool to interpret experimental data; ii) has limited power in detecting false science; and iii) first and foremost, is not the result of a simple “click of a mouse”, but should be the result of accurate research planning by experienced and knowledgeable users.

Rett syndrome (RTT) is a severe genetic disorder resulting from mutations in the X-linked methyl-CpG-binding protein 2 (MECP2) gene. Recently, a zebrafish carrying a mecp2-null mutation has been developed with the resulting phenotypes exhibiting defective sensory and thigmotactic responses, and abnormal motor behavior reminiscent of the human disease. Here, we performed a proteomic analysis to examine protein expression changes in mecp2-null vs. wild-type larvae and adult zebrafish. We found a total of 20 proteins differentially expressed between wild-type and mutant zebrafish, suggesting skeletal and cardiac muscle functional defects, a stunted glycolysis and depleted energy availability. This molecular evidence is directly linked to the mecp2-null zebrafish observed phenotype. In addition, we identified changes in expression of proteins critical for a proper redox balance, suggesting an enhanced oxidative stress, a phenomenon also documented in human patients and RTT murine models. The molecular alterations observed in the mecp2-null zebrafish expand our knowledge on the molecular cascade of events that lead to the RTT phenotype.We performed a proteomic study of a non-mammalian vertebrate model (zebrafish, Danio rerio) for Rett syndrome (RTT) at larval and adult stages of development. Our results reveal major protein expression changes pointing out to defects in energy metabolism, redox status imbalance, and muscle function, both skeletal and cardiac. Our molecular analysis grants the mecp2-null zebrafish as a valuable RTT model, triggering new research approaches for a better understanding of the RTT pathogenesis and phenotype expression. This non-mammalian vertebrate model of RTT strongly suggests a broad impact of Mecp2 dysfunction.

Mutations in the cyclin-dependent kinase-like 5 gene cause a clinical variant of Rett syndrome (CDKL5-RTT). A role for the acute-phase response (APR) is emerging in typical RTT caused by methyl-CpG-binding protein 2 gene mutations (MECP2-RTT). No information is, to date, available on the inflammatory protein response in CDKL5-RTT. We evaluated, for the first time, the APR protein response in CDKL5-RTT. Protein patterns in albumin- and IgG-depleted plasma proteome from CDKL5-RTT patients were evaluated by two-dimensional gel electrophoresis/mass spectrometry. The resulting data were related to circulating cytokines and compared to healthy controls or MECP2-RTT patients. The effects of omega-3 polyunsaturated fatty acids (ω-3 PUFAs) were evaluated. CDKL5-RTT mutations resulted in a subclinical attenuated inflammation, specifically characterized by an overexpression of the complement component C3 and CD5 antigen-like, both strictly related to the inflammatory response. Cytokine dysregulation featuring a bulk increase of anti-inflammatory cytokines, predominantly IL-10, could explain the unchanged erythrocyte sedimentation rate and atypical features of inflammation in CDKL5-RTT. Omega-3 PUFAs were able to counterbalance the pro-inflammatory status. For the first time, we revealed a subclinical smouldering inflammation pattern in CDKL5-RTT consisting in the coexistence of an atypical APR coupled with a dysregulated cytokine response.

Objective The aim of this study was to test the value of F2-isoprostane (F2-IsoP) levels in the pericoronal gingiva as a potential predictor of oral disability after mandibular third molar (M3M) surgery. Study design Thirteen patients (5 male, 8 female) with partially impacted M3M were enrolled. Pericoronal soft tissues histology was obtained, gingival F2-IsoPs were measured by gas chromatography/negative-ion chemical ionization tandem mass spectrometry, and health-related quality of life (HRQOL) scores were assessed. Results A variable degree of pericoronitis was evident before the M3M surgery, with a mean level of pericoronal F2-IsoPs of 634.8 ± 193.5 pg/mg of tissue. A significant positive correlation between pericoronal F2-IsoPs and HRQOL was observed in male (rs = 0.440; P = .0011), but not female (rs = −0.062; P = .5124) patients. Conclusions Levels of F2-IsoPs in the pericoronal soft tissues are correlated with the degree of oral disability after M3Ms surgery in male, but not female, patients.

