Restless legs syndrome in Wilson's disease: frequency, characteristics, and mimics.

OBJECTIVE: To determine characteristics, clinical significance, frequency, and mimics of restless legs syndrome (RLS) in a cohort of Wilson's disease (WD, n = 42/f = 18), compared to healthy, matched controls. MATERIALS AND METHODS: Structured clinical interviews (patients and caregiving family members), repeated neurological examinations (afternoon and presleep), comprehensive laboratory tests, WD-, RLS-, and sleep-specific rating scales, and video-polysomnography. RESULTS: Thirteen patients with WD (13/42 = 31.0%) clearly fulfilled the five diagnostic criteria of RLS; in eight patients (19.1%), the burden of RLS was clinically significant. The RLS was of moderate severity, equally distributed among sexes, manifested mainly in the evening and before falling asleep, and had developed mostly after clinical manifestation of WD (time elapsed 10.2 ± 14.5 years), still at a young mean age (27.5 ± 11.5 years). The known RLS-associated features were absent (normal iron and kidney parameters) or rare (positive family history, polyneuropathy). Compared to WD patients without RLS, patients with RLS were significantly elder and had suffered longer from WD. WD-specific RLS mimics as well as RLS confounding motor comorbidities (dystonia, tremor, chorea) were frequent and a diagnostic challenge; in difficult cases, the differentiation was reached by clinical observation of the motor behavior in the evening or at nighttime. CONCLUSION: RLS was frequent in this cohort of WD and might be causally related to WD. RLS should be included in the diagnostic work-up of WD. In complex motor disorders, differential diagnosis of RLS might require evening/nighttime examination and video-polysomnography. In WD patients with a clinically significant RLS, treatment with dopaminergic substances may be considered.

Interleukin-18 and interferon-gamma polymorphisms in Brazilian human immunodeficiency virus-1-infected patients presenting with lipodystrophy syndrome.

Cytokines play important roles in the pathogenesis of lipodystrophy syndrome (LS). Single nucleotide polymorphisms (SNPs) at positions -607(C/A) and -137(C/G) in the promoter region of the interleukin-18 (IL-18) gene and at position +874(T/A) of the interferon-gamma (IFN-gamma) gene are related to the expression of these cytokines. To examine whether IL-18 and IFN-gamma polymorphisms are associated with LS, these SNPs were genotyped in 88 human immunodeficiency virus (HIV)-infected patients presenting LS, 79 HIV-infected without LS, and 133 healthy controls. The -607A allele, -607AA genotype, and -137G/-607A and -137C/-607A haplotypes in the IL-18 gene were over-represented in HIV patients presenting LS. The -137G/-607C haplotype was associated with protection against LS. These results indicate that the -607(C/A) SNP is associated with LS development in HIV-infected patients.

TNF microsatellite alleles may confer protection against the development of lipodystrophy syndrome in Brazilian HIV patients.

The aim of this study was to evaluate the frequency of TNFa-e microsatellites and the promoter region (TNF-308 and TNF-238) in HIV/AIDS-infected patients presenting or not lipodystrophy syndrome (LS). The design is the genetic case-control association study. Microsatellite and the TNF promoter region polymorphisms were amplified by PCR and submitted to polyacrylamide gel electrophoresis. The genotypes and allele frequencies for 67 HIV-positive patients with lipodystrophy were compared with 50 HIV-positive patients with no evidence of lipodystrophy and with 131 healthy HIV-negative individuals. The presence of the TNFa5 allele could provide HIV/AIDS patients with protection against developing LS. The presence of TNF-308G allele, as well as of its homozygote TNF-308GG, were associated with susceptibility to developing LS. In addition, the presence of the haplotype TNFe3-d3-238G-308A-c1-a5-b7 suggests protection against developing that syndrome. This study highlights that polymorphic sites spanning the region nearby the TNF locus are associated with LS development in HIV/AIDS patients.

A genetic polymorphism of matrix metalloproteinase 9 (MMP-9) affects the changes in circulating MMP-9 levels induced by highly active antiretroviral therapy in HIV patients.

