Association between sex, systemic iron variation and probability of Parkinson's disease.

OBJECTIVE: Iron homeostasis appears altered in Parkinson's disease (PD). Recent genetic studies and meta-analyses have produced heterogeneous and inconclusive results. In order to verify the possible role of iron status in PD, we have screened some of the main metal gene variants, evaluated their effects on iron systemic status, and checked for possible interactions with PD. MATERIALS AND METHODS: In 92 PD patients and 112 healthy controls, we screened the D544E and R793H variants of the ceruloplasmin gene (CP), the P589S variant of the transferrin gene (TF), and the H63D and C282Y variants of the HFE gene, encoding for homologous proteins, respectively. Furthermore, we analyzed serum concentrations of iron, copper and their related proteins. RESULTS: The genetic investigation revealed no significant differences in allelic and genotype distributions between patients and controls. Two different multivariable forward stepwise logistic models showed that, when the effect of sex is considered, an increase of the probability of having PD is associated with low iron concentration and Tf-saturation. CONCLUSIONS: This study provides new evidence of the involvement of iron metabolism in PD pathogenesis and reveals a biological effect of sex.

Wearing-off detection in clinical practice: the wearing off real practice key (WORK-PD) study in Parkinson's disease.

OBJECTIVE: Verifying the validity and feasibility of the WOQ-19 as a useful tool in routine clinical practice and in management of patients. METHODS: 532 consecutive Parkinson's disease (PD) patients were recruited from 6 different neurological outpatient units, specialized in movement disorders, of central Italy. Inclusion criteria were diagnosis of PD and any current pharmacological treatment of PD while exclusion criteria were evident cognitive or depressive impairment, infusion with dopamine agonists or Duodopa, or Deep Brain Stimulation therapy. Patients were asked to complete the Italian version of WOQ-19 before the neurological visit. A medical form for the collection of demographic and clinical data of patients and for the evaluation of comprehensibility and usability the WOQ-19 was filled by the neurologist during the visit. RESULTS: Our data confirmed that WOQ-19 was able to identify WO in 69% of patients, a percentage similar to the recently reported in the Italian WOQ-19 validation study. Motor symptoms were more frequent than non-motor symptoms (80% vs. 20%). Patients who experienced WO had a higher age of PD onset, more severe disease, longer disease duration and were more likely to be female. CONCLUSIONS: The WOQ-19 was understandable for the patient, easily administered and suitable for routine outpatient use. It could be also particularly useful in clinical practice in the early identification of non-motor symptoms, often under reported by patients and revealed only with clinical support.

Functional and structural balances of homologous sensorimotor regions in multiple sclerosis fatigue.

Fatigue in multiple sclerosis (MS) is a highly disabling symptom. Among the central mechanisms behind it, an involvement of sensorimotor networks is clearly evident from structural and functional studies. We aimed at assessing whether functional/structural balances of homologous sensorimotor regions-known to be crucial for sensorimotor networks effectiveness-decrease with MS fatigue increase. Functional connectivity measures at rest and during a simple motor task (weak handgrip of either the right or left hand) were derived from primary sensorimotor areas electroencephalographic recordings in 27 mildly disabled MS patients. Structural MRI-derived inter-hemispheric asymmetries included the cortical thickness of Rolandic regions and the volume of thalami. Fatigue symptoms increased together with the functional inter-hemispheric imbalance of sensorimotor homologous areas activities at rest and during movement, in absence of any appreciable parenchymal asymmetries. This finding supports the development of compensative interventions that may revert these neuronal activity imbalances to relieve fatigue in MS.

Inflammation and iron metabolism in adult patients with epilepsy: does a link exist?

