Structural brain anomalies in Cri-du-Chat syndrome: MRI findings in 14 patients and possible genotype-phenotype correlations.

INTRODUCTION: Cri-du-Chat Syndrome (CdCS) is a genetic condition due to deletions showing different breakpoints encompassing a critical region on the short arm of chromosome 5, located between p15.2 and p15.3, first defined by Niebuhr in 1978. The classic phenotype includes a characteristic cry, peculiar facies, microcephaly, growth retardation, hypotonia, speech and psychomotor delay and intellectual disability. A wide spectrum of clinical manifestations can be attributed to differences in size and localization of the 5p deletion. Several critical regions related to some of the main features (such as cry, peculiar facies, developmental delay) have been identified. The aim of this study is to further define the genotype-phenotype correlations in CdCS with particular regards to the specific neuroradiological findings. PATIENTS AND METHODS: Fourteen patients with 5p deletions have been included in the present study. Neuroimaging studies were conducted using brain Magnetic Resonance Imaging (MRI). Genetic testing was performed by means of comparative genomic hybridization (CGH) array at 130 kb resolution. RESULTS: MRI analyses showed that isolated pontine hypoplasia is the most common finding, followed by vermian hypoplasia, ventricular anomalies, abnormal basal angle, widening of cavum sellae, increased signal of white matter, corpus callosum anomalies, and anomalies of cortical development. Chromosomal microarray analysis identified deletions ranging in size from 11,6 to 33,8 Mb on the short arm of chromosome 5. Then, we took into consideration the overlapping and non-overlapping deleted regions. The goal was to establish a correlation between the deleted segments and the neuroradiological features of our patients. CONCLUSIONS: Performing MRI on all the patients in our cohort, allowed us to expand the neuroradiological phenotype in CdCS. Moreover, possible critical regions associated to characteristic MRI findings have been identified.

7q11.23 microduplication syndrome: neurophysiological and neuroradiological insights into a rare chromosomal disorder.

BACKGROUND: The phenotypical consequence of the heterozygous chromosome 7q11.23 interstitial microdeletion is the Williams-Beuren syndrome, a very well-known genetic multi-systemic disorder. Much less is known about the reverse condition, the heterozygous interstitial microduplication of 7q11.23 region. The first molecular cytogenetic description was published in 2005, and only after several years were the reported patients numerous enough to attempt a description of a common phenotype. METHOD: By using a broad multidisciplinary approach, we investigated 12 patients with this rare genetic anomaly. Ten of them harboured the duplication of the classical Williams-Beuren syndrome region and two a slightly larger duplication. Upon a detailed description of the clinical and psychological features, we used electroencephalography and magnetic resonance imaging to explore neurophysiological function and brain structures. RESULTS: We analysed the clinical, psychological, neuroradiological and neurophysiological features of 12 yet-unpublished individuals affected by this rare genetic anomaly, focusing specifically on the last two aspects. Several structural abnormalities of the central nervous system were detected, like ventriculomegaly, hypotrophic cerebellum, hypotrophic corpus callosum and hypoplastic temporal lobes. Although only one of 12 individuals suffered from seizures during childhood, three others had abnormal electroencephalography findings prominent in the anterior brain regions, without any visible seizures to date. CONCLUSION: Taken together, we enlarged the yet-underrepresented cohort in the literature of patients affected by 7q11.23 microduplication syndrome and shed further light on neuroradiological and neurophysiological aspects of this rare genetic syndrome.

Occupational Exposure to Chromium of Assembly Workers in Aviation Industries.

