Expression patterns of the immune checkpoint ligand CD276 in urothelial carcinoma.

BACKGROUND: CD276 is an immune checkpoint molecule. Elevated CD276 expression by urothelial carcinoma is associated with poor prognosis, but little is known about its expression across different tumor stages. We therefore investigated CD276 expression in bladder cancer (BC) cells and in tissue samples of BC stages from pT2 to pT4. METHODS: CD276 expression was explored in 4 urothelial cancer cell lines and 4 primary normal urothelial cell populations by quantitative RT-PCR, Western blot and flow cytometry. CD276 was investigated in bladder tumors from 98 patients by immunohistochemistry using a score (0-300) incorporating both, staining intensity and area of CD276 staining. Normal appearing urothelium in the bladder of the same patients served as controls. RESULTS: The urothelial carcinoma cell lines expressed significantly higher levels of CD276 on transcript (p < 0.006), total protein levels (p < 0.005), and on the cell surface (p < 0.02) when compared to normal urothelial cells. In pT2-T4 tumor tissue samples, CD276 was overexpressed (median score 185) when compared to corresponding healthy tissues from the same patients (median score 50; p < 0.001). No significant differences in CD276 expression were recorded in late, locally advanced ≥ pT3a tumors (median score 185) versus organ-confined < pT3a tumors (median score 190), but it was significantly lower in the normal urothelial tissue associated with ≥ pT3a tumors (median score 40) versus < pT3a tumors (median score 80; p < 0.05). CONCLUSION: CD276 expression is significantly elevated in urothelial carcinoma cells in all stages but varies between individuals considerably. Reduced CD276 expression in normal urothelial cells may imply that these cells would be protected from CD276-mediated immuno therapies.

Enzymatic Reactions in a Lab-on-Valve System: Cholesterol Evaluations.

The micro sequential injection analysis / lab-on-valve (µSIA-LOV) system is a miniaturized SIA system resulting from the implementation of a lab-on-valve (LOV) atop of the selection valve. It integrates the detection cell and the sample processing channels into the same device, promoting the reduction of reagent consumption and waste generation, the improvement of the versatility, and the reduction of the time of analysis. All of these characteristics are really relevant to the implementation of enzymatic reactions. Additionally, the evaluation of cholesterol in serum samples is widely relevant in clinical diagnosis, since higher values of cholesterol in human blood are actually an important risk factor for cardiovascular problems. An automatic methodology was developed based on the µSIA-LOV system in order to evaluate its advantages in the implementation of enzymatic reactions performed by cholesterol esterase, cholesterol oxidase and peroxidase. Considering these reactions, the developed methodology was also used for the evaluation of cholesterol in human serum samples, showing reliable and accurate results. The developed methodology presented detection and quantification limits of 1.36 and 4.53 mg dL-1 and a linear range up to 40 mg dL-1. This work confirmed that this µSIA-LOV system is a simple, rapid, versatile, and robust analytical tool for the automatic implementation of enzymatic reactions performed by cholesterol esterase, cholesterol oxidase, and peroxidase. It is also a useful alternative methodology for the routine determinations of cholesterol in real samples, even when compared with other automatic methodologies.

Assessment of essential elements and chemical contaminants in thirteen fish species from the Bay Aratu, Bahia, Brasil / Avaliação de elementos químicos essenciais e contaminantes em treze espécies de peixes da Baía de Aratu, Bahia, Brasil

Abstract Concentrations of ten elements (Cd, Cr, Cu, Fe, Ni, Pb, Se, Sr, V and Zn) were determinate in muscle tissues of 13 fish species from Aratu Bay, Bahia, Brazil by inductively coupled plasma optical emission spectrometry. The accuracy and precision of our results were checked by using two certified reference materials: BCR-422 cod muscle and SRM 1566b oyster tissue. The average trace element concentrations in the fish species varied in the following ranges, in μg g–1: 0.03-0.8 for Cr; 2.0-33.7 for Cu, 2.4-135.1 for Fe, 1.6-25.6 for Se; 1.6-35.1 for Sr; and 2.8-40.5 for Zn. The Diaptereus rhombeus (carapeba) specie presented the highest concentrations of Se, Cu and Fe. Chromium and Se were present at levels above the limit of tolerance allowed by the National Agency of Sanitary Vigilance (ANVISA). The results were also evaluated using the multivariate analysis techniques: principal component analysis (PCA) and hierarchical cluster analysis (HCA).