PURPOSE: Inflammation has been shown to play a key role in epilepsy, and may also affect both the iron status and metabolism. Consequently, a relationship between iron metabolism and neuronal excitability and seizures could be expected. METHODS: We aimed at characterizing in 37 adult patients affected by focal epilepsy during the interictal period serum inflammatory cytokines, such as interleukin 6 (IL-6), IL-6 soluble receptor (IL6-sR), interleukin 1 (IL-1), IL-1 receptor-antagonist (IL-1RA), tumor necrosis factor-α (TNF-α), and markers of iron status and metabolism: hemoglobin concentration (Hgb), mean corpuscular volume (MCV), hematocrit (Hct) red blood cell (RBC) count, serum iron and copper concentrations, ceruloplasmin (iCp), the ceruloplasmin enzymatic activity (eCp), the specific ceruloplasmin activity (eCp/iCp), total ferroxidase activity, transferrin (Tf), serum ferritin (SF), Tf saturation (Sat-Tf), and ratio of ceruloplasmin to transferrin (Cp/Tf). We investigated the correlations between these biological markers as well their relationship with patients' clinical features. A group of 43 healthy subjects had the same serologic measurements to serve as controls. RESULTS: Our findings showed in the group of patients with epilepsy an increase of IL-6 (p=0.026) and a decrease of TNF-α (p=0.002) with respect to healthy subjects. For the first time, we also detected significant changes in iron metabolism as an increase of Cp/Tf (p=0.011) and a decrease of Tf (p=0.031), possibly driven by cytokine modifications and consistent with inflammation as acute phase and antioxidant activity markers. Accordingly, TNF-α positively correlated with Tf (p=0.005). Finally, a significant positive correlation between seizures frequency and eCp (p=0.046) and inversely with Hgb (p=0.038) and Hct (p=0.041), and an inverse correlation between TNF-α and the duration of epilepsy (p=0.021) was detected. CONCLUSIONS: Our findings demonstrate a relevant relationship between epilepsy and systemic inflammation, with a consistent link between seizures, inflammatory cytokines (IL-6 and TNF-α) and iron regulation and metabolism, as acute phase and antioxidant markers.

Effects of hemochromatosis and transferrin gene mutations on peripheral iron dyshomeostasis in mild cognitive impairment and Alzheimer's and Parkinson's diseases.

Deregulation of iron metabolism has been observed in patients with neurodegenerative diseases. We have carried out a molecular analysis investigating the interaction between iron specific gene variants [transferrin (TF, P589S), hemochromatosis (HFE) C282Y and (H63D)], iron biochemical variables [iron, Tf, ceruloplasmin (Cp), Cp:Tf ratio and % of Tf saturation (% Tf-sat)] and apolipoprotein E (APOE) gene variants in 139 Alzheimer's disease (AD), 27 Mild Cognitive Impairment (MCI), 78 Parkinson's disease (PD) patients and 139 healthy controls to investigate mechanisms of iron regulation or toxicity. No difference in genetic variant distributions between patients and controls was found in our Italian sample, but the stratification for the APOEε4 allele revealed that among the APOEε4 carriers was higher the frequency of those carriers of at least a mutated TF P589S allele. Decreased Tf in both AD and MCI and increased Cp:Tf ratio in AD vs. controls were detected. A multinomial logistic regression model revealed that increased iron and Cp:Tf ratio and being man instead of woman increased the risk of having PD, that increased values of Cp:Tf ratio corresponded to a 4-fold increase of the relative risk of having MCI, while higher Cp levels were protective for PD and MCI. Our study has some limitations: the small size of the samples, one ethnic group considered, the rarity of some alleles which prevent the statistical power of some genetic analysis. Even though they need confirmation in larger cohorts, our data suggest the hypothesis that deregulation of iron metabolism, in addition to other factors, has some effect on the PD disease risk.

Longitudinal prognostic value of serum "free" copper in patients with Alzheimer disease.

BACKGROUND: Serum copper not bound to ceruloplasmin ("free") appears slightly elevated in patients with Alzheimer disease (AD). We explored whether a deregulation of the free copper pool can predict AD clinical worsening. METHODS: We assessed levels of copper, iron, zinc, transferrin, ceruloplasmin, peroxides, total antioxidant capacity, free copper, and apolipoprotein E genotype in 81 patients with mild or moderate AD, mean age 74.4, SD = 7.4 years, clinically followed up after 1 year. The association among biologic variables under study and Mini-Mental State Examination (MMSE) (primary outcome), activities of daily living (ADL), and instrumental activities of daily living (IADL) (secondary outcomes) performed at study entry and after 1 year were analyzed by multiple regression. RESULTS: Free copper predicted the annual change in MMSE, adjusted for the baseline MMSE by means of a linear regression model: it raised the explained variance from 2.4% (with only sex, age, and education) to 8.5% (p = 0.026). When the annual change in MMSE was divided into < 3 or > or = 3 points, free copper was the only predictor of a more severe decline (predicted probability of MMSE worsening 23%: odds ratio = 1.23; 95% confidence interval = 1.03-1.47; p = 0.022). Hyperlipidemic patients with higher levels of free copper seemed more prone to worse cognitive impairment. Free copper at baseline correlated with the ADL and IADL clinical scales scores at 1 year. CONCLUSIONS: These results show an association between copper deregulation and unfavorable evolution of cognitive function in Alzheimer disease. Further research is needed to establish whether copper is an independent risk factor for cognitive decline.

