Central nervous system involvement in late-onset Pompe disease: clues from neuroimaging and neuropsychological analysis.

BACKGROUND AND PURPOSE: Late-onset Pompe disease (LOPD) is a rare, multisystem disorder that is well established to mainly impair skeletal muscle function. Systematic studies exploring brain functions in LOPD are lacking. The aim of this study was to detect morphological and functional brain alterations as well as neuropsychological impairment in LOPD. METHODS: We studied 21 patients (10 male, mean age 49 ± 18.4 years) with defined diagnosis of LOPD, divided into two groups: one with pre-symptomatic hyperCKemia with no muscle weakness and the second with limb-girdle muscle weakness. All patients underwent 3T magnetic resonance imaging (MRI) to obtain morphological/angiographic evaluation as well as normalized cortical brain volume and resting-state functional MRI. Fazekas score was applied to quantify white matter lesions, whereas Smoker's criteria were used to examine dolichoectasia. A complete neuropsychological assessment was performed. RESULTS: The MRI data showed that 12/21 patients (57%) demonstrated signs of cerebral vasculopathy, with a Fazekas score >2 in 67%. According to Smoker's criteria, 11/21 patients (52%) had a dolichoectasia of the vertebrobasilar system; an intracranial aneurysm was detected in 3/21 patients (14%). Resting-state functional MRI demonstrated significantly decreased brain connectivity in the salience network with a more relevant reduction in the bilateral middle and superior frontal gyrus. Gray matter atrophy correlated with age and disease duration. A mild impairment in executive functions was also identified. CONCLUSIONS: In this LOPD cohort the results showed morphological and functional brain alterations with mild neuropsychological dysfunction, mainly in the limb-girdle muscle weakness group. Cerebrovascular alterations seemed to be not related to common risk factors, suggesting a major role of enzymatic deficiency in the pathogenesis of brain abnormalities.

Cortical reorganization in multiple sclerosis after intrathecal baclofen therapy.

Our objective was to assess the role of Intrathecal Baclofen Therapy (ITB) in the cortical reorganization in a patient affected by multiple sclerosis (MS) undergoing physical therapy. We reported a case of a woman affected by MS and severe spasticity, who performed an fMRI examination, before and after the ITB implantation. The subject showed controlateral motor cortex activation after motor task. After a month of ITB implantation, patient showed ipsilateral and controlateral motor cortex activation although with a broader extension. fMRI examination supported the hypothesis of a central influence in patients who undergo physiotherapy and therapy with ITB.

Necrosis score, surgical time, and transfused blood volume in patients treated with preoperative embolization of intracranial meningiomas. Analysis of a single-centre experience and a review of literature.

PURPOSE: Several authors have demonstrated that preoperative embolization of meningiomas reduces blood loss during surgery. However, preoperative embolization is still under debate. Aim of this study is the retrospective evaluation of necrosis score, surgical time, and transfused blood volume, on patients affected by intracranial meningiomas treated with preoperative embolization before surgery, compared with a control group treated only with surgery. METHOD: Twenty-eight patients with meningiomas were subjected to a preoperative embolization with polyvinyl alcohol (PVA). These patients were divided into two groups: group 1, patients with preoperative embolization performed at least 7 days before surgery; and group 2, patients with preoperative embolization performed less than 7 days before surgery. A statistical evaluation was made by comparing necrosis score, surgical time, and transfused blood volume of these groups. Then, we compared these parameters also with group 3, which included patients with surgically treated meningioma who did not undergo preoperative embolization. RESULTS: Surgery time and transfused blood volume were significantly lower in patients who had been embolized at least 7 days before definitive surgery. Furthermore, large confluent areas of necrosis were significantly more frequent in patients with a larger time span between embolization and surgery. CONCLUSION: Preoperative embolization with PVA in patients with intracranial meningiomas is safe and effective, as it reduces the volume of transfused blood during surgical operation. However, patients should undergo surgery at least 7 days after embolization, as a shorter time interval has been correlated with a longer surgical time and a higher transfused blood volume.

Differences between conventional and nonconventional MRI techniques in Parkinson's disease.