Resumo Foram determinadas as concentrações de dez elementos (Cd, Cr, Cu, Fe, Ni, Pb, Se, Sr, V and Zn) em tecidos de músculos de treze espécies de peixes da Baía de Aratu, Bahia, Brasil, utilizando a espectrometria de emissão ótica com plasma indutivamente acoplado. A precisão e exatidão dos resultados foram analisadas utilizando dois materiais de referência certificada: BCR-422 cod muscle e o SRM 1566b oyster tissue. A concentração média dos elementos traço nas espécies de peixes variaram nas seguintes faixas, em μg g−1: Cr = 0,03-0,8; Cu = 2,0-33,7, Fe = 2,4-135,1, Se = 1,6-25,6; Sr = 1,6-35,1; Zn = 2.8-40.5. A espécie Diaptereus rhombeus (carapeba) apresentou as maiores concentrações de Se, Cu e Fe. Cromo e Se apresentaram níveis acima do limite de tolerância estabelecido pela Agência Nacional de Vigilância Sanitária (ANVISA). Os resultados ainda foram avaliados usando técnicas de análises multivariadas: análise de componente principal (PCA) e análise de agrupamento hierárquico (HCA).

Assessment of essential elements and chemical contaminants in thirteen fish species from the Bay Aratu, Bahia, Brasil.

Concentrations of ten elements (Cd, Cr, Cu, Fe, Ni, Pb, Se, Sr, V and Zn) were determinate in muscle tissues of 13 fish species from Aratu Bay, Bahia, Brazil by inductively coupled plasma optical emission spectrometry. The accuracy and precision of our results were checked by using two certified reference materials: BCR-422 cod muscle and SRM 1566b oyster tissue. The average trace element concentrations in the fish species varied in the following ranges, in µg g-1: 0.03-0.8 for Cr; 2.0-33.7 for Cu, 2.4-135.1 for Fe, 1.6-25.6 for Se; 1.6-35.1 for Sr; and 2.8-40.5 for Zn. The Diaptereus rhombeus (carapeba) specie presented the highest concentrations of Se, Cu and Fe. Chromium and Se were present at levels above the limit of tolerance allowed by the National Agency of Sanitary Vigilance (ANVISA). The results were also evaluated using the multivariate analysis techniques: principal component analysis (PCA) and hierarchical cluster analysis (HCA).

Distribution of trace elements in tissues of shrimp species Litopenaeus vannamei (Boone, 1931) from Bahia, Brazil.

In this study, concentrations of trace elements in tissues of shrimp species (Litopenaeus vannamei) from farming and zone natural coastal located in the northeastern Brazil were investigated. The elements determination was performed by optical emission spectrometry with inductively coupled plasma (ICP OES). The following ranges of concentrations in the tissues were obtained in µg g-1 dry weight: Al: 13.4-886.5, Cd: 0.93-1.80; Cu: 24.8-152; Fe: 3.2-410.9; Mn: 0.36-24.4; Se: 0.094-9.81 and Zn: 20.3-109.4. The shrimp muscle can be a good iron source (about 88.9 mg-1g dry weight). The distribution of Se concentration in tissues showed much variation between locations, and the concentration levels found in shrimp muscles of wild samples were high, where its levels in 67% of muscle and 50% of others tissues samples exceeded the ANVISA limit, indicating evidence of selenium bioaccumulation. Significant correlation was observed between the following pairs of elements: Fe-Zn (r= -0.70), Mn-Cu (r= -0.74), Se-Cu (r= -0.68), Se-Mn (r= 0.82) in the muscles; Fe-Al (r= 0.99), Mn-Al (r= 0.62), Mn-Fe (r= 0.62), Se-Al (r = 0.88), Se-Fe (r= 0.87), Se-Mn (r= 0.58) in the exoskeleton and Cu-Zn (r = 0.68), Al-Cu (r= 0.88), Fe-Cu (r= 0.95) and Fe-Al (r= 0.97) in the viscera.