'Free' copper in serum of Alzheimer's disease patients correlates with markers of liver function.

Non-ceruloplasmin bound copper ('free') seems slightly elevated in Alzheimer's disease (AD) patients. To test the hypothesis of a correlation between 'free' copper and liver function in AD. We evaluated 51 AD patients and 53 controls through typical tests for chronic liver disease (AST, ALT, gamma-GT, Albumin, prothrombin time - PT-, bilirubins), along with copper, ceruloplasmin, iron, cholesterol in the serum and apolipoprotein E epsilon4 (APOE4) genotype. Absolute serum copper and 'free' copper were higher, albumin was lower and PT longer in AD patients than in controls. 'Free' copper correlated negatively with markers of liver function, in that albumin and albumin/PT ratio (r = -0.43, p = 0.004), and positively with direct bilirubin. Copper and 'free' copper were higher in the APOE4 carriers. These results suggest that abnormalities in copper metabolism might have an effect on liver function in AD.

What is the relationship among atherosclerosis markers, apolipoprotein E polymorphism and dementia?

Evidence suggests the important role of vascular factors both in vascular dementia (VaD) and Alzheimer disease (AD) pathogenesis. However, the relationship between apolipoprotein E (APOE) polymorphism and markers of atherosclerosis is still controversial. The aim of the study was to investigate the interplay between APOE polymorphisms and atherosclerosis in patients with AD and VaD. In this cross-sectional study, 101 demented (68 AD and 33 VaD) patients underwent APOE genotyping and neck vessel ultrasound to evaluate carotid artery disease [intima-media thickness (IMT) and plaques]. Patients with AD carrying epsilon4 allele presented increased IMT values with respect to non-epsilon4 carriers and VaD patients, whereas no relation was found between APOE polymorphisms and the presence or grade of carotid plaques both in AD and VaD patients. The epsilon4 APOE allele may promote intima-media thickening, interacting with other factors contributing to AD development.

Excess of nonceruloplasmin serum copper in AD correlates with MMSE, CSF [beta]-amyloid, and h-tau.

OBJECTIVE: To assess whether serum copper in Alzheimer disease (AD) correlates with cognitive scores, beta-amyloid, and other CSF markers of neurodegeneration. METHODS: The authors studied copper, ceruloplasmin, total peroxide, and antioxidants levels (TRAP) in serum; beta-amyloid in plasma; and copper, beta-amyloid, h-tau, and P-tau in the CSF of 28 patients with AD and 25 healthy controls, in relation to clinical status. RESULTS: Serum copper (p < 0.0001), peroxides (p = 0.002), a copper fraction unexplained by ceruloplasmin (p < 0.0001), and CSF h-tau (p = 0.001) were increased in AD, whereas serum TRAP (p = 0.03) and CSF beta-amyloid were decreased (p < 0.0001). Plasma beta-amyloid increased with age in healthy controls (r = 0.6; p = 0.05). CSF markers of AD correlated with serum copper variables. CSF copper was partially dependent on the serum copper fraction unexplained by ceruloplasmin (t = 2.2, p = 0.04). CSF beta-amyloid seemed to be related to serum copper (r = -0.46; p = 0.002). Mini-Mental Status Examination scores correlated positively with beta-amyloid (r = 0.46, p = 0.002) and inversely with copper unexplained by ceruloplasmin (r = -0.45, p = 0.003). CONCLUSIONS: The authors' results confirm the existence of changes in copper component distribution, particularly the copper fraction unexplained by ceruloplasmin and support the hypothesis of a beta-amyloid and copper connection in Alzheimer disease.