Magnetic resonance imaging (MRI) provides an in vivo assessment of cortical and subcortical regions affected in Parkinson's disease (PD). This review summarizes the most important conventional and non-conventional MRI techniques applied in this field. Standard neuroimaging techniques have played a marginal role in the diagnosis and follow-up of PD, essentially being used only to discriminate atypical syndromes from PD, to exclude secondary causes such as vascular lesions, and to confirm the absence of specific imaging features found in atypical parkinsonisms. However, non-conventional MRI techniques, i.e. new neuroimaging approaches such as magnetic resonance spectroscopy, diffusion tensor imaging, and functional MRI, may allow the detection of structural, functional and metabolic changes useful not only for differential diagnosis, but also for early diagnosis and outcome and treatment monitoring in PD. In addition, we illustrate the advantages of high-field MRI over lower magnetic fields, highlighting the great potential of advanced neuroimaging techniques.

Proton magnetic resonance spectroscopy (H-MRS) in chronic schizophrenia. A single-voxel study in three regions involved in a pathogenetic theory.

The cognitive dysmetria theory suggests a disconnectivity between the dorsolateral prefrontal cortex, thalami and vermis to explain the pathophysiology of schizophrenia. This study investigated the metabolic integrity of this neurologic circuit in patients with schizophrenia using proton magnetic resonance spectroscopy (H-MRS). Twenty-two patients with schizophrenia and twelve control subjects were studied. Metabolites concentrations were evaluated by a single-voxel technique in the prefrontal cortex, thalami and vermis. To our knowledge, this is the first H-MRS experience with concomitant evaluation of these regions in schizophrenic patients. We found no significant statistical difference in N-AA, Cho and Cr absolute concentrations and N-AA/Cho, N-AA/Cr and Cho/Cr ratios between the schizophrenic patients and control group. At the vermis, we found a constant spectrum with low levels of N-AA and higher levels of Cho and Cr. Our experience does not clearly support or refute the cognitive dysmetria theory. The consistency of metabolic findings in the cerebellar vermis could represent an important datum, highlighting the specificity of metabolic and functional activity in this region.

Heavy metals affect regulatory volume decrease (RVD) in nematocytes isolated from the jellyfish Pelagia noctiluca.

The environmental contamination caused by heavy metals raises the question of their effect on biological systems. Among bio-indicators useful to monitor the toxicological effects of these chemicals, Cnidarians offer a unique model. Cnidarians possess highly specialized stinging cells, termed nematocytes, which respond to hyposmotic solution with well established homeostatic parameters as an acute osmotic phase (OP), leading to cell swelling, and then a slower regulatory volume decrease (RVD) phase, causing cell shrinkage. Here we report the effect of 65% artificial sea water (ASW) containing heavy metals, such as Cd, La, Co, Cu and Zn (concentrations comprised between 100 and 0.1 µM) on both OP and RVD in nematocytes isolated from the jellyfish Pelagia noctiluca by 605 mM NaSCN plus 0.01 mM Ca(2+). The exposure of the cells to Co and La inhibited RVD but not OP. However, Cu, Cd and Zn prevented the OP in a dose-dependent manner and, hence, also the detection of RVD. These results suggest that, in isolated nematocytes, heavy metal pollutants impair RVD either directly or indirectly through interference with the OP, thus negating RVD. Although further studies need to clarify the exact mechanisms whereby heavy metals exert their toxicity, it is evident that nematocytes of Cnidarians could serve as a model for ecotoxicological investigations.

Nematocytes' activation in Pelagia noctiluca (Cnidaria, Scyphozoa) oral arms.