The main cause of Rett syndrome (RTT), a pervasive development disorder almost exclusively affecting females, is a mutation in the methyl-CpG binding protein 2 (MeCP2) gene. To date, no cure for RTT exists, although disease reversibility has been demonstrated in animal models. Emerging evidence from our and other laboratories indicates a potential role of oxidative stress (OS) in RTT. This review examines the current state of the knowledge on the role of OS in explaining the natural history, genotype-phenotype correlation, and clinical heterogeneity of the human disease. Biochemical evidence of OS appears to be related to neurological symptom severity, mutation type, and clinical presentation. These findings pave the way for potential new genetic downstream therapeutic strategies aimed at improving patient quality of life. Further efforts in the near future are needed for investigating the yet unexplored "black box" between the MeCP2 gene mutation and subsequent OS derangement.

Objective Rett syndrome (RTT) is a neurological disorder and a leading cause of mental retardation in females. It is caused by mutations in methyl-CpG-binding protein 2 (MeCP2) gene and more rarely in cyclin-dependent kinase-like 5 (CDKL5) and forkhead box protein G1 (FOXG1) genes. Increased oxidative stress (OS) has been documented in MeCP2-RTT patients. Here, we evaluated the levels of 4-hydroxynonenal plasma protein adducts (4HNE-PAs) in MeCP2-, CDKL5-, and FOXG1-RTT and in their clinical variants. Design and methods 4HNE-PAs were determined by Western blot in plasma from healthy subjects and RTT patients. Results 4HNE-PAs levels were increased in MeCP2- and CDKL5-related RTT but not in FOXG1-related RTT. Conclusion These results showed that OS is present in RTT clinical variants and could play a key role in RTT pathogenesis. Under the OS point of view FOXG1-related RTT appears to be distinct from the MeCP2/CDKL5, suggesting a distinct mechanism involved in its pathogenesis.

An increased oxidative stress and a decreased life span of erythrocytes (RBCs) are reported in patients with diabetes. Aim of this study was to assess in RBCs from patients with type 2 diabetes whether downstream effector mechanisms of apoptosis, such as activation of caspase-3, is operative, and whether an iron-related oxidative imbalance, occurring inside RBCs and in plasma, could be involved in caspase-3 activation. In 26 patients with type 2 diabetes and in 12 healthy subjects, oxidative stress was evaluated by means of different markers; non-protein-bound iron, methemoglobin and glutathione were determined in RBCs, and non-protein-bound iron was also determined in plasma. Erythrocyte caspase-3 activation was evaluated by an immunosorbent enzyme assay. Arterial hypertension, demographic and standard biochemical data were also evaluated. The results show, for the first time, that type 2 diabetic RBCs put into motion caspase-3 activation, which is significantly higher than in control RBCs. Such an effector mechanism of "eryptosis" was positively correlated to blood glucose levels and to the increased plasma NPBI level. Caspase-3 activation was also positively correlated to occurrence of arterial hypertension. The results suggest that an extracellular oxidative milieu can be responsible for erythrocyte caspase-3 activation in patients with type 2 diabetes. In turn, caspase-3 activation can be envisaged as a novel mechanism which, by impairing the maintenance of erythrocyte shape and function, might contribute to the shortened life span of RBCs from patients with type 2 diabetes and to hemorheological disorders observed in these patients.

Background Breathing disorders in Rett Syndrome (RS) have been generally attributed to severe autonomic and/or brain stem dysfunction, and, to our knowledge, no information regarding lung morphologic characteristics exists to date. The aim of the present study was to determine if there are RS-associated pulmonary abnormalities. Methods A total of 27 female subjects (age, M ± SD: 12.6 ± 5.9 y; age range: 3-32 y) with gene-encoding, methyl-CpG-binding-protein-2-mutation-confirmed RS underwent high-resolution CT (HRCT) scans of the thorax. A volumetric acquisition was used, and isotropic data sets were acquired with thin collimation ( Results Abnormal HRCT scan findings were observed in 15 of 27 (55.5%) cases, consisting of centrilobular nodules (10/15, 66.7%), thickening of the bronchial walls (8/15, 53.33%), and patchy ground-glass opacities (4/15, 26.7%), with upper lobe predominance. In addition, bronchiolectasis were found in nine of 15 (60%) patients. Conclusions Pulmonary lesions, respiratory bronchiolitis-associated interstitial lung disease-like lesions, are present on imaging studies in about half of typical patients with RS. Further research is needed to clarify the epidemiologic characteristics and the pathogenesis of these previously unrecognized pulmonary abnormalities.