We examined whether two functional polymorphisms (g.-1562C>T and g.-90(CA)14-24) in the matrix metalloproteinase (MMP)-9 gene or MMP-9 haplotypes affect the circulating levels of pro-MMP-9 and pro-MMP-9/TIMP-1 (tissue inhibitor of metalloproteinase-1) ratios in AIDS patients, and modulate alterations in these biomarkers after highly active antiretroviral therapy (HAART). We studied 82 patients commencing HAART. Higher pro-MMP-9 concentrations and pro-MMP-9/TIMP-1 ratios were found in CT/TT patients compared with CC patients. HAART decreased pro-MMP-9 levels and pro-MMP-9/TIMP-1 ratios in CT/TT patients, it did not modify pro-MMP-9 levels and it increased pro-MMP-9/TIMP-1 ratios in CC patients. The g.-90(CA)14-24 polymorphism, however, produced no significant effects. Moreover, we found no significant differences in HAART-induced changes in plasma pro-MMP-9, TIMP-1 and pro-MMP-9/TIMP-1 ratios when different MMP-9 haplotypes were compared. These findings suggest that the g.-1562C>T polymorphism affects pro-MMP-9 levels in patients with AIDS and modulates the alterations in pro-MMP-9 levels caused by HAART, thus possibly affecting the risk of cardiovascular complications.

Protective role of γδ T cells in cigarette smoke and influenza infection.

Airborne pathogens commonly trigger severe respiratory failure or death in smokers with lung disease. Cigarette smoking compromises the effectiveness of innate immunity against infections but the underlying mechanisms responsible for defective acquired immune responses in smokers remains less clear. We found that mice exposed to chronic cigarette smoke recovered poorly from primary Influenza A pneumonia with reduced type I and II interferons (IFNs) and viral-specific immunoglobulins, but recruited γδ T cells to the lungs that predominantly expressed interleukin 17A (IL-17A). Il-17a-/- mice exposed to smoke and infected with Influenza A also recruited γδ T cells to the lungs, but in contrast to wild-type mice, expressed increased IFNs, made protective influenza-specific antibodies, and recovered from infection. Depletion of IL-17A with blocking antibodies significantly increased T-bet expression in γδ T cells and improved recovery from acute Influenza A infection in air, but not smoke-exposed mice. In contrast, when exposed to smoke, γδ T cell deficient mice failed to mount an effective immune response to Influenza A and showed increased mortality. Our findings demonstrate a protective role for γδ T cells in smokers and suggest that smoke-induced increase in IL-17A inhibits the transcriptional programs required for their optimal anti-viral responses. Cigarette smoke induces IL-17A expression in the lungs and inhibits γδ T-cell-mediated protective anti-viral immune responses.

Assessment of indicators of vitamin A status in non-cirrhotic chronic hepatitis C patients.

Subjects with chronic liver disease are susceptible to hypovitaminosis A due to several factors. Therefore, identifying patients with vitamin deficiency and a requirement for vitamin supplementation is important. Most studies assessing vitamin A in the context of hepatic disorders are conducted using cirrhotic patients. A cross-sectional study was conducted in 43 non-cirrhotic patients with chronic hepatitis C to evaluate markers of vitamin A status represented by serum retinol, liver retinol, and serum retinol-binding protein levels. We also performed the relative dose-response test, which provides an indirect estimate of hepatic vitamin A reserves. These vitamin A indicators were assessed according to the stage of liver fibrosis using the METAVIR score and the body mass index. The sample study was predominantly composed of male subjects (63%) with mild liver fibrosis (F1). The relative dose-response test was <20% in all subjects, indicating vitamin A sufficiency. Overweight or obese patients had higher serum retinol levels than those with a normal body mass index (2.6 and 1.9 µmol/L, respectively; P<0.01). Subjects with moderate liver fibrosis (F2) showed lower levels of serum retinol (1.9 vs 2.5 µmol/L, P=0.01) and retinol-binding protein levels compared with those with mild fibrosis (F1) (46.3 vs 67.7 µg/mL, P<0.01). These results suggested an effect of being overweight on serum retinol levels. Furthermore, more advanced stages of liver fibrosis were related to a decrease in serum vitamin A levels.