Nematocytes' discharge is triggered to perform both defense and predation strategies in cnidarians and occurs under chemico-physical stimulation. In this study, different compounds such as amino acids and proteins (mucin, albumin, poly-L: -lysine, trypsin), sugars and N-acetylate sugars (N-acetyl neuraminic acid, N-acetyl galactosamine, sucrose, glucose, agarose and trehalose), nucleotides (ATP and cAMP), were tested as chemosensitizers of nematocyte discharge in the oral arms of the scyphozoan Pelagia noctiluca, particularly abundant in the Strait of Messina (Italy). Excised oral arms were submitted to a combined chemico-physical stimulation by treatment with different compounds followed by mechanical stimulation by a non-vibrating test probe. Discharge induced by a chemico-physical stimulation was more significant than that obtained after mechanical stimulation alone. A chemosensitizing mechanism, with a dose-dependent effect, was observed after treatment with sugars, amino compounds such as glutathione, nucleotides and mucin, according to that already seen in sea anemones. Such findings suggest that, though Anthozoa and Scyphozoa exhibit different divergence times during the evolutionary process, the discharge activation exhibits common features, probably derived from their last common ancestor.

Pancreatic and extrapancreatic lesions in patients with intraductal papillary mucinous neoplasms of the pancreas: a single-centre experience.

PURPOSE: This study was performed to describe the possible presence of extrapancreatic neoplasms in patients with intraductal papillary mucinous neoplasm (IPMN) and to evaluate whether the extrapancreatic tumours were synchronous or metachronous to IPMNs. MATERIALS AND METHODS: One hundred and forty-two patients (56 men and 86 women; mean age 69.5 years, range 37-98) with IPMN diagnosed using the Sendai criteria were enrolled. Six patients (4.2%) had type I, 103 (72.5%) type II and 33 (23.2%) type III IPMN. All patients were studied using the following imaging techniques: ultrasonography (US), multidetector computed tomography (MDCT) and magnetic resonance cholangiopancreatography (MRCP). RESULTS: Pancreatic IPMN was localised in the head in 43 patients (30.3%), in the body in 13 (9.2%), in the tail in ten (7.0%), in the head-body in 17 (12.0%), in the body-tail in 15 (10.6%) diffuse throughout the gland in 44 (31.0%). The mean size of the cystic lesions was [(mean+/-standard deviation (SD)] 1.9+/-1.9 cm (range 0.5-8.0 cm). Twenty patients (14.1%) had associated pancreatic or extrapancreatic diseases. In evaluating the distribution of pancreatic or extrapancreatic diseases according to IPMN type, we found that this was not significantly different among types I, II and III (p=0.776). CONCLUSIONS: The majority of pancreatic and extrapancreatic cancers occur before the diagnosis of IPMNs is made and is not related to the type of IPMN.

One century sedimentary record of Hg and Pb pollution in the Sagua estuary (Cuba) derived from 210Pb and 137Cs chronology.

The vertical distribution of Hg and Pb were determined in a sediment core collected from the Sagua estuary (North Cuba) that receives input from the Sagua river, one of the most polluted rivers discharging into the Cuban coastal environment. Depth profiles of metal concentrations were converted to time-based profiles using the (210)Pb dating method and confirmed with the (137)Cs fallout peak. The mean mass accumulation rate was estimated to be 0.17+/-0.04 g cm(-2)y(-1) (mean sediment accumulation rate 0.52+/-0.13 cm y(-1)) and the core bottom was estimated to date back about 130 years. The historical sedimentary record showed a strong enrichment of mercury concentrations in the past decades, caused by the incomplete treatment of industrial wastes from a chlor-alkali plant with mercury-cell technology in the Sagua river basin. Lead fluxes to sediments showed a gradual increase from the 1920s to present, which agrees with a population increase in Sagua la Grande City. Fluxes of both metals have increased the past 25 years, with values reaching a maximum of 0.5 and 3.9 microg cm(-2)y(-1) for Hg and Pb, respectively.

Factors altering the haemolytic power of crude venom from Aiptasia mutabilis (Anthozoa) nematocysts.