Aircraft are constructed by modules that are covered by a "primer" layer, which can often contain hexavalent chromium [Cr(VI)], known carcinogen to humans. While the occupational exposure to Cr(VI) during aircraft painting is ascertained, the exposure assessment of assembly workers (assemblers) requires investigations. Three biological monitoring campaigns (BM-I,II,III) were performed in an aviation industry, on homogeneous groups of assemblers (N = 43) and controls (N = 23), by measuring chromium concentrations in end-shift urine collected at the end of the working week and the chromium concentration difference between end- and before-shift urines. BM-I was conducted on full-time workers, BM-II was performed on workers after a 3-4 day absence from work, BM-III on workers using ecoprimers with lower Cr(VI) content. Samples were analyzed by atomic absorption spectroscopy and mean values were compared by T-test. Even if Cr concentrations measured during BM-I were lower than Biological Exposure Indices by ACGIH, statistically significant differences were found between urinary Cr concentrations of workers and controls. Despite 3-4 days of absence from work, urinary chromium concentrations measured during BM-II were still higher than references from nonoccupationally exposed populations. In the BM-III campaign, the obtained preliminary results suggested the efficacy of using ecoprimers. The healthcare of workers exposed to carcinogenic agents follows the principle of limiting the exposure to "the minimum technically possible". The obtained results evidence that assemblers of aviation industries, whose task does not involve the direct use of primers containing Cr(VI), show an albeit slight occupational exposure to Cr(VI), that must be carefully taken into consideration in planning suitable prevention measures during risk assessment and management processes.

Immune-inflammatory responses and oxidative stress in Alzheimer's disease: therapeutic implications.

Alzheimer's disease (AD) is a heterogeneous and progressive neurodegenerative disease which in Western society mainly accounts for clinical dementia. AD has been linked to inflammation and oxidative stress. Neuro-pathological hallmarks are senile plaques, resulting from the accumulation of several proteins and an inflammatory reaction around deposits of amyloid, a fibrillar protein, Abeta, product of cleavage of a much larger protein, the beta-amyloid precursor protein (APP) and neurofibrillary tangles. Inflammation clearly occurs in pathologically vulnerable regions of AD and several inflammatory factors influencing AD development, i.e. environmental factors (pro-inflammatory phenotype) and/or genetic factors (pro-inflammatory genotype) have been described. Irrespective of the source and mechanisms that lead to the generation of reactive oxygen species, mammalian cells have developed highly regulated inducible defence systems, whose cytoprotective functions are essential in terms of cell survival. When appropriately activated, each one of these systems has the possibility to restore cellular homeostasis and rebalance redox equilibrium. Increasing evidence, support the notion that reduction of cellular expression and activity of antioxidant proteins and consequent augment of oxidative stress are fundamental causes for ageing processes and neurodegenerative diseases., including AD. The better understanding of different molecular and cellular inflammatory mechanisms is crucial for complete knowledge of AD pathophysiology, hence for its prevention and drug therapy. Accordingly, two lines of preventive therapeutics can be outlined, the first based on anti-inflammatory drugs, the second one on anti-oxidative properties.

Association between the polymorphisms of TLR4 and CD14 genes and Alzheimer's disease.

Alzheimer's disease (AD) is a heterogeneous and progressive neurodegenerative disease which in Western society mainly accounts for clinical dementia. Inflammation plays a key role in AD and dissecting the genetics of inflammation may provide an answer to the possible treatment. Hence, the better understanding of different molecular and cellular inflammatory mechanisms is crucial for complete knowledge of AD pathophysiology, and for its prevention and drug therapy. Accordingly, in the present study we evaluated whether the pro-inflammatory polymorphisms of lipopolysaccaride-receptors, +896A/G Toll-Like Receptor (TLR4) and -260C/T CD14, are risk factors for AD. The study included both 626 AD patients (427 women and 199 men; age range: 53-98 years; mean age: 74.88+/-8.44) from Northern Italy and age and gender matched controls. Our results demonstrate that the +896A/G TLR4 single nucleotide polymorphism (SNP) is associated with AD, whereas no association has been observed with -260C/T CD14 SNP. Furthermore, no differences have been observed evaluating the combined presence of +896A+TLR4/-260T+CD14 "high responder"(proinflammatory-profile). However, our results showing the involvement of TLR4 in AD pathophysiology, strengthen the suggestion that systemic inflammation plays a key role in AD. Carriers of high responder SNP, affected by mild cognitive impairment might, be the ideal target for a preventive treatment with biologics as monoclonal antibodies directed against the pro-inflammatory cytokines to decrease the level of systemic inflammation involved in AD pathophysiology.

Quantification of sirolimus and everolimus by immunoassay techniques: test specificity and cross-reactivity evaluation.