Distribution of trace elements in tissues of shrimp species Litopenaeus vannamei (Boone, 1931) from Bahia, Brazil / Distribuição de elementos traço em tecidos do camarão Litopenaeus vannamei (Boone, 1931) da Bahia, Brasil

Abstract In this study, concentrations of trace elements in tissues of shrimp species (Litopenaeus vannamei) from farming and zone natural coastal located in the northeastern Brazil were investigated. The elements determination was performed by optical emission spectrometry with inductively coupled plasma (ICP OES). The following ranges of concentrations in the tissues were obtained in µg g–1 dry weight: Al: 13.4-886.5, Cd: 0.93-1.80; Cu: 24.8-152; Fe: 3.2-410.9; Mn: 0.36-24.4; Se: 0.094-9.81 and Zn: 20.3-109.4. The shrimp muscle can be a good iron source (about 88.9 mg–1g dry weight). The distribution of Se concentration in tissues showed much variation between locations, and the concentration levels found in shrimp muscles of wild samples were high, where its levels in 67% of muscle and 50% of others tissues samples exceeded the ANVISA limit, indicating evidence of selenium bioaccumulation. Significant correlation was observed between the following pairs of elements: Fe-Zn (r= –0.70), Mn-Cu (r= –0.74), Se-Cu (r= –0.68), Se-Mn (r= 0.82) in the muscles; Fe-Al (r= 0.99), Mn-Al (r= 0.62), Mn-Fe (r= 0.62), Se-Al (r = 0.88), Se-Fe (r= 0.87), Se-Mn (r= 0.58) in the exoskeleton and Cu-Zn (r = 0.68), Al-Cu (r= 0.88), Fe-Cu (r= 0.95) and Fe-Al (r= 0.97) in the viscera.

Resumo Esse estudo teve como objetivo avaliar as concentrações de elementos traço em tecidos da espécie de camarão Litopenaeus vannamei coletadas da zona costeira e de carciniculturas localizadas no nordeste do Brasil. Os elementos químicos foram determinados por espectrômetro de emissão óptica com plasma indutivamente acoplado (ICP OES). Foram encontradas as seguintes faixas de concentrações desses elementos nos tecidos (em mg g–1 peso seco): Al: 13,4-886,5; Cd: 0,93-1,80; Cu: 24,8-152; Fe: 3,2-4109; Mn: 0,36-24,4; Se: 0,094-9,81 and Zn: 20,3-109,4. O músculo do camarão investigado pode ser uma boa fonte de ferro (cerca de 88.9 mg-1g peso seco). A distribuição da concentração de Se nos tecidos apresentou muita variação entre as localidades, com níveis acima do estabelecido pela ANVISA para 67% dos musculos e 50% dos outros tecidos investigados, indicando evidências de bioacumulação do selênio. Houve correlações significativas entre os seguintes pares de elementos:: Fe-Zn (r= –0,70), Mn-Cu (r= –0,74), Se-Cu (r= –0,68), Se-Mn (r= 0,82) nos músculos, Fe-Al (r= 0,99), Mn-Al e Mn-Fe (r= 0,62), Se-Al (r = 0,88), Se-Fe (r= 0,87), Se-Mn (r= 0,58) no exoesqueleto e Cu-Zn (r = 068), Al-Cu (r= 0,88), Fe-Cu (r= 0,95) and Fe-Al (r= 0,97) nas vísceras.

Distribution of glial cells in the auditory brainstem: normal development and effects of unilateral lesion.