The effect of different agents upon the haemolytic power of Aiptasia mutabilis crude venom was studied inhuman erythrocytes to determine its toxicity and stability. Nematocysts were isolated from acontia of the Anthozoan A. mutabilis and submitted to sonication for extracting crude venom. Aliquots of venom were tested in 0.05% erythrocyte suspensions in the presence of various factors such as proteases (papain,collagenase, trypsin, alpha-chymotrypsin); cations (Ca2+, Mg2+, Ba2+, K+ and Cu2+), osmotic protectants as polyethylenglycole (PEG) of different MW and antioxidant compounds (GSH, cysteine and ascorbic acid).Results demonstrate the dose-response of the haemolytic effect of A. mutabilis. Haemolysis by the crude venom was prevented by Ca2+, Ba2+ and Cu2+ treatment, and to a minor extent by Mg2+ and K+. Papain and PEG with a molecular mass exceeding 1000 Da also prevented haemolysis. These findings are consistent with a pore-forming mechanism of crude venom in erythrocytes rather than an oxidative damage at the employed doses.

Effect of various factors on Pelagia noctiluca (Cnidaria, Scyphozoa) crude venom-induced haemolysis.

The haemolytic power of isolated nematocysts from the scyphozoan Pelagia noctiluca was studied with attention to the effect of osmotic protectants as carbohydrates at different MW, cations as Mg2+, Ca2+, Ba2+,Cu2+, K+; proteases as collagenase, trypsin, alpha-chymotrypsin, papain; and antioxidants. Crude venom was at first obtained by sonication of holotrichous-isorhiza nematocysts previously isolated from oral arms of P. noctiluca and then haemolytically tested upon human erythrocytes. Osmotic protectants were effective in inhibiting the haemolytic power depending on their molecular weight so that total inhibition of crude venom-induced haemolysis was observed after PEG treatment (polyethyleneglycol 6000Da). Amongst divalent cations only Ba2+ and Cu2+ significantly inhibited the haemolytic power of crude venom. Proteases seem not to alter the haemolytic activity while antioxidant compounds only slightly reduced the haemolytic power. Such findings may suggest a pore-forming mechanism for P. noctiluca crude venom rather than an oxidative damage to the cell membrane.

The unusual toxicity and stability properties of crude venom from isolated nematocysts of Pelagia noctiluca (Cnidaria, Scyphozoa).

We have firstly investigated the toxicological activity by hemolytic assay of crude extract obtained by sonication of holotrichous isorhiza isolated nematocysts of the Scyphozoan Pelagia noctiluca, collected in the Strait of Messina. The hemolytic activity was both time- and dose-dependent on fish, rabbit, chicken and human red blood cells. At lowest doses rabbit and chicken erythrocytes were the most sensitive, whereas those of eel were the most resistant to the crude extract. Different storage conditions, such as -20 degrees C, -80 degrees C for up to 6 months and lyophilization, did not affect the stability of crude venom. Moreover, neither treatment at 4 degrees C, 20 degrees C and 37 degrees C for different time periods ranging between 30 min and 24 h, nor harsh thermal treatment at 80 degrees C and 100 degrees C affected the hemolytic power. The crude venom resulted even stable towards proteolysis and alkaline pH values.

Total and partial digestion of sediments for the evaluation of trace element environmental pollution.

Four different sample treatment methods for the determination of trace elements have been compared: a total digestion with HNO3-H2O2-HF using microwave, and three different standardized methods of fractionation: BCR three-steps sequential extraction, USEPA standard 3050B and ISO standard 11466. The four treatment methods were applied to the determination of Cu and Ni in four samples collected in different areas of Cienfuegos Bay (Cuba). The location of samples and the analytes were selected on the basis of results obtained by previous studies. Analyses following total digestion and BCR three-steps procedure were performed by inductively coupled plasma mass spectroscopy whereas analyses following EPA and ISO procedures were performed by flame atomic absorption spectroscopy. The results obtained have been compared with an estimated anthropic fraction evaluated in each sampling point as the difference between the total concentration and an estimated background concentration level. The BCR three-steps provided the best approximation of the estimated anthropic fraction and was therefore applied also in the determination of Pb and Cd for further consideration.

Estrogens counteract the masculinizing effect of tributyltin in zebrafish.