Nerve cells are very susceptible to hypoxia responsive for mitochondrial dysfunctions involved in the subsequent oxidative stress, apoptosis and necrosis. In this paper, we examined the effect of 12 h incubation of U-373 MG astrocytes in hypoxic environment (73% N(2): 2% O(2): 5% CO(2), v:v) by evaluating cell proliferation, modifications of NO and ATP production, intracellular Ca(2+) concentration [Ca(2+)](i), membrane potential, desferoxamine-chelatable free iron, esterified F2-isoprostanes levels and the production of phosphorylated ERK. The same parameters were evaluated also after a following re-oxygenation period of 24 h. Immediately after hypoxia the NO concentration increased significantly and returned to values similar to those of controls after the re-oxygenation period. At the same time, ATP levels remained similar to controls and the cell proliferation significantly decreased. This involved a significant increase of [Ca(2+)](i) immediately after hypoxia and the value remained significantly elevated after the following re-oxygenation period. Moreover, after hypoxia, astrocytes were slightly although not significantly depolarized. Indeed iron and F2-isoprostanes levels increased significantly after hypoxia. Finally ERK proteins increased slowly and not significantly after hypoxia and the same trend was observed after the re-oxygenation period. On the whole, our results indicate that 2% O(2) hypoxia induces a moderate oxidative stress, well tolerated by U-373 MG cells, remaining the ATP production, mitochondrial membrane potential and activated ERK proteins, similar to the values of controls.

Nerve cells are very responsive to weak pulsed electromagnetic fields (EMFs). Such non-ionizing radiation, with frequencies of 0–300 Hz and 0.1–100 mT, can affect several cellular activities, with unusual dose–response characteristics. The present study examined the effect of a 2-h exposure of synaptosomes on a system generating a peak magnetic field of 2 mT. We evaluated the changes of the synaptosomal mitochondrial respiration rate and ATP production, membrane potential, intrasynaptosomal Ca2+ concentration, and the release of free iron and F2-isoprostanes. O2 consumption and ATP production remained unchanged in exposed synaptosomes. The intrasynaptosomal Ca2+ concentration decreased slowly and no depolarization of the synaptosomal membrane was detected. Finally, the release of free iron and F2-isoprostanes by synaptosomal suspensions also remained unchanged after EMF exposure. These results indicate that the physiological behavior of cortical synaptosomes was unaffected by weak pulsed EMFs.

Oxidative stress due to free radical formation and initiation of abnormal oxidative reactions is involved in several diseases of newborns, such as hypoxic-ischemic encephalopathy. Melatonin, an endogenously produced indoleamine primarily formed in the pineal gland, is a potent free radical scavenger as well as an indirect antioxidant. The present study was conducted to evaluate the formation of oxidative damage mediators and the possible effect of melatonin treatment in a model of hypoxic-ischemic encephalopathy in 7-day-old rats. Pups were subjected to permanent ligation of the right common carotid artery and exposed for 2.5 hr to a nitrogen-oxygen mixture (92% and 8%, respectively) (hypoxia-ischemia, HI). Melatonin was injected intraperitoneally to a group of rats at the dose of 15 mg/kg 30 min before starting the ischemic procedure (HI-Melatonin). After 24 hr of treatment, in homogenized cerebral cortex, desferoxamine (DFO)-chelatable free iron, total F(2)-isoprostanes and total F(4)-neuroprostanes, originating from the free radical-catalyzed peroxidation of arachidonic and docosahexaenoic acids, respectively, were determined. HI induced a significant increase in DFO-chelatable iron, total F(2)-isoprostanes and F(4)-neuroprostanes in both right and left side of the cerebral cortex. In HI-Melatonin-treated animals the levels of free iron, F(2)-isoprostanes, and F(4)-neuroprostanes were significantly lower than that in HI rats and the values were similar to controls. These data show the important neuroprotective role of melatonin in reducing oxidative damage resulting from HI. Melatonin could represent a potential safe approach to perinatal brain damage in humans.

Nonprotein-bound iron (NPBI) and F2-isoprostanes, reliable markers of oxidative stress, are increased in plasma of newborns and inversely correlated to the gestational age. Because NPBI represents a pro-oxidant stimulus in plasma, we test the hypothesis that the entity of lipid peroxidation is related with NPBI concentrations. Plasma levels of free, esterified, and total F2-isoprostanes were investigated in relation to NPBI levels in 59 newborns and 16 healthy adults. The pro-oxidant role of iron was ascertained in vitro, by measuring all the forms of F2-isoprostanes after incubation with ammonium iron sulfate. Significant positive correlations were found between NPBI and total as well as esterified F2-isoprostanes in plasma of the newborns. The addition of ammonium iron sulfate induced a marked increase in all the forms of F2-isoprostanes after 2 hours of incubation. The higher NPBI concentration, the higher F2-isoprostanes levels. An increase NPBI dose dependent in total F2-isoprostanes formation was observed in dialyzed low density lipoprotein from adult plasma. The results clearly show that once NPBI is generated, whatever its source, it is capable of inducing oxidative stress. NPBI-induced oxidative stress may contribute to the morbidity in preterm infants that are particularly susceptible to free radical damage.