Auditory brainstem networks facilitate sound source localization through binaural integration. A key component of this circuitry is the projection from the ventral cochlear nucleus (VCN) to the medial nucleus of the trapezoid body (MNTB), a relay nucleus that provides inhibition to the superior olivary complex. This strictly contralateral projection terminates in the large calyx of Held synapse. The formation of this pathway requires spatiotemporal coordination of cues that promote cell maturation, axon growth, and synaptogenesis. Here we have examined the emergence of distinct classes of glial cells, which are known to function in development and in response to injury. Immunofluorescence for several astrocyte markers revealed unique expression patterns. Aldehyde dehydrogenase 1 family member L1 (ALDH1L1) was expressed earliest in both nuclei, followed by S100ß, during the first postnatal week. Glial fibrillary acidic protein (GFAP) expression was seen in the second postnatal week. GFAP-positive cell bodies remained outside the boundaries of VCN and MNTB, with a limited number of labeled fibers penetrating into the margins of the nuclei. Oligodendrocyte transcription factor 2 (OLIG2) expression revealed the presence of oligodendrocytes in VCN and MNTB from birth until after hearing onset. In addition, ionized calcium binding adaptor molecule 1 (IBA1)-positive microglia were observed after the first postnatal week. Following hearing onset, all glial populations were found in MNTB. We then determined the distribution of glial cells following early (P2) unilateral cochlear removal, which results in formation of ectopic projections from the intact VCN to ipsilateral MNTB. We found that following perturbation, astrocytic markers showed expression near the ectopic ipsilateral calyx. Taken together, the developmental expression patterns are consistent with a role for glial cells in the maturation of the calyx of Held and suggest that these cells may have a similar role in maturation of lesion-induced connections.

Focused microwave-induced combustion for digestion of botanical samples and metals determination by ICP OES and ICP-MS.

The advantages and shortcomings of focused microwave-induced combustion (FMIC) for digestion of plant samples were studied. The effects of sample mass, absorbing solution, oxygen gas flow-rate, and time of reflux step on recoveries of major, minor and trace metals were systematically evaluated. Afterwards, Al, Ba, Ca, Co, Cr, Cu, Mg, Mn, Ni, Sr, V, and Zn were determined by inductively coupled plasma optical emission spectrometry (ICP OES) and by inductively coupled plasma mass spectrometry (ICP-MS). The main advantages of FMIC when compared to microwave-assisted wet digestion (MAWD) and focused-microwave-assisted wet digestion (FMAWD) are the possibility to digest larger masses of samples (up to 3g) using shorter heating times and diluted nitric acid solution for absorbing all analytes. Using the selected experimental conditions for FMIC, residual carbon content was lower than 0.7% for all samples and relative standard deviation (RSD) varied from 1.5 to 14.1%. Certified reference materials (NIST 1515 apple leaves and NIST 1547 peach leaves) were used for checking accuracy and determined values for all metals were in agreement with certified values at a 95% confidence level. No statistical difference (ANOVA, 95% of confidence level) was observed for results obtained by FMIC, FMAWD, and MAWD. Limits of detection were lower when using FMIC in the range of 0.02-0.15 µg g(-1) for ICP OES and 0.001-0.01 µg g(-1) for ICP-MS, which were about 3 and 6 times lower than the values obtained by FMAWD and MAWD, respectively. It is important to point out that FMIC was a suitable sample preparation method for major, minor and trace metals by both determination techniques (ICP OES and ICP-MS). Additionally, since it allows lower LODs (because up to 3g of sample can be digested) and diluted acid solutions are used (without any further dilution), the use of ICP-MS is not mandatory.

Flow methodology for methanol determination in biodiesel exploiting membrane-based extraction.

