Invasive Lactococcus lactis producing mycobacterial Hsp65 ameliorates intestinal inflammation in acute TNBS-induced colitis in mice by increasing the levels of the cytokine IL-10 and secretory IgA.

AIMS: To investigate the anti-inflammatory activity of an invasive and Hp65-producing strain Lactococcus lactis NCDO2118 FnBPA+ (pXYCYT:Hsp65) in acute 2,4,6-trinitrobenzene sulphonic acid (TNBS)-induced colitis in mice as an innovative therapeutic strategy against Crohn's disease (CD). METHODS AND RESULTS: The pXYCYT:Hsp65 plasmid was transformed into the L. lactis NCDO2118 FnBPA+ strain, resulting in the L. lactis NCDO2118 FnBPA+ (pXYCYT:Hsp65) strain. Then, the functionality of the strain was evaluated in vitro for Hsp65 production by Western blotting and for invasion into Caco-2 cells. The results demonstrated that the strain was able to produce Hsp65 and efficiently invade eukaryotic cells. Subsequently, in vivo, the anti-inflammatory capacity of the recombinant strain was evaluated in colitis induced with TNBS in BALB/c mice. Oral administration of the recombinant strain was able to attenuated the severity of colitis by mainly reducing IL-12 and IL-17 levels and increasing IL-10 and secretory immunoglobulin A levels. CONCLUSIONS: The L. lactis NCDO2118 FnBPA+ (pXYCYT:Hsp65) strain contributed to a reduction in inflammatory damage in experimental CD. SIGNIFICANCE AND IMPACT OF THE STUDY: This study, which used L. lactis for the production and delivery of Hsp65, has scientific relevance because it shows the efficacy of this new strategy based on therapeutic protein delivery into mammalian enterocytes.

Lactococcus lactis carrying a DNA vaccine coding for the ESAT-6 antigen increases IL-17 cytokine secretion and boosts the BCG vaccine immune response.

AIMS: A regimen utilizing Bacille Calmette-Guerin (BCG) and another vaccine system as a booster may represent a promising strategy for the development of an efficient tuberculosis vaccine for adults. In a previous work, we confirmed the ability of Lactococcus lactis fibronectin-binding protein A (FnBPA+) (pValac:ESAT-6), a live mucosal DNA vaccine, to produce a specific immune response in mice after oral immunization. In this study, we examined the immunogenicity of this strain as a booster for the BCG vaccine in mice. METHODS AND RESULTS: After immunization, cytokine and immunoglobulin profiles were measured. The BCG prime L. lactis FnBPA+ (pValac:ESAT-6) boost group was the most responsive group, with a significant increase in splenic pro-inflammatory cytokines IL-17, IFN-γ, IL-6 and TNF-α compared with the negative control. CONCLUSIONS: Based on the results obtained here, we demonstrated that L. lactis FnBPA+ (pValac:ESAT-6) was able to increase the BCG vaccine general immune response. SIGNIFICANCE AND IMPACT OF THE STUDY: This work is of great scientific and social importance because it represents the first step towards the development of a booster to the BCG vaccine using L. lactis as a DNA delivery system.

Simvastatin modulates gene expression of key receptors in zebrafish embryos.