Recently, it has been demonstrated that the biocide tributyltin (TBT) can interfere with fish sex differentiation, leading to a bias of sex toward males. On the contrary, it is well known that estrogenic compounds can induce fish feminization. Yet, the combined effects of mixtures of androgenic and estrogenic compounds on fish sex differentiation have never been investigated before, even though in the environment animals are frequently exposed to both groups of xenobiotics. Therefore, in order to investigate whether exposure to estrogenic compounds can block the masculinizing effect of TBT, 5 days post-fertilization zebrafish (Danio rerio) larvae were exposed for a four month period to TBT and to the synthetic estrogen-ethinylestradiol (EE2). The fish were fed a diet containing TBT at nominal concentrations of 25 and 100 ng TBT/g, and two groups of animals were also dosed with TBT plus EE2 at nominal water concentration of 3.5 ng/L, using a flow-through design. As expected, fish exposed to TBT showed a bias of sex toward males (62.5% males in control tanks and 86% and 82% in TBT 25 and TBT 100 ng TBT/g, respectively). Co-exposure to EE2 completely blocked the masculinizing effect of TBT, with 7% males in the TBT 25 ng/g + EE2 treatment and 0% in the EE2 alone and in the TBT 100 ng/ + EE2 exposed groups. These results clearly indicate that EE2, at environmentally relevant concentrations, can block the TBT masculinizing effects in zebrafish, which suggests that in the aquatic environment the presence of estrogens may neutralize the fish masculinizing effect of TBT. Our findings highlight the need of testing the combined effects of contaminants, as single exposure studies may not be sufficient to predict the effects of mixtures of xenobiotics with antagonistic properties.

New insights into the mechanism of imposex induction in the dogwhelk Nucella lapillus.

In an attempt to clarify the mechanism(s) of tributyltin-mediated imposex induction in females of the neogastropod Nucella lapillus, dogwhelks collected in an almost imposex free population were exposed to several treatments for a 3 month-period, and the effects on imposex induction and testosterone/estradiol levels were evaluated. As a positive control, tributyltin (50 ng TBT Sn/L) clearly induced imposex and led to a significant increase in the severity of the phenomenon. In contrast, although a selective P450 aromatase inhibitor (formestane at 0.3 mg/L) was capable of imposex induction, it failed to increase its severity. A vertebrate androgen receptor (AR) antagonist (cyproterone acetate at 1.25 mg/L) in combination with TBT completely blocked the imposex induction capacity of TBT. On the other hand, an estrogen receptor antagonist (tamoxifen at 0.3 mg/L) rendered no effect. The determination of steroid levels in female specimens revealed that TBT induces an elevation of free testosterone (but not the total amount, free+esterified), while the co-administration of the anti-androgen and TBT was able to rescue the increase of free testosterone levels. Despite a minor decrease in the amount of testosterone-fatty acid esters in the TBT group, significant differences in esterified testosterone were not found among treatments. On the contrary, free estradiol levels were elevated in the TBT, anti-androgens and TBT plus anti-androgens groups. These results indicate that free estradiol biosynthesis in TBT-exposed females does not seem to be affected. Overall, our results demonstrate that a selective aromatase inhibitor can induce imposex in N. lapillus but not to a similar extent of TBT, which may suggest the involvement of other mechanism in imposex induction, besides aromatase inhibition. Additionally, the study points to the involvement of AR receptors in imposex induction.

Butyltins concentration levels and imposex occurrence in snails from the Sicilian coasts (Italy).

Two sampling campaigns have been carried out along the Sicilian coasts within the frame of a European project (Hic-TBT) financed by the LIFE programme. Snails and sediments have been sampled at 5 stations, characterised by very low (Ustica natural sea reserve), low (Ustica harbour), intermediate (Castellamare and Termini Imerese) and high shipping densities (Palermo). Biological and chemical analyses were carried out in order to evaluate the incidence of imposex occurring in snails, and the concentration levels of organotins in sediment and snail samples. Despite the low organotin concentration levels in sediment and snail samples, the results show that imposex is widespread around Sicily. In four locations almost all female snails were affected by imposex at advanced development stages, even in areas with a fairly low shipping density. Only in the natural sea reserve of Ustica Island imposex was relatively less severe. Finally a logarithmic correlation between TBT and imposex stages was observed.