A methodology based in flow analysis and membrane-based extraction has been applied to the determination of methanol in biodiesel samples. A hydrophilic membrane was used to perform the liquid-liquid extraction in the system with the organic sample fed to the donor side of the membrane and the methanol transfer to an aqueous acceptor buffer solution. The quantification of the methanol was then achieved in aqueous solution by the combined use of immobilised alcohol oxidase (AOD), soluble peroxidase and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS). The optimization of parameters such as the type of membrane, the groove volume and configuration of the membrane unit, the appropriate organic solvent, sample injection volume, as well as immobilised packed AOD reactor was performed. Two dynamic analytical working ranges were achieved, up to 0.015% and up to 0.200% (m/m) methanol concentrations, just by changing the volume of acceptor aqueous solution. Detection limits of 0.0002% (m/m) and 0.007% (m/m) methanol were estimated, respectively. The decision limit (CCalpha) and the detection capacity (CCbeta) were 0.206 and 0.211% (m/m), respectively. The developed methodology showed good precision, with a relative standard deviation (R.S.D.) <5.0% (n=10). Biodiesel samples from different sources were then directly analyzed without any sample pre-treatment. Statistical evaluation showed good compliance, for a 95% confidence level, between the results obtained with the flow system and those furnished by the gas chromatography reference method. The proposed methodology turns out to be more environmental friendly and cost-effective than the reference method.

Newly generated dentate granule cells from epileptic rats exhibit elongated hilar basal dendrites that align along GFAP-immunolabeled processes.

Previous studies showed that neurogenesis occurs in the dentate gyrus of the adult rodent. Recent evidence suggests that the resulting newly born neurons integrate into pre-existing hippocampal circuitry. Newly born neurons in the developing and adult dentate gyrus exhibit a transient basal dendrite. In adult pilocarpine-induced epileptic rats, basal dendrites persist and are ectopically located in the hilus where they receive synaptic input from mossy fiber axons. We hypothesize that these hilar basal dendrites are derived from newly born neurons that are born after the pilocarpine-induced seizures. To test this hypothesis, the length of basal dendrites from epileptic rats was compared with that from control rats using doublecortin immunocytochemistry, which labels newly born neurons and their processes for up to 3 weeks after their genesis. The data on hilar basal dendrites in pilocarpine animals indicate that those from newly born neurons are significantly longer than those found in the control rats. We also demonstrate that 20% of newly born neurons in the epileptic rat have a basal dendrite that enters the hilus at an angle greater than 30 degrees from its cell body as compared with <2% in the control rats. Lastly, we provide evidence that the hilar basal dendrites in the epileptic rats are adjacent to glial fibrillary acidic protein-labeled astrocytic processes in the hilus and suggest that an ectopic glial scaffold in the hilus is involved with the formation of hilar basal dendrites. In conclusion, the data show that newly born neurons from epileptic rats have longer hilar basal dendrites and their formation might relate to gliosis which occurs as a result of hilar neuronal cell loss after status epilepticus.

Gene expression profiling in skeletal muscle of Zucker diabetic fatty rats: implications for a role of stearoyl-CoA desaturase 1 in insulin resistance.

AIMS/HYPOTHESIS: Insulin resistance in skeletal muscle is a hallmark of type 2 diabetes. Therefore, we sought to identify and validate genes involved in the development of insulin resistance in skeletal muscle. MATERIALS: Differentially regulated genes in skeletal muscle of male obese insulin-resistant, and lean insulin-sensitive Zucker diabetic fatty (ZDF) rats were determined using Affymetrix microarrays. Based on these data, various aspects of glucose disposal, insulin signalling and fatty acid composition were analysed in a muscle cell line overexpressing stearoyl-CoA desaturase 1 (SCD1). RESULTS: Gene expression profiling in insulin-resistant skeletal muscle revealed the most pronounced changes in gene expression for genes involved in lipid metabolism. Among these, Scd1 showed increased expression in insulin-resistant animals, correlating with increased amounts of palmitoleoyl-CoA. This was further investigated in a muscle cell line that overexpressed SCD1 and accumulated lipids, revealing impairments of glucose uptake and of different steps of the insulin signalling cascade. We also observed differential effects of high-glucose and fatty acid treatment on glucose uptake and long-chain fatty acyl-CoA profiles, and in particular an accumulation of palmitoleoyl-CoA in cells overexpressing SCD1. CONCLUSIONS/INTERPRETATION: Insulin-resistant skeletal muscle of ZDF rats is characterised by a specific gene expression profile with increased levels of Scd1. An insulin-resistant phenotype similar to that obtained by treatment with palmitate and high glucose can be induced in vitro by overexpression of SCD1 in muscle cells. This supports the hypothesis that elevated SCD1 expression is a possible cause of insulin resistance and type 2 diabetes.