Down syndrome (DS, trisomy 21) is the leading cause of chromosomal-related intellectual disability. At an early age, adults with DS develop with the neuropathological hallmarks of Alzheimer's disease, associated with a chronic oxidative stress. To investigate if non-protein bound iron (NPBI) can contribute to building up a pro-oxidative microenvironment, we evaluated NPBI in both plasma and erythrocytes from DS and age-matched controls, together with in vivo markers of lipid peroxidation (F2-isoprostanes, F2-dihomo-isoprostanes, F4-neuroprostanes) and in vitro reactive oxygen species (ROS) formation in erythrocytes. The serum iron panel and uric acid were also measured. Second, we explored possible correlation between NPBI, lipid peroxidation and cognitive performance. Here, we report NPBI increase in DS, which correlates with increased serum ferritin and uric acid. High levels of lipid peroxidation markers and intraerythrocyte ROS formations were also reported. Furthermore, the scores of Raven's Colored Progressive Matrices (RCPM) test, performed as a measure of current cognitive function, are inversely related to NPBI, serum uric acid, and ferritin. Likewise, ROS production, F2-isoprostanes, and F4-neuroprostanes were also inversely related to cognitive performance, whereas serum transferrin positively correlated to RCPM scores. Our data reveal that increased availability of free redox-active iron, associated with enhanced lipid peroxidation, may be involved in neurodegeneration and cognitive decline in DS. In this respect, we propose chelation therapy as a potential preventive/therapeutic tool in DS.

Rett syndrome (RTT) is a devastating neurodevelopmental disease, previously included into the autistic spectrum disorders, affecting almost exclusively females (frequency 1:10,000). RTT leads to intellective deficit, purposeful hands use loss and late major motor impairment besides featuring breathing disorders, epilepsy and increased risk of sudden death. The condition is caused in up to 95% of the cases by mutations in the X-linked methyl-CpG binding protein 2 (MECP2) gene. Our group has shown a number of previously unrecognized features, such as systemic redox imbalance, chronic inflammatory status, respiratory bronchiolitis-associated interstitial lung disease-like lung disease, and erythrocyte morphology changes. While evidence on an intimate involvement of MeCP2 in the immune response is cumulating, we have recently shown a cytokine dysregulation in RTT. Increasing evidence on the relationship between MeCP2 and an immune dysfunction is reported, with, apparently, a link between MECP2 gene polymorphisms and autoimmune diseases, including primary Sjogren's syndrome, systemic lupus erythematosus, rheumatoid arthritis, and systemic sclerosis. Antineuronal (i.e., brain proteins) antibodies have been shown in RTT. Recently, high levels of anti-N-glucosylation (N-Glc) IgM serum autoantibodies [i.e., anti-CSF114(N-Glc) IgMs] have been detected by our group in a statistically significant number of RTT patients. In the current review, the Authors explore the current evidence, either in favor or against, the presence of an autoimmune component in RTT.

Contains fulltext : 153430.pdf (Publisher’s version ) (Open Access) Haematopoietic progenitor cells show special sensitivity to mitochondrial DNA (mtDNA) mutagenesis, which suggests that increased mtDNA mutagenesis could underlie anemias. Here we show that elevated mtDNA mutagenesis in mice with a proof-reading deficient mtDNA polymerase (PolG) leads to incomplete mitochondrial clearance, with asynchronized iron loading in erythroid precursors, and increased total and free cellular iron content. The resulting Fenton chemistry leads to oxidative damage and premature destruction of erythrocytes by splenic macrophages. Our data indicate that mitochondria actively contribute to their own elimination in reticulocytes and modulate iron loading. Asynchrony of this sequence of events causes severe mitochondrial anaemia by depleting the organism of red blood cells and the bone marrow of iron. Our findings account for the anaemia development in a progeroid mouse model and may have direct relevance to the anemias associated with human mitochondrial disease and ageing.