Reproducibility of food atopy patch tests over time in the general child population

Background  The atopy patch test (APT) is no longer an experimental method; it is increasingly being used as a standard diagnostic tool for the characterization of patients with aeroallergen- and food-triggered disorders. Some technical aspects of this test still remain to be answered. We aimed to study the reproducibility of this test over time in the general child population.
Methods  In a general population of 118 children, we investigated the reproducibility of duplicate APTs with four food allergens in their native form, which were repeated at set intervals from the first test: 7 days (group 1), 14 days (group 2), and 21 days (group 3).
Results  We observed very poor reproducibility on both sides of the back in all three studied subgroups. The reproducibility rates and Cohen's κ values did not improve when we did not consider the side of the back. There were no differences in the prevalence of atopy between the subjects with reproducible and nonreproducible APT results. All three groups studied showed no difference in the prevalence rates of atopy. There was no relationship between APT and skin prick test positivity for the same allergen. Questionnaire-derived data about previous food-related reactions did not help in the evaluation of the doubtful nonreproducible APT results with food allergens.
Conclusions  Our results show that the reproducibility of food APTs is poor and unsatisfactory over time, and there is an urgent need for the development of optimal, stable, and good-quality APT testing substances.     

Organization of the state space of a simple recurrent network before and after training on recursive linguistic structures.

Recurrent neural networks are often employed in the cognitive science community to process symbol sequences that represent various natural language structures. The aim is to study possible neural mechanisms of language processing and aid in development of artificial language processing systems. We used data sets containing recursive linguistic structures and trained the Elman simple recurrent network (SRN) for the next-symbol prediction task. Concentrating on neuron activation clusters in the recurrent layer of SRN we investigate the network state space organization before and after training. Given a SRN and a training stream, we construct predictive models, called neural prediction machines, that directly employ the state space dynamics of the network. We demonstrate two important properties of representations of recursive symbol series in the SRN. First, the clusters of recurrent activations emerging before training are meaningful and correspond to Markov prediction contexts. We show that prediction states that naturally arise in the SRN initialized with small random weights approximately correspond to states of Variable Memory Length Markov Models (VLMM) based on individual symbols (i.e. words). Second, we demonstrate that during training, the SRN reorganizes its state space according to word categories and their grammatical subcategories, and the next-symbol prediction is again based on the VLMM strategy. However, after training, the prediction is based on word categories and their grammatical subcategories rather than individual words. Our conclusion holds for small depths of recursions that are comparable to human performances. The methods of SRN training and analysis of its state space introduced in this paper are of a general nature and can be used for investigation of processing of any other symbol time series by means of SRN.     

Induction of biologically active antineutrophil cytoplasmic antibodies by immunization with human apoptotic polymorphonuclear leukocytes

Translocation of intracellular components to the cell surface during the priming or apoptosis of polymorphonuclear leukocytes (PMN) is an important mechanism for interaction of antineutrophil cytoplasmic antibodies (ANCA) with these antigens. To test the capacity of apoptotic PMN to trigger production of ANCA, six groups of mice were immunized with either live or apoptotic lymphocytes, or with live, apoptotic, formalin-fixed, or lysed PMN. Mice immunized with both live and apoptotic neutrophils developed high titers of antibodies which gave a granular cytoplasmic immunofluorescent pattern. These antibodies were specific for lactoferrin and myeloperoxidase. Following a second intravenous infusion of apoptotic PMNs, mice developed anti-PR3 antibodies. Vasculitis lesions were not found in mice which developed ANCA. The ANCA-containing IgG fraction induced superoxide production by human PMNs. These results support the hypothesis that neutrophil-specific antigens presented on the cell membranes of apoptotic PMN may induce ANCA in the proper conditions.     

MIR204 n.37C>T variant as a cause of chorioretinal dystrophy variably associated with iris coloboma,early-onset cataracts and congenital glaucoma

Four members of a three-generation Czech family with early-onset chorioretinal dystrophy were shown to be heterozygous carriers of the n.37C>T in MIR204. The identification of this previously reported pathogenic variant confirms the existence of a distinct clinical entity caused by a sequence change in MIR204. Chorioretinal dystrophy was variably associated with iris coloboma, congenital glaucoma, and premature cataracts extending the phenotypic range of the condition. In silico analysis of the n.37C>T variant revealed 713 novel targets. Additionally, four family members were shown to be affected by albinism resulting from biallelic pathogenic OCA2 variants. Haplotype analysis excluded relatedness with the original family reported to harbour the n.37C>T variant in MIR204. Identification of a second independent family confirms the existence of a distinct MIR204-associated clinical entity and suggests that the phenotype may also involve congenital glaucoma.     

The impairment of lysyl oxidase in keratoconus and in keratoconus-associated disorders

Keratoconus (KC) is an eye disease characterized by the progressive thinning and protrusion of the cornea, which results in the loss of visual acuity. This disorder remains poorly understood, although recent studies indicate the involvement of genetic and environmental factors. Recently, we have found that the distribution of the cross-linking enzyme lysyl oxidase (LOX) is markedly decreased in about 63 % of keratoconic specimens. Similarly, LOX activity is significantly reduced by 38 % compared to control tissue. Nearly 70 systemic disorders have been reported in association with KC, most of them affecting the extracellular matrix. In this review we attempted to ascertain whether any KC-associated diseases exhibit signs that may reflect LOX impairment. We hypothesized that very similar changes in the extracellular matrix, particularly at the level of collagen metabolism, including LOX impairment in mitral leaflets, may reflect an association between KC and mitral valve prolapse. Moreover, this putative association is supported by the high frequency of Down syndrome in both diseases. Among other disorders that have been found to coincide with KC, we did not find any in which the LOX enzyme may be directly or indirectly impaired. On the other hand, in cases where KC is present along with other connective tissue disorders (Marfan syndrome, Ehlers–Danlos syndrome and others), KC may not arise as a localized manifestation, but rather may be induced as the result of a more complex connective tissue disorder.     

Surface characterization of dental Y-TZP ceramic after air abrasion treatment

Objective

The aim of this study was to characterize the surface of Y-TZP after abrasion with various airborne particles.

Methods

The Y-TZP blanks were cut into 44 discs and sintered according to the manufacturer's instructions. The specimens were treated as follows: (a) control specimens, (b) abraded with 50 μm alumina, (c) abraded with 110 μm alumina, (d) abraded with 30 μm silica-coated alumina, (e) abraded with 110 μm silica-coated alumina, (f) abraded with 110 μm alumina followed by 110 μm silica-coated alumina particles. Airborne abrasion was performed at a pressure of 2.5 bar for 15 s/cm². The Y-TZP was characterized using X-ray photoelectron spectroscopy (XPS), field emission scanning electron microscopy (FESEM) and X-ray diffraction analysis (XRD).

Results

Surface morphology of Y-TZP ceramic was changed after the airborne abrasion process compared to the control specimens. The grain boundaries disappeared and part of the airborne particles are embedded and/or rested on the ceramic surfaces. The elemental composition of the Y-TZP surface after the airborne abrasion process depended on the type and size of these particles. The concentration of Si resulted higher after the airborne abrasion process with 110 μm alumina followed by 110 μm silica-coated alumina particles in comparison to the specimens abraded with 110 μm silica-coated alumina particles. The ratio of elements normalized by yttrium for these specimens was: [Zr]/[Y]/[Al]/[Si] = 15.2/1.0/26.0/73.6, respectively.

Conclusion

The change of grain topography occurred during each impact process. Silica nano-particles covered not only loosely the abraded ceramic surface after abrasion process, but the release of kinetic energy in form of thermal energy resulted in melting of the ceramic surface and in the formation of zirconium silicate.     

Human Hepatic Alcohol Dehydrogenase and Human Erythrocyte Catalase Do Not Metabolize the Cytochrome P-4502E1 Substrate, Chlorzoxazone

Studies of cytochrome P-4502E1 (CYP2E1)-mediated oxidation of ethanol have been hampered by the lack of a suitable probe for in vivo human studies. Chlorzoxazone, a prescribed skeletal muscle relaxant, is metabolized to 6-hydroxychlorzoxazone by CYP2E1 and has been advocated as a specific probe of this enzyme on the basis of microsomal studies. The applications of this probe may include delineating the contribution of CYP2E1 to in vivo human ethanol metabolism. However, the activity of nonmicrosomal enzymes in metabolizing chlorzoxazone is unknown. Alcohol dehydrogenase (ADH), predominantly a hepatic cytosolic enzyme, may be more important than CYP2E1 in the oxidation of ethanol to acetaldehyde. The contribution of catalase in the in vivo oxidation of ethanol to acetaldehyde is controversial. To determine if either of these enzymes metabolizes chlorzoxazone and whether ethanol oxidation by either enzyme is inhibited by chlorzoxazone or its metabolite, multiple in vitro studies were performed. ADH enzyme kinetics were performed with human recombinant β₁β₁ and β₃β₃ ADH with ethanol and chlorzoxazone (0.5 to 2.5 mM). Neither ADH isoenzyme exhibited NAD⁺-dependent oxidation of chlorzoxazone, but displayed Michaelis-Menten kinetics for ethanol with K_m values of 89 μM and 34 mM, for β₁β₁, and β₃β₃, respectively. Typical in vivo concentrations of chlorzoxazone and its metabolite, 6-hydroxychlorzoxazone, did not alter β₁β₁, or β₃β₃ ADH-mediated oxidation of ethanol to acetaldehyde. Studies of human hepatic nonmicrosomal enzyme activity were expanded to include all nonmicrosomal NAD⁺-dependent hepatic enzymes by starch gel electrophoresis assessment. Human hepatic enzymatic activity in the presence of chlorzoxazone was similar to that observed in the control sample (no added substrate), suggesting a lack of metabolism by NAD⁺-dependent enzymes. Similarly, human erythrocyte catalase, in the presence of a hydrogen peroxide generating system, did not metabolize chlorzoxazone. Furthermore, neither chlorzoxazone nor 6-hydroxychlorzoxazone altered the catalase-induced formation of acetaldehyde from ethanol. These data are consistent with chlorzoxazone as a specific probe of CYP2E1 that may be useful to alcohol researchers.     

Cytotoxicity of acetylcholinesterase reactivators evaluated in vitro and its relation to their structure

The development of acetylcholinesterase reactivators, i.e., antidotes against organophosphorus poisoning, is an important goal of defense research. The aim of this study was to compare cytotoxicity and chemical structure of five currently available oximes (pralidoxime, trimedoxime, obidoxime, methoxime, and asoxime) together with four perspective oximes from K-series (K027, K074, K075, and K203). The cytotoxicity of tested substances was measured using two methods – colorimetric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay and impedance based real-time cytotoxicity assay – in three different cell lines (HepG2, ACHN, and NHLF). Toxicity was subsequently expressed as toxicological index IC₅₀. The tested compounds showed different cytotoxicity ranging from 0.92 to 40.06?mM. In HepG2 cells, K027 was the least and asoxime was the most toxic reactivator. In ACHN and NHLF cell lines, trimedoxime was the compound with the lowest adverse effects, whereas the highest toxicity was found in methoxime-treated cells. The results show that at least five structural features affect the reactivators’ toxicity such as the number of oxime groups in the molecule, their position on pyridinium ring, the length of carbon linker, and the oxygen substitution or insertion of the double bond into the connection chain. Newly synthetized oximes with IC₅₀?≥?1?mM evaluated in this three cell lines model might appear suitable for further testing.     

Standardized extracts of flavonoids increase the viability of PC12 cells treated with hydrogen peroxide: effects on oxidative injury

Oxidative stress plays an important role in cell death associated with many diseases. In the present study, concentration-dependence of hydrogen peroxide on rat pheochromocytoma (PC12) cell viability was studied. Preventive effects of antioxidants on the viability of these cells treated with 2 mM hydrogen peroxide were compared. Trolox and Stobadine, as chain-breaking antioxidants were studied in comparison with standardized extracts of flavonoids of Ginkgo biloba and Pycnogenol, known as agents effective in several diseases. All antioxidants increased the viability of hydrogen peroxide-treated PC12 cells. Flavonoid extracts were more effective than Trolox and Stobadine. Antioxidants were most effective if present after the oxidative treatment. As expected, the preloading with antioxidants was without effect on cell viability. Correlations between viability increase induced by antioxidants, and content of oxidation products of proteins and lipids were studied at concentrations of antioxidants mostly effective in preventing cell death: Trolox (10 microM), Stobadine (30 microM), Ginkgo biloba (160 microg/ml), Pycnogenol (100 microg/ml). In these concentrations, antioxidants did not statistically significantly decrease the content of protein carbonyls, with exception of Stobadine, which had no effect. Ginkgo biloba, Trolox and Stobadine intensively decreased the content of malondialdehyde, a product of lipid peroxidation. Pycnogenol was without any preventive effect. Concentrations of antioxidants with a large effect on viability of PC12 cells were not effective in preventing oxygen radical-induced injury of proteins. Antioxidants prevented the oxidative injury of lipids more effectively than that of proteins.