Metabisulphite-induced occupational asthma in a radiographer.

There is epidemiological evidence for an increased prevalence of occupational asthma among radiographers. However, the causes of darkroom disease are not yet defined. A 37-year-old female radiographer reported work-related asthma approximately 2 yrs after starting work in a local hospital. She was atopic and showed bronchial hyperresponsiveness to methacholine. Occupational-type exposure with a fixing agent, but not with a developer, produced an immediate-type asthmatic reaction. As the fixing agent contained sodium metabisulphite (SMBS), a substance known to cause asthma, bronchial challenges with SMBS were performed in the patient and nine asthmatic controls. The patient showed a positive bronchial immediate reaction on 2 separate days after inhalation of 48 and 96 microg SMBS, and one control also showed a significant fall in forced expiratory volume in one second after inhalation of 12 microg SMBS. The positive reaction in the control subject argues for a greater susceptibility of both persons to SMBS or its reaction product sulphur dioxide, rather than for a new occupational allergen. It is concluded that sodium metabisulphite exposure should be recognised as a cause of darkroom asthma.

Differential hepatic gene expression in a polygenic mouse model with insulin resistance and hyperglycemia: evidence for a combined transcriptional dysregulation of gluconeogenesis and fatty acid synthesis.

New Zealand obese (NZO) mice exhibit severe insulin resistance of hepatic glucose metabolism. In order to define its biochemical basis, we studied the differential expression of genes involved in hepatic glucose and lipid metabolism by microarray analysis. NZOxF1 (SJLxNZO) backcross mice were generated in order to obtain populations with heterogeneous metabolism but comparable genetic background. In these backcross mice, groups of controls (normoglycemic/normoinsulinemic), insulin-resistant (normoglycemic/hyperinsulinemic) and diabetic (hyperglycemic/hypoinsulinemic) mice were identified. At 22 weeks, mRNA was isolated from liver, converted to cDNA, and used for screening of two types of cDNA arrays (high-density filter arrays and Affymetrix oligonucleotide microarrays). Differential gene expression was ascertained and assessed by Northern blotting. The data indicate that hyperinsulinemia in the NZO mouse is associated with: (i) increased mRNA levels of enzymes involved in lipid synthesis (fatty acid synthase, malic enzyme, stearoyl-CoA desaturase) or fatty acid oxidation (cytochrome P450 4A14, ketoacyl-CoA thiolase, acyl-CoA oxidase), (ii) induction of the key glycolytic enzyme pyruvate kinase, and (iii) increased mRNA levels of the gluconeogenic enzyme phosphoenolpyruvate carboxykinase. These effects were enhanced by a high-fat diet. In conclusion, the pattern of gene expression in insulin-resistant NZO mice appears to reflect a dissociation of the effects of insulin on genes involved in glucose and lipid metabolism. The data are consistent with a hypothetical scenario in which an insulin-resistant hepatic glucose production produces hyperinsulinemia, and an enhanced insulin- and substrate-driven lipogenesis further aggravates the deleterious insulin resistance of glucose metabolism.

[Airways and lung disease by metal working fluids]. / Atemwegs- und Lungenerkrankungen durch Kühlschmierstoffe.

Metal working fluids (MWF) are used in the manufacturing process to cool and lubricate machined parts. MWF contain the coolant, mostly water-oil emulsions with soluble semi-synthetic or synthetic oils, but also emulsifiers and additives. Reaction products and both bacterial and fungal contamination occur during use. In the present review we listed literature data of dose-effect-relationships with a focus on biological substances. Most studies describe obstructive airways disease due to irritant effects of MWF. Immunoglobulin E mediated airways disease or hypersensitivity pneumonitis have rarely been described. There is no doubt that MWF exert adverse effects on human airways, but there is few data concerning dose-effect relationships. The role of bacterial and fungal contamination or endotoxin exposure cannot be defined by the available information. However, there is some evidence that biological components in MWF may represent the most hazardous substances.