The possible cross-reactivity of immunoassays with structurally-related drugs was investigated. Innofluor Certican (FPIA) calibrators were measured by using IMx Sirolimus assay (MEIA) and MEIA Sirolimus calibrators were analysed by using FPIA Certican assay. Drug concentrations were measured in 95 and 100 samples from renal transplanted patients (RTP) on sirolimus or everolimus treatment by using immunoassays and LC/ESI-MSMS. A high cross-reactivity was found both for MEIA and FPIA. High correlation degrees, confirmed by the Bland-Altman and the Eksborg tests, were found between drug concentrations measured in real samples by both immunoassays (r = 0.909 and r = 0.970, respectively). LC/ESI-MSMS analysis of samples containing sirolimus showed no positivity for everolimus. Similarly, samples from patients on treatment with everolimus resulted negative as far as regards sirolimus. MEIA and FPIA could be considered mutually reliable and accurate alternatives for the specific-drug immunoassay. It should be noticed that in patients switching from one drug to the other unreal overestimation of the blood levels of the current administered immunosuppressant can occur.

[Antiblastic drug exposure: statistical analysis of the data coming from environmental monitoring of a site devoted to drug production]. / Esposizione a farmaci antiblastici: analisi statistica dei dati di monitoraggio ambientale in una sala di allestimento dei farmaci.

An environmental monitoring strategy was carried out for the determination of surface concentrations of cyclophosphamide (CP), ifosfamide (IF) and 5-fluorouracil (5-FU) in a drug preparation room of an oncology ward. Analytes were determined by wipe tests, liquid-liquid extraction with diatomaceous earths and GC/MSMS or HPLC/UV analysis. The analysed 249 samples showed concentrations of CP, IF and 5-FU varying in the ranges 0.020-18.83, 0.100-26.96, 0.740-208.9 microg/dm2, respectively. It is noteworthy that the 9.3% (CP), 18.6% (IF) and 76.3% (5-FU) of the investigated surfaces showed high contamination levels, with analytes amount higher than 0.5 microg/dm2 and a progressive contamination decrement going from workbenches, floor, hood planes and other examined surfaces (interphone, telephone etc.). A significant correlation (rhos = 0.303, p = 0.001) between the measured analyte concentration and the analyte handled amount was found only in the case of IF, and a diffuse contamination (traces of all the three analytes) was found on all investigated surfaces, even when analytes were not been used during the sampling days.

Mono- and diiodocyclophosphamide as possible internal standards for cyclophosphamide quantification: characterization by ion trap multi-stage mass spectrometry and effects of iodine-chlorine substitution on the fragmentation pattern.

Hospital personnel involved in antineoplastic drug preparation and administration to patients are exposed to large amounts of these drugs. Labour legislation indicates the necessity of planning monitoring strategies aimed at prevention and/or reduction of drug exposure. Monitoring strategies consist of quantitative determinations of indicators, present in environmental and biological matrices. Among the antineoplastic drugs widely used, cyclophosphamide (CP) has been identified as a suitable indicator of potential exposure to mixtures of antineoplastic drugs. Many literature methods for quantitative analysis of CP involve either liquid (LC) or gas chromatography (GC) with mass spectrometry (MS), both of which require use of a suitable internal standard. The present work focuses on the synthesis of mono- and diiodocyclophosphamide (CPI and CPI(2)) to be used as internal standard. These compounds were analyzed by GC/EI-MS/MS and LC/ESI-MS(n) using ion trap mass spectrometry. The product ion mass spectra are interpreted in terms of proposed structures of fragment ions. Iodine-chlorine substitution resulted in a weakening of the carbon-halogen bond with a noteworthy influence on the ion fragmentation processes. The proposed suitability of CPI and CPI(2) as internal standards was based on similarities to CP as regards ionization and fragmentation processes. The results obtained suggest that CPI could be used as internal standard for CP quantification by LC/ESI-MS/MS, and CPI(2) for GC/EI-MS/MS analyses.

Evidence that a dodecamer duplication in the gene HOPA in Xq13 is not associated with mental retardation.