Repetitive transcranial magnetic stimulation (rTMS) at high and low frequency: an efficacious therapy for major drug-resistant depression?

OBJECTIVE: Repetitive transcranial magnetic stimulation (rTMS) is proposed for the treatment of drug-resistant depression. Studies performed in accordance with evidence-based medicine (EBM) are scarce, particularly in seeking optimal treatment and evaluation parameters. We aimed to test various types of rTMS in a large sample of depressed patients following EBM rules and to investigate treatment-related changes in plasma levels of neurotransmitters involved in depression. METHODS: Seventy-one drug-resistant depressed patients were randomly assigned to low (1 Hz) or high (17 Hz) rate TMS, applied for 5 days over the left dorsolateral prefrontal cortex (L-DLPFC). Patients were separated into two study designs. One group (20 patients) received only active treatment, while the other entered a double-blind, placebo-controlled, crossover design. Pre- and post-treatment blood samples were taken for evaluation of plasma levels of dopamine and serotonin. RESULTS: After a week of treatment patients had a measurable benefit. However, overall the placebo stimulation did not differ significantly from real stimulation, nor were differences observed between the two rates of rTMS. The only difference emerged when the real stimulation was applied at 17 Hz following placebo treatment. Plasma levels of neurotransmitters between active and placebo rTMS were similar. CONCLUSIONS: Using the treatment schedule of 1 week, although a clinical improvement after active treatment was indeed observed, this was both clinically and biochemically indistinguishable from that seen in the placebo arm. SIGNIFICANCE: This suggests that most of the previous emphasis, for short period of treatment, should be tempered down and that further work is required in order to verify whether optimal stimulation and evaluation parameters for TMS-treatment of depression beyond the placebo effect may be found following EBM rules.

Allelic variation in the human prodynorphin gene promoter and schizophrenia.

Experimental and clinical studies suggest an involvement of the opioid neuropeptide system in schizophrenia. In particular, the prodynorphin (PDYN), the precursor of the dynorphin opioid peptides, has been shown to play an important role in several aspects of human mental diseases. Recently, a functional polymorphism in the promoter of PDYN gene has been described. We studied the possible relationship between this polymorphism and schizophrenia and we found no significant difference in allelic and genotype distributions between schizophrenic patients and control subjects. However, we observed a significant interactive effect with the receptor 3 of dopamine gene (DRD3); in particular, the frequency of subjects carrying PDYN allele 3 being also homozygotes for DRD3 Gly allele (of Ser9Gly polymorphism) was significantly greater in patients than controls. We conclude that PDYN gene polymorphism alone does not alter the risk for schizophrenia but, by an epistatic interaction with the Gly allele of DRD3 gene, may contribute to the susceptibility to this disorder.

Degradation of imazosulfuron in soil.

Imazosulfuron, 1-(2-chloroimidazo[1,2-alpha]pyridin-3-ylsulfonyl)-3- (4,6-dimethoxypyrimidin-2-yl)urea, is a new post-emergence sulfonylurea herbicide applied once per growing season, highly active at low application levels, used to control most annual and perennial broad-leaf weeds and sedges in paddy rice (75-95 g AI ha-1) and turf (500-1000 g AI ha-1), and characterised by low mammalian toxicity. The degradation of imazosulfuron in a sandy loam soil (10-15 cm depth) has been investigated under aerobic and anaerobic laboratory conditions to evaluate its likely environmental fate. Test soils were treated with imazosulfuron in acetonitrile to give a final concentration of 0.2 mg kg-1 (100 g ha-1). The treated soils were extracted with methylene chloride + acetonitrile + 1 M ammonium hydroxide (4 + 15 + 1 by volume) and analysed by HPLC. Recoveries from spiked soils ranged from 80% to 95% (RSD 0.5-2.5%; n = 3) for imazosulfuron and its metabolites 2-chloroimidazo[1,2-alpha]pyridin-3-sulfonamide (IPSN), 1-(2-chloroimidazo[1,2-alpha]pyridin-3-ylsulfonyl)-3-(4-hydroxy-6- methoxypyrimidin-2-yl)urea (HMS) and 2-amino-4,6-dimethoxypyrimidine (ADPM). The limits of quantitation ranged from 0.001 mg kg-1 to 0.005 mg kg-1. Chemical cleavage of the sulfonylurea bond to give ADPM and IPSN was the main degradation pathway in aerobic conditions, whereas a remarkable production of HMS under anaerobic conditions suggested that degradation was due to micro-organisms which had the ability to demethylate imazosulfuron. In aerobic and anaerobic conditions, imazosulfuron disappeared from the soil with a half-life of approximately 70 (90% CL 62-75; r2 = 0.98) and 4 (90% CL 3.2-4.2; r2 = 0.97) days, respectively.