Changes in low molecular weight DNA fragmentation in white blood cells after diisocyanate exposure of workers.

The pathogenesis of diisocyanate-induced asthma is still largely unknown. Recently, it has been shown that thiol-redox homeostasis of human airway epithelial cells may be altered after in vitro exposure to diisocyanates. In the present study, low molecular weight (LMW) DNA fragmentation patterns in white blood cells (WBCs) were assessed on 16 industrial workers with work-related asthma, before and after chamber challenges with one of three commonly used diisocyanates in concentrations up to 30 ppb. LMW-DNA fragmentation changes were evaluated after 15 h incubation of WBCs embedded in agarose plugs in lysis buffer with or without hydrogen peroxide (H2O2). Increased LMW-DNA fragmentation occurred in WBCs taken at 30 min or 19 h after the end of the chamber challenge in both subjects with positive and in 8 of 14 subjects with negative challenges. In contrast, no change in LMW-DNA fragmentation was seen in WBCs taken at the same time intervals from 11 non-exposed controls. There was no association between changes in DNA fragmentation patterns and possible confounding factors such as age, smoking status, atopy, medication, duration of occupational exposure and period since exposure cessation. These results indicate that diisocyanate exposure can induce DNA fragmentation. Similarities in the increased amounts of WBC LMW-DNA fragments following diisocyanate exposure with the DNA fragmentation after plugs lysis in buffer with H2O2 support the hypothesis that diisocyanates change the intracellular redox steady-state. Whether this effect plays any role in isocyanate-induced asthma has to be investigated in larger epidemiological studies.

Evaluation of long-term toxicity in patients after cisplatin-based chemotherapy for non-seminomatous testicular cancer.

BACKGROUND: Because of the increasing number of long-term survivors of metastatic testicular germ-cell cancer, a general concern has been secondary morbidities, especially cardiovascular risk factors. PATIENTS AND METHODS: Thirty-two patients treated with cisplatin- and doxorubicin-containing chemotherapy > or = 13 years before the time of analyses were evaluated for neuro-, oto-, pulmonary-, vascular- and gonadal toxicity including evaluation of myocardial damage and cardiovascular risk factors and analysis of microcirculation. RESULTS: Thirty percent of the patients showed abnormal left ventricle function. Elevated follicle stimulating hormone (FSH) and luteinising hormone (LH) levels in 75% of patients were often associated with low testosterone levels. Elevated total cholesterol levels were found in 82% and higher triglyceride levels in 44% of patients, most of them were overweight. About 25% of the patients developed diastolic arterial hypertension after chemotherapy. Reduced hearing was confirmed in 23% of patients, especially at frequencies higher than 3000 Hz. Moreover, 53% of patients presented transient evoked otoacoustic emissions. In 38% of patients non-symptomatic neuropathy was detected, in 28% symptomatic neuropathy, and in 6% disabling polyneuropathy. In 80% of patients with neuropathic symptoms additional morphological and functional abnormalities were found by nailfold capillary videomicroscopy, compared to only 57% of the patients without neuropathic symptoms. CONCLUSIONS: Patients cured by cisplatin-based chemotherapy for metastatic testicular cancer have to be cognizant of their unfavorable cardiovascular risk profile, that might be a greater risk than developing a relapse or second malignancy.

Carboxyl residues in the iron-sulfur protein are involved in the proton pumping activity of P. denitrificans bc(1) complex.

A study is presented on chemical modification of the three subunit Paracoccus denitrificans bc(1) complex. N-(Ethoxycarbonyl)-2-ethoxy-1,2-dihydroquinoline (EEDQ) treatment caused a loss of the proton pumping activity of liposome-reconstituted bc(1) complex. A similar effect, which is referred to as the decoupling effect, resulted upon reaction of N,N'-dicyclohexylcarbodiimide (DCCD) with the complex. Direct measurement of the binding of EEDQ to the complex subunits, performed in the presence of the fluorescent hydrophobic nucleophile 4'-[(aminoacetamido)methyl]fluorescein (AMF), showed that the iron-sulfur protein (ISP) and cytochrome c(1) were labeled by EEDQ, whereas cytochrome b was not. Tryptic digestion and sequencing analysis of the fluorescent fragment of the ISP revealed this to consist of a segment with six acidic residues, among which the highly conserved aspartate 160 is present. Analogous experiments on DCCD binding showed that all the three subunits of the complex were labeled. However, DCCD concentration dependence of carboxyl residue modification in the individual subunits and of proton pumping activity showed that the decrease of the H(+)/e(-) ratio correlated only with the modification of the ISP. Tryptic digestion of labeled ISP and sequencing analysis of the fluorescent fragment gave results superimposable upon those obtained with EEDQ. Chymotryptic digestion and sequencing analysis of the single fluorescent fragment of cytochrome b showed that this fragment contained glutamate 174 and aspartate 187. We conclude that, in the P. denitrificans bc(1) complex, carboxyl residues in cytochrome b do not appear to be critically involved in the proton pump mechanism of the complex.

Transcriptional activation of TRADD mediates p53-independent radiation-induced apoptosis of glioma cells.

Survival of patients with Glioblastoma Multiforme (GM), a highly malignant brain tumor, remains poor despite concerted efforts to improve therapy. The median survival of patients with GM has remained approximately 1 year regardless of the therapeutic approach. Since radiation therapy is the most effective adjuvant therapy for GM and nearly half of GM tumors harbor p53 mutations, we sought to identify genes that mediate p53-independent apoptosis of GM cells in response to ionizing radiation. Using broad-scale gene expression analysis we found that following radiation treatment, TRADD expression was induced in a uniquely radiosensitive GM cell line but not in radioresistant GM cell lines. TRADD over-expression killed GM cells and activated NF-kappa B. We found that blocking the TRADD-mediated pathway using a dominant-negative mutant of FADD (FADD-DN) enhanced radiation resistance of GM cells, as reflected in both susceptibility to apoptosis and clonogenic survival following irradiation. Conversely, stable expression of exogenous TRADD enhanced radiation-induced apoptosis of GM cell lines, reflecting the biological significance of TRADD regulation in p53-independent apoptosis. These findings generate interest in utilizing TRADD in gene therapy for GM tumors, particularly in light of its dual function of directly inducing rapid apoptosis and sensitizing GM cells to standard anti-neoplastic therapy.

Bacillus subtilis spore film dosimeters in personal dosimetry for occupational solar ultraviolet exposure.

OBJECTIVE: Ultraviolet (UV) radiation is noted to be one of the most important risk factors for nonmelanoma and melanoma skin cancers. The recent development of a spore film test chamber containing spores of Bacillus subtilis resulted in a new method of UV measurement with a spectral sensitivity profile similar to erythema-weighted data calculated from spectroradiometric measurements. METHODS: The practical application of dosimeters was tested on 11 persons for 43 days, under different conditions of UV exposure in five different geographical regions. Four professional lifeguards at a public swimming pool carried dosimeters attached to their shoulders or to their caps, for 11 days. Three mountain guides attached dosimeters laterally to their heads on 27 different occasions of mountaineering activity in different mountain regions. Four ski instructors carried lateral head dosimeters during eight days of skiing in the Alps. RESULTS: The life guards received daily UV exposures ranging from 3.6 to 9.5 minimal erythema doses (MED) (mean 5.9, SD +/- 1.9). The mountain guides had personal daily UV exposures of from 4.4 to 17.1 MED (11.9 +/- 3.9) and ski instructors from 2.8 to 8.8 MED (6.1 +/- 1.8). CONCLUSIONS: Bacillus subtilis spore film dosimeters can be applied effectively for personal solar UV measurements of occupationally exposed persons. such as lifeguards. mountain guides and ski instructors. UV levels in these occupations exceed international limits of exposure.