A recent study suggested that a dodecamer duplication in exon 42 of the HOPA gene in Xq13 may be a significant factor in the etiology of X-linked mental retardation. In an effort to investigate this possibility, we determined the incidence of the dodecamer duplication in cohorts of non-fragile X males with mental retardation from three countries, cohorts of fragile X males from two countries, 43 probands from families with X-linked mental retardation and control cohorts from three countries. The duplication was found in 3.6-4.0% of male patients from two non-fragile X groups (Italy and South Carolina), in 1.2% from another non-fragile X group (South Africa), but in no male patients from families with X-linked mental retardation (South Carolina). The dodecamer duplication was also found in several white males with fragile X syndrome from France (5%) and South Africa (22.2%). Additionally, the duplication was found in 1.5% of South Carolinian newborn males, 2.5% South Carolinian male college students, 5% Italian male controls and 4.5% of the white South African controls. None of the black South African non-fragile X individuals with mental retardation, the fragile X or the control samples tested carried the duplication, suggesting that the duplication is rare in the black South African population. The incidence of the duplication was not significantly different between any of the groups in the study. Therefore, results of our studies in four different populations do not corroborate the findings of the previous study, and indicate that the HOPA dodecamer duplication does not convey an increased susceptibility to mental retardation.

Genetic variations in human fetal globin gene microsatellites and their functional relevance.

Short tandem repeats are abundantly present within the genome. They are commonly used as polymorphic markers but their potential functional role is poorly documented. Several of these microsatellites have been described within the beta-globin locus and some could be involved in controlling gene expression. Our purpose was to investigate the extent and significance of the (TG)n(CG)m dinucleotide repeat polymorphisms in the two gamma-globin gene IVS2s. Two groups of subjects were studied: a group of 63 beta-thalassaemic patients presenting either with a severe Cooley's anaemia (n=50) or with thalassaemia intermedia (TI, n=13), and a control group of 60 unrelated healthy individuals. A high heterogeneity of the polymorphic repeats was demonstrated, extending the range of the published alleles from 13 to 22 and allowing a first attempt at making a phenotype/genotype correlation. One specific allele, (TG)13 in the Agamma-gene, was highly enriched in the TI patients (46.1% vs 2.9% of the Cooley's anaemia cases, P < 0.0002, and 23.3% in the normal controls, P < 0.008) and preferentially observed in TI patients with a high haemoglobin F (Hb F). Transient transfection assays in K562 cells, with the growth hormone gene as a reporter, showed a positive regulatory action mediated by a (TG)13-containing 243 nt IVS2 fragment. Finally, a first set of mobility shift experiments with erythroid (K562 and MEL) and nonerythroid (HeLa) cell lines showed binding of erythroid component(s) in this DNA region and the binding pattern was modified upon induction of MEL cells by DMSO. Thus, our in vivo and in vitro data raise the question of a possible contribution of the gamma-gene IVS2s polymorphic microsatellites to the variable Hb F synthesis in the major haemoglobinopathies: a well known, puzzling and still unanswered question.

New mutations in XNP/ATR-X gene: a further contribution to genotype/phenotype relationship in ATR/X syndrome. Mutations in brief no. 176. Online.

The molecular causes of ATR-X syndrome reside in mutations involving the XNP/ATR-X gene, which maps in the Xq13.3 region. Mutational analysis of this gene in two unrelated affected patients allowed us to identify two new molecular defects in two distinct regions of the gene. The first is a A-->G splice mutation in the acceptor site of the intron 11 that removes most of the 3' part of the protein, including the helicase domains and the glutamic acid stretch. Three cryptic acceptor splice sites are activated by this point mutation with consequent production of three types of abnormal mRNA: two with intronic insertions and a smaller one, approximately 10% of the total transcript, which is shorter than normal mRNA by one amino acid residue (E). Since the physiopathological characteristics of the patient carrying the splice mutation do not exhibit severe urogential abnormalities despite the lack of the -COOH end of the protein, a residual function of this third transcript is to be suspected. The second encountered nucleotide change (G-->T) leads to an R246L amino acid substitution in the putative zinc finger DNA-binding domain in the -NH2 terminal part of the protein.