Liquid chromatographic determination of imazosulfuron in drinking water and in soil using ultraviolet detection.

Reversed-phase liquid chromatography (LC) is used to determine a relatively new sulfonylureic herbicide, imazosulfuron, 1-(2-chloroimidazo-[1,2-a] pyridin-3-ylsulfonyl)-3-(4,6-dimethoxy-2-pyrimidinyl)-urea (TH-913), in drinking water and in soil. TH-913 is extracted from water using solid-phase extraction on C18 bonded silica. Soil samples (20 g) are extracted with 300 ml of methanol-water (50:50) and the acidified extracts are transferred onto Sep-Pak C18 and processed as described for water samples. Off-line desorption is done with 20 ml of methanol-water (50:50). The eluate is evaporated to dryness, the residue dissolved in acetonitrile and analysed by LC with UV detection at 238 nm. The recoveries of TH-913 from water were over 95% (at 0.05 microg/l level) and from soil over 90% (at 0.005 mg/kg level).

Phase II of doxorubicin/taxol in metastatic breast cancer. Argentine Multicenter Taxol Group.

PURPOSE: To assess the response rate, survival, and toxicity of Taxol (paclitaxel) as 1-h infusion plus doxorubicin as first-line treatment for patients with metastatic breast cancer (MBC). PATIENTS AND METHODS: Seventy-six patients with untreated MBC were recruited. All of them had measurable disease and were evaluable for toxicity. Fifty-five percent of the patients had visceral involvement. The dose of doxorubicin was fixed at 50 mg/m2 as a short intravenous infusion, followed by 200 mg/m2 of Taxol as a 1-h intravenous infusion. Doxorubicin was administered during the first seven cycles, continuing with Taxol only up to a maximum of ten cycles. RESULTS: Neutropenia was the most important toxicity: 30% grade 3 and 18% grade 4. Only 2 patients showed a decrease in the left ventricular ejection fraction (LVEF) which caused discontinuing the treatment. No clinical congestive heart failure (CHF) was observed. Seventy-four patients were eligible for response evaluation: 10 (14%) achieved complete response (CR) and 46 (62%) achieved partial response (PR). The mean duration of response was 13.47+/-1.35 months (95% confidence interval (CI): 10.82; 16.12) and the mean survival was 21.50+/-1.42 months (95% CI: 18.72; 24.29). CONCLUSION: The overall response (OR) rate was 76%. No CHF was assessed and 2 patients stopped treatment due to LVEF decrease. Although doxorubicin 50 mg/m2 followed by Taxol 200 mg/m2 in 1-h intravenous infusion presents a toxicity profile which demands a close follow-up, it represents a convenient outpatient schedule with similar activity rate compared to longer Taxol infusions.

Uracil/tegafur plus oral calcium folinate in advanced breast cancer.

Uracil and tegafur (in a molar ratio of 4:1 [UFT]) has proven activity against breast cancer and is delivered in an easy-to-administer oral formulation. Orzel, which combines UFT with the oral biomodulator, calcium folinate, may provide even greater antitumor efficacy against breast cancer. Here, we describe the preliminary results of this phase II trial investigating the feasibility of 250 mg/m2/day of UFT plus 45 mg/day of oral calcium folinate administered to highly pretreated patients with advanced breast cancer. The results indicate a highly tolerable regimen and an overall response rate of 27.8% in a group of poor-prognosis patients. These findings warrant continued investigation.

Occurrence and metabolism of anandamide and related acyl-ethanolamides in ovaries of the sea urchin Paracentrotus lividus.

Cannabinoid receptors have been described in sea urchin sperm and shown to mediate inhibition of sperm acrosome reaction. Anandamide (arachidonoyl-ethanolamide), the mammalian physiological ligand at the cannabinoid CB1 receptor, has been subsequently found to effect this inhibition. Here we present data showing that ovaries from the sea urchin Paracentrotus lividus contain anandamide and two related acyl-ethanolamides, as well as enzymatic activities potentially responsible for their biosynthesis and degradation. Pilot experiments carried out with either ovaries or spermatozoa, extracted from both P. lividus and Arbacea lixula and radiolabelled with [14C]ethanolamine, showed that in sexually mature ovaries of both species significant levels of radioactivity were incorporated into a lipid component with the same chromatographic behaviour as anandamide. Lipid extracts from P. lividus ovaries were purified and analysed by gas chromatography/mass spectrometry which showed the presence of low but measurable amounts of anandamide, palmitoyl- and stearoyl-ethanolamides. The extracts were also found to contain lipid components with the same chromatographic behaviour as the N-acyl-phosphatidyl-ethanolamines, the phospholipid precursors of acyl-ethanolamides in mammalian tissues, and capable of releasing anandamide, palmitoyl- and stearoyl-ethanolamides upon digestion with S. chromofuscus phospholipase D. Accordingly, whole homogenates from P. lividus contained an enzymatic activity capable of converting synthetic [3H]N-arachidonoyl-phosphatidyl-ethanolamine into [3H]anandamide. Finally, mature ovaries of P. lividus were shown also to contain an amidohydrolase activity which catalyses the hydrolysis of anandamide and palmitoyl-ethanolamide to ethanolamine. This enzyme displayed subcellular distribution, pH/temperature dependency profiles and sensitivity to inhibitors similar but not identical to those of the previously described 'anandamide amidohydrolase' from mammalian tissues. These data support the hypothesis, formulated in previous studies, that anandamide or related metabolites may be oocyte-derived cannabimimetic regulators of sea urchin fertility.

Novel diterpenoid diacylglycerols from marine molluscs: potent morphogens and protein kinase C activators.

Five novel 1,2-sn-diacylglycerols with diterpenoid acyl moieties in the sn-1 position were isolated and characterized, together with the corresponding 1,3-sn-diacylglycerols, from three species of dorid nudibranchs molluscs. Their potent activity as morphogens in vivo in the Hydra tentacle regeneration assay and their parallel activity as activators of rat brain protein kinase C (PKC) in vitro are reported here. Our findings promote the use of these compounds as useful molecular probes for both in vivo and in vitro studies on the participation of PKC in cell development.

High performance liquid chromatographic analysis of quinolone antibacterial agents.

Eleven quinolone antibacterials have been analyzed by HPLC. The effects of different pH values and organic modifiers on chromatographic behaviour are discussed. Moreover the influence on separation mechanism of different counter-ions at various concentrations in the eluent has been investigated. Two different chromatographic methods have been selected, that allow the simultaneous analysis of quinolones containing or not a piperazinyl moiety, respectively.

Application of solid-phase partition cartridges in the determination of fungicide residues in vegetable samples.

Disposable, ready-to-use cartridges filled with a macroporous diatomaceous material are used to extract in a single step fungicide residues with dichloromethane from aqueous acetone extracts of vegetables. This procedure takes the place of some functions (such as separating funnel partition, drying over anhydrous sodium sulphate and clean-up) usually performed by separate steps in classical schemes. Fourteen fungicides (dichloran, vinclozolin, chlorthalonil, triadimefon, dichlofluanide, procymidone, hexaconazole, captan, folpet, ditalimfos, iprodione, captafol, pyrazophos and fenarimol) were determined using the described procedure with recoveries between 83 and 107% at spiking levels ranging for the different compounds from 0.04 to 0.40 mg/kg. Crops subjected to the described procedure included lettuce, strawberry, apple, yellow pepper and peach, and gave extracts containing a mass of co-extractives between 5 and 30 mg. Compared with classical schemes, the described procedure is simple, less labour intensive, allows parallel handling of several extracts and does not require preparation and maintenance of equipment. Troublesome emulsions such as those frequently observed in separating funnel partitioning do not occur.