Late emergence of A594V and L595W mutations related to ganciclovir resistance in a patient with HCMV retinitis and long-term HIV progression.

The emergence of ganciclovir (GCV) resistance during the treatment of human cytomegalovirus (HCMV) infection is a serious clinical challenge, and is associated with high morbidity and mortality. In this case report, we describe the emergence of two consecutive mutations (A594V and L595W) related to GCV resistance in a patient with HCMV retinitis and long-term HIV progression after approximately 240 days of GCV use. Following the diagnosis of retinitis, the introduction of GCV did not result in viral load reduction. The detected mutations appeared late in the treatment, and we propose that other factors (high initial HCMV load, previous GCV exposure, low CD4+ cell count), in addition to the presence of resistance mutations, may have contributed to the treatment failure of HCMV infection in this patient.

Lectin-mediated effects on HIV type 1 infection in vitro.

Lectins with specificity for terminal mannose residues and anti-mannan antibodies neutralize HIV-1 infection in vitro. This is assumed to be caused by binding of the agents to the viral glycoproteins. In this study we show that one such agent, the Galanthus nivalis lectin (GNA), also blocks infection at the target cell level. To explore the effect of GNA on HIV infection we used the two HIV-1 isolates LAV and NDK, representing in the first case a prototype virus and in the latter case a highly cytopathic virus, which spreads preferentially via cell-to-cell contact. MT-4 cells were used as target cells and infection was determined from the occurrence of syncytia. Cell-to-cell infection was studied with CEM cells persistently infected with the two virus isolates. GNA, at concentrations in the nanogram per milliliter range, neutralized the HIV-1 isolates LAV, NDK, and MN as well as HIV-2ROD. Pretreatment of cells with the lectin, before addition of virus, or of infected cells, also blocked infection. This effect was more pronounced with HIV-1NDK than with HIV-1LAV. Mannosidase treatment of the target cells abolished the GNA effect on HIV-1NDK infection. It is concluded that GNA inhibits infection of several HIV isolates. It neutralizes infection by binding to the virion but also blocks infection at the target cell level. The latter effect may be different for different virus isolates. Mannosyl residuals at the cell surface are targets for GNA modulation of infection with the cytopathic HIV-1NDK. These do not represent essential virus receptors.

The use of approximation formulae in calculations of acid-base equilibria.

Details of diagrams recently presented for the regions where approximation formulae give the pH of mono- and diprotic acids with an error of less than 0.02 pH unit are discussed.

Acid-base ionization equilibria of 1-(2-carbamylethyl) imidazole and its 2-methyl and 2-ethyl derivatives.

The acid and base ionization constants of 1-(2-carbamylethyl)-2-alkylimidazoles, as well as the acid ionization constants of acrylamide and acetamide, have been determined by potentiometry. Concentration constants at 10.0, 25.0 and 40.0 degrees and I = 0.1M (KNO(3)) were measured. From plots of log K vs. 1/T the thermodynamic parameters (DeltaH and DeltaS) of the ionization reactions were calculated.

Simultaneous determination of the acid and basic ionization constants of imidazole.

The acid and base ionization constants of imidazole (LH) have been determined simultaneously by potentiometry. Concentration constants were obtained at 10.0, 15.0, 25.0 and 40.0 degrees with I = 0.1M (KNO(3)), the values (and standard deviations) at 25.0 degrees being pK(a1)(LH(2)(+)) = 7.002 +/- 0.006 and pK(a2)(LH(2)(+)) = 12.588 +/- 0.004. The following values were obtained for the corresponding thermodynamic parameters: DeltaH(1) =38 and DeltaH(2) = 38 kJ mole ; DeltaS(1) = -8 and DeltaS(2)= -112 J.mole(-1).K(-1).

Urea potentiometric biosensor based on urease immobilized on chitosan membranes.

Potentiometric biosensors based on urease (E.C. 3.5.1.5.) immobilized on chitosan membranes coupled to all-solid-state nonactin ammonium ion selective electrodes are described. The enzyme was immobilized on the chitosan membranes by four procedures: (A) adsorption; (B) adsorption followed by reticulation with dilute aqueous glutaraldehyde solution; (C) activation with glutaraldehyde followed by contact with the enzyme solution; and (D) activation with glutaraldehyde, contact with the enzyme solution and reduction of the Schiff base with sodium borohydride. The response characteristics of the biosensors obtained with these enzymatic membranes were determined and compared. The biosensor with best response characteristics, obtained by procedure (B), showed the following characteristics of response to urea: (i) linearity in the 10(-4) to 10(-2) M range; (ii) slope of up to 56 mV per decade; (iii) response time between 30 s and 2 min; and (iv) lifetime of 2 months. This biosensor was tested in the determination of urea in blood serum samples.

A combination of synchronous fluorescence spectroscopy with chemometric treatment and internal standards in non-aqueous potentiometric titrations of fulvic acids.

The acid properties of a soil fulvic acid (sfua) were characterized by potentiometric titration with tetrabutylammonium hydroxide in two non-aqueous solvents with high acid-base resolution power N,N-dimethylformamide (DMF) and acetonitrile. Synchronous fluorescence spectroscopy (SyF) was also used to monitor directly the sfua status during the potentiometric titration in DMF. The potentiometric titration curves showed no clear end-point and the analysis of the sets of spectra obtained at increasing neutralization degree, with a self-modeling curve resolution method (SIMPLISMA), revealed the existence of two components with featureless concentration profiles. Internal standards (maleic, salicylic and p-hydroxylbenzoic acids) were used to determine the amounts of acid groups with different acid strengths in the two non-aqueous solvents. It was shown that the variations observed in the SyF spectra sets of the internal standards are not correlated with those observed in the sfua data. The splitting of the sfua groups in the non-aqueous titration curves seems to be forced artificially depending on the standards used.

Monitoring of molecular transformations in acid-base reactions by evolving factor analysis of Fourier transform infrared spectral data.

An evolving factor analysis procedure with concentration constraints (gradient concentration window) was applied to the analysis of data sets of aqueous Fourier transform infrared (FT-IR) spectra of carboxylic acids (acetic, malonic and succinic acids) collected in experiments with varying pH. Besides the calculation of the number of acid-base systems, this procedure allowed the calculation of the FT-IR spectra of the acid-base species present in equilibrium as well as the corresponding pK(a) values.

Flow-through tubular iodide and bromide selective electrodes based on epoxy resin heterogeneous membranes.

Tubular all-solid-state iodide and bromide selective electrodes with channels drilled through the crystalline heterogeneous membranes have been prepared for use in flow-injection analysis (FIA). The membranes, made with AgX/Ag(2)S powdered mixtures (X = I, Br) dispersed in a non-conductive epoxy resin, were assembled inside a hollow cylindrical support of conductive epoxy resin. The response characteristics of these detectors, in a low-dispersion FIA system, have been evaluated. Both show a Nernstian response over the range between 5 x 10(-5) and O.1M X(-), good reproducibility and fast response, which allows a sampling rate of 60/hr. Operational pH ranges from 2.5 to 11 and from 3 to 10 were obtained for the iodide and bromide electrodes respectively. Iodide must be absent in determinations with the bromide electrode.

Conductimetric analysis of the ion binding properties of three leaf extracts of chestnut (Castanea sativa), Eucalyptus (Eucalyptus globulus) and oak (Quercus robur).

Humic materials extracted from tree leaves of chestnut (Castanea sativa), eucalyptus (Eucalyptus globulus) and oak (Quercus robur) were analyzed by performing conductimetric titrations. Values between about 84 and 236 muS cm(-1) for the molar conductivity and between 0.42 and 0.74 for the charge distribution parameter were obtained when the concentrations of the extract are increased from 40 to 100 mg 1(-1). These variations were explained by using the counterion condensation theory, and the distance between the charged groups of the polyions, the volume of the counterion condensation and the Debye-Hückel potential were also calculated.

Comparison of the simplex and Powell methods with a weighted response function for the optimization of FIA systems.

An experimental comparison between the relative performances of the Powell and simplex methods for the optimization of a Flow Injection Analysis (FIA) system for the determination of nitrite in water is reported. An evaluation of the advantages of using a weighted linear combination of two variables (related to sensitivity and sample throughput rate) as the response function in the guidance of the optimization procedures towards different practical requirements is also included. Both methods proved to be effective for the optimization, none having shown definite advantages over the other. The use of a weighted response function in these optimization methods proved to be useful for assessing the versatility of FIA systems.

Fluorescence quenching of anthropogenic fulvic acids by Cu(II), Fe(III) and UO(2)(2+).

The quenching of the fluorescence of three anthropogenic fulvic acids (FA) provoked by Cu(II) (pH 6.0), Fe(III) (pH 4.0) and UO(2)(2+) (pH 3.5), was analyzed by a non-linear method and by Stern-Volmer plots. The FA samples were extracted from composted sewage sludges (csFA), composted municipal wastes (mwFA) and composted livestock wastes (lsFA). Synchronous-scan fluorescence (SyF) spectra were collected as a function of metal ion concentration. Spectral data were treated by a self-modeling mixture analysis method (SIMPLISMA) to detect the SyF spectral band with the strongest quenching and to calculate the corresponding quenching profile. The analysis of these profiles by a non-linear method allowed the estimation of conditional stability constants (K) and of the percentage of non-complexing fluorophores. The same quantitative information was obtained by the modified Stern-Volmer equation taking into account the existence of fluorophores that do not participate in the complexation. Good agreement was found between the results of the two procedures. The logK calculated by the non-linear method were (standard deviation in parenthesis): csFA, Cu(II), 4.22 (5); Fe(III), 5.0 (1); UO(2)(2+), 5.2 (2); mwFA, Cu(II), 4.21 (3); Fe(III), 5.6 (2); UO(2)(2+), 4.7 (3); lsFA, Cu(II), 4.51 (8); Fe(III), 5.5 (2); UO(2)(2+), 3.6 (2).

Reduced anti-Mycobacterium tuberculosis antibody response in tuberculosis patients with acquired immunodeficiency syndrome.

1. When the sera of patients with tuberculosis were tested for anti-Mycobacterium tuberculosis IgG using an indirect ELISA, the test was positive for 94.1% of the samples from patients not having AIDS (N = 51), but for only 37.5% of the samples from patients with AIDS (N = 16). 2. False-positive results were obtained for 7.3% of patients not infected with HIV (N = 96) and for 4.7% of patients infected with HIV (N = 64). 3. In most serum samples obtained from patients with tuberculosis and AIDS after the beginning of specific treatment there was a reduction of the ELISA absorbance at 490 nm with time. 4. These results indicate that serological tests for the detection of anti-M. tuberculosis IgG in patients with AIDS are of limited value for the diagnosis of tuberculosis, most likely as a consequence of the underlying immune defect of the patients.

Sediments as monitors of heavy metal contamination in the Ave river basin (Portugal): multivariate analysis of data.

The concentrations of heavy metals (Cd, Cr, Cu, Ni, Pb, Zn) were determined in river sediments collected at the Ave river basin (Portugal) to obtain a general classification scenery of the pollution in this highly polluted region. Multivariate data analysis techniques of clustering, principal components and eigenvector projections were used in this classification. Five general areas with different polluting characteristics were detected and several individual heavy metal concentration abnormalities were detected in restricted areas. A good correlation between the overall metal contamination determined by multivariate analysis and metal pollution indexes for all sampling stations was obtained. Some preliminary experiments showed that the metal concentrations normalised to the volatile matter content in the sediment fraction with grain size <63 microm seems to be an adequate method for assessing metal pollution.