Evaluation of performance of time-saving extraction devices in the BCR three-step sequential extraction procedure.

Two alternative extraction methods--a routine ultrasonic bath and a microwave oven--were developed and optimized for their eventual exploitation in the three-stage sequential extraction procedure proposed by the European Standards, Measurements and Testing (S, M & T) Programme, formerly Bureau Communitaire de Reference (BCR), for the operationally defined speciation of Cd, Cr, Cu, Ni and Zn. The conventional BCR three-stage sequential extraction procedure was modified at each stage, applying ultrasonic or microwave devices in order to shorten the required 16 h of shaking in all three steps. The experimental tests and the optimization of the operating parameters were carried out on a highly homogenized estuarine sediment reference material (RM S7) prepared by the Joint Research Centre (JRC) of ISPRA. Extractable metal contents were measured by inductively coupled plasma-mass spectrometry (ICP-MS) and the results obtained in each fraction by all three methods were statistically compared for all the studied elements. The conventional and newly developed alternative extraction methods were finally compared by the analysis of BCR 601, which is certified for the three-step BCR sequential extraction procedure. According to the statistical evaluation of the results, the proposed accelerated sequential extraction methods are valid alternatives to conventional shaking, with much shorter extraction times.

Comparison between different leaching tests for the evaluation of metal release from fly ash.

Comparison of three different leaching tests, each characterized by different conditions, e.g. liquid/solid ratio, pH, leachant, stirring conditions, duration time, has been performed within the activities of Analeach, financed by the European Union in the "Cooperation with third Countries" programme (INCO). The comparison was performed on four highly homogenized fly ash reference materials, of different origin, provided by the Joint Research Centre of Ispra. Seven different elements (Cd, Cr, Cu, Ni, Pb, V, and Zn) were considered in this study. The influence of the main experimental conditions and of the matrix on metal release was studied. Basic characterization of the materials was also performed.

A new mussel certified reference material (CRM 477) for the quality control of butyltin determination in the marine environment.

The analytical techniques used for the determination of butyltin compounds in biological tissues are generally based on a succession of steps (e.g., extraction, derivatization, separation, detection) which are all prone to systematic errors. An interlaboratory programme performed at the EU level and based on a stepwise approach has enabled one to identify and remove most of these errors and to improve considerably the state of the art so that certification of a mussel reference material has been made possible. This paper describes the preparation of this mussel reference material containing monobutyltin (MBT), dibutyltin (DBT) and tributyltin (TBT), the homogeneity and stability studies and the analytical work performed for the certification of the contents of MBT [Sn(C4H9)3+], DBT [Sn(C4H9)2(2+)] and TBT [Sn(C4H9)3+]. The results obtained by a group of expert EU laboratories are discussed along with the methods used to certify the mass fractions (based on dry mass) of monobutyltin (1.50 +/- 0.28 mg kg-1 as MBT cations), dibutyltin (1.54 +/- 0.12 mg kg-1 as DBT cations) and tributyltin (2.20 +/- 0.19 mg kg-1 as TBT cations). The paper also describes an attempt to certify the content of triphenyltin in this material which, although reasonable agreement was found among the participants' results, failed owing to the instability of this compound. This new material will be of great support to ensure the quality control of butyltin determination in shellfish, as carried out in environmental monitoring programmes.

Determination of PAH in atmospheric particulates in the area of the Italian base in Antarctica: report on monitoring activities during the last three scientific expeditions.

Concentrations of polycyclic aromatic hydrocarbons (PAH) in atmospheric particulates have been determined in the frame of the control activities for environmental protection during the VI (1990-1991), VII (1991-1992) and VIII (1992-1993) Italian expeditions in Antarctica. Samples were collected at the four cardinal points at about 200 m from the central area of the Italian scientific base. Data show low levels of the individual PAH (95% in the range 1-50 pg m(-3)) confirming the absence, up to now, of significant contamination from the base. These results could be used as a reference for the assessment of the air pollution due to combustion sources present in the base. Any possible variation in the base's activities in the future should be analysed and evaluated with respect to these findings.