Nuclear receptors (NR) are involved in the regulation of several metabolic processes and it is well known that these constituents may be modulated by different chemicals classes, including pharmaceuticals that may activate or antagonize NR. In mammals, some pharmaceuticals modulate the transcription of pregnane X receptor, Pxr, peroxisome proliferator activated receptor, Ppars, and aryl hydrocarbon receptor, Ahr, affecting mRNA expression of genes belonging to various regulatory pathways, including lipid metabolism and detoxification mechanisms. The aim of this study was to determine the effects of simvastatin (SIM), an anticholesterolemic drug, on selected NR and AhR mRNA transcription levels during zebrafish early development. Embryos were collected at different development stages (0, 2, 6, 14, 24, 48, and 72 hr post fertilization (hpf)) and mRNA of all target NR was detected at all time points. Embryos (1 and 24 hpf) were exposed to different concentrations of SIM (5 or 50 µg/L) in two differing assays with varying exposure times (2 or 80 hr). The transcription levels of ahr2, raraa, rarab, rarga, pparαa, pparß1, pparγ, pxr, rxraa, rxrab, rxrbb, rxrga, rxrgb, as well as levels of cholesterol (Chol) were measured after exposure. SIM exerted no marked effect on Chol levels, and depending upon exposure duration mRNA levels of NR and AhR either increased or decreased. After 2 hr SIM treatment in 24 hpf embryos, transcription of ppars, pxr, and ahr was up-regulated, while after 80 hr mRNA levels of pxr and ahr were decreased with no marked changes in ppars. Data demonstrate that SIM produced alterations in gene expression of NR which are involved in varying physiological functions and that may disturb regulation of different physiological processes which might impair fish survival and ecosystems regeneration.

Effects of pharmaceuticals and personal care products (PPCPs) on multixenobiotic resistance (MXR) related efflux transporter activity in zebrafish (Danio rerio) embryos.

Certain ATP binding cassette (ABC) transporter proteins, such as zebrafish Abcb4, are efflux pumps acting as a cellular defence against a wide range of different, potentially toxic chemical compounds thus mediating so called multixenobiotic resistance (MXR). Certain chemicals target MXR proteins and, as so called chemosensitisers, inhibit the activity of these proteins thus increasing the toxicity of other chemicals that would normally be effluxed. In this study 14 pharmaceuticals and personal care products (PPCPs) that are being increasingly detected in aquatic systems, were assessed for interference with the MXR system of zebrafish (Danio rerio). Concentration dependent effects of test compounds were recorded with the dye accumulation assay using zebrafish embryos and in ATPase assays with recombinant zebrafish Abcb4. In the dye accumulation assay embryos at 24h post fertilisation (hpf) were exposed to 8µm rhodamine 123 along with test compounds for 2h. The rhodamine 123 tissue levels upon the exposure served as a measure for MXR transporter efflux activity of the embryo (low rhodamine levels - high activity; high levels - low activity). The known ABC protein inhibitors MK571, vinblastine and verapamil served as positive controls. All tested PPCPs affected rhodamine 123 accumulation in embryos. For seven compounds rhodamine tissue levels were either both decreased and increased depending on the compound concentration indicating both stimulation and inhibition of rhodamine 123 efflux by those compounds, only increased (inhibition, six compounds) or only decreased (stimulation, one compound). Recombinant zebrafish Abcb4 was obtained with the baculovirus expression system and PPCPs were tested for stimulation/inhibition of basal transporter ATPase activity and for inhibition of the transporter ATPase activity stimulated with verapamil. Eight of the tested PPCPs showed effects on Abcb4 ATPase activity indicating that their effects in the dye accumulation assay may have indeed resulted from interference with Abcb4-mediated rhodamine 123 efflux. Slight stimulatory effects were found for musk xylene, nerol, isoeugenol, α-amylcinnamaldehyde, α-hexylcinnamaldehyde and simvastatin indicating Abcb4 substrate/competitive inhibitor properties of those compounds. Likewise, decreases of the verapamil-stimulated Abcb4 ATPase activity by diclofenac and fluoxetine may indicate competitive transporter inhibition. Sertraline inhibited the basal and verapamil-stimulated Abcb4 ATPase activities suggesting its property as non-competitive Abcb4 inhibitor. Taken together, our finding that chemically diverse PPCPs interfere with MXR efflux activity of zebrafish indicates that (1) efflux transporters may influence bioaccumulation of many PPCPs in fish and that (2) many PPCPs may act as chemosensitisers. Furthermore, it appears that interference of PPCPs with efflux activity in zebrafish embryos is not only from effects on Abcb4 but also on other efflux transporter subtypes.

Influence of the Nd:YAG laser pulse duration on the temperature of primary enamel.

The aim of this study is to evaluate the temperature change on specimens of primary enamel irradiated with different pulse duration of Nd:YAG laser. Fifteen sound primary molars were sectioned mesiodistally, resulting in 30 specimens (3.5 × 3.5 × 2.0 mm). Two small holes were made on the dentin surface in which K-type thermocouples were installed to evaluate thermal changes. Specimens were randomly assigned in 3 groups (n = 10): A = EL (extra long pulse, 10.000 µs), B = LP (long pulse, 700 µs), and C = SP (short pulse, 350 µs). Nd:YAG laser (λ = 1.064 µm) was applied at contact mode (10 Hz, 0.8 W, 80 mJ) and energy density of 0.637 mJ/mm(2). Analysis of variance (ANOVA) was performed for the statistical analysis (P = 0.46). Nd:YAG laser pulse duration provided no difference on the temperature changes on primary enamel, in which the following means were observed: A = EL (23.15°C ± 7.75), B = LP (27.33°C ± 11.32), and C = SP (26.91°C ± 12.85). It can be concluded that the duration of the laser pulse Nd:YAG increased the temperature of the primary enamel but was not influenced by different pulse durations used in the irradiation.

Metabolic syndrome prevalence is increased in rheumatoid arthritis patients and is associated with disease activity.

OBJECTIVES: To evaluate the prevalence of metabolic syndrome (MetS) in patients with rheumatoid arthritis (RA) vs. controls, and to verify possible associations of MetS with specific disease-related factors. METHODS: The subjects were 283 RA patients and 226 healthy controls, frequency matched by age and sex. MetS was defined according to National Cholesterol Education Program (NCEP) criteria. Disease activity was evaluated with the Disease Activity Score using 28 joints (DAS28). A standardized clinical evaluation was performed and cardiovascular risk factors were assessed. RESULTS: The criteria for MetS were met by 39.2% RA patients vs. 19.5% in the control group (p < 0.001). Increased waist circumference, elevated blood pressure (BP), and fasting glucose were more frequent in RA patients than controls (p < 0.001 for all associations). By multiple logistic regression analysis (adjusted for age, sex, and years at school), the risk of having MetS was significantly higher for RA patients than for controls [odds ratio (OR) 1.87, 95% confidence interval (CI) 1.17-3.00, p = 0.009]. The DAS28 was significantly higher in RA patients with MetS than in those without MetS (3.59 ± 1.27 vs. 3.14 ± 1.53; p = 0.01). Disease duration, the presence of rheumatoid factor, and extra-articular manifestations were similar for patients with and without MetS. CONCLUSIONS: MetS frequency was higher in RA patients than in controls. Among RA patients, MetS was associated with disease activity. The higher prevalence of cardiovascular risk factors in RA suggests that inflammatory processes play a notable role in the development of cardiovascular disease (CVD), and indicates that tight control of systemic inflammatory activity and CVD modifiable risk factors should be recommended.

Different patterns of dermatological presentations in patients co-infected with human immunodeficiency virus and hepatitis C virus (HCV), and those infected with HCV alone.

BACKGROUND: Hepatitis C virus (HCV) and human immunodeficiency virus (HIV) share the same transmission routes. About 30% of HIV-positive patients are co-infected with HCV. Of the various HCV-related extrahepatic events, those involving the skin may be the first sign of infection. AIM: To specify the skin presentations in patients co-infected with HIV and HCV (co-infected patients; CP) and compare them with those found in patients with HCV mono-infection (mono-infected patients; MP). METHODS: This was a cross-sectional study, in which the studied population consisted of MP and CP from a tertiary hospital in the South of Brazil, who underwent complete skin examination and laboratory tests. RESULTS: In total, 201 patients were assessed, of whom 108 were CP, and 93 were MP. Pruritus tended to be more common in MP. MP also had significantly more dermatological conditions (mean of 5.2) than CP (mean of 4.5). In total, 104 different skin diseases were identified. There was a higher prevalence of infectious diseases and pigmentation disorders, such as verruca vulgaris and facial melasma, in CP, whereas trunk and facial telangiectasias, palmar erythema, and varicose veins were more common in MP. CONCLUSION: We found a high prevalence of skin conditions both in MP and in CP; however, the patterns of the dermatological conditions were different. CP were found to have significantly fewer skin lesions than MP, but had a higher prevalence of infectious and pigmentation disorders. By contrast, vascular conditions were more common in MP.

Assessment of aerobic capacity during swimming exercise in ob/ob mice.

Obesity is a highly prevalent condition associated with several diseases. Physical exercise has been considered as a non-pharmacological tool in the treatment of obesity. However, several aspects underlying exercise evaluation and prescription in obesity and associated pathologies are still under investigation. Although many research involving exercise have been performed in animal models, there is a lack of protocols for aerobic capacity assessment in obese animals, such as the ob/ob mice. This study aimed the following: (i) to verify the possibility of determining the lactate threshold (LT) on swimming exercise in ob/ob mice and in non-obese heterozygote mice (ob/OB), through visual inspection (vLT) and polynomial adjustment (pLT); and (ii) to verify if the LT determined through these protocols corresponds to the maximal lactate steady state (MLSS). Eight ob/ob and ten ob/OB mice performed an incremental exercise test to determine vLT and pLT as well as constant-load exercise bouts to determine MLSS. There were no within-group differences between vLT, pLT and MLSS [ob/ob: ~5.3% body weight (BW); ob/OB: ~3·6%BW] with a high agreement among protocols. In conclusion, the identification of the LT and MLSS intensities was possible for both groups. These data suggest that the proposed protocols may be used in new research on the effects of different exercise intensities on some aspects of obesity.

Primary malignant melanoma of the uterine cervix--case report and review.

INTRODUCTION: Malignant melanoma (MM) represents 1% of all cancers and has an incidence of 3-7% in the female genital tract, the majority of cases being reported in the vulva and vagina. CASE REPORT: A 75-year-old white female had a history of vaginal bleeding due to a 1.5 cm exophytic and ulcerated cervical lesion. Incisional biopsy was taken and sent for histopathological examination, which revealed MM of the cervix, confirmed by immunohistochemistry. Exclusion of the primitive tumor in other sites was made and after FIGO staging (IB1) the patient underwent a radical hysterectomy with bilateral salpingo-oophorectomy and pelvic lymphadenectomy. Three months later, hepatic and osseous metastases were detected, and the patient underwent chemotherapy and palliative radiotherapy with no success. DISCUSSION: Primary MM of the cervix should be considered in the differential diagnosis of cervical malignancies. Early diagnosis is essential in order to warrant a better prognosis, although there are no cases of cure described.

Expectations and final evaluation of complete dentures by patients, dentist and dental technician.

There is a poor association between the dentist's evaluation of denture quality and patients' satisfaction with their dentures. Possible differences between dental professionals and patients' expectations might help explain differences in outcome evaluations. This study compared scores given by a dentist, a dental technician and patients for their expectations before and their final evaluation after complete dentures treatment. Twenty completely edentulous patients, a prosthodontist and a dental technician provided scores for the expected aesthetic and functional results of their dentures based on a visual analogue scale at baseline. Post-treatment completion ratings were given after adjustments, by dentist and patients. The dental technician provided post-treatment completion ratings after completing the dentures. The patients had higher expectations than the dental technician and the dentist perceived for both aesthetic and function (P < 0.001). The patients also presented higher post-treatment completion ratings than the dental professionals perceived for final aesthetics (P = 0.016, Kendall's W = 0.207) and function (P = 0.002, Kendall's W = 0.303). Only the dentist presented a statistically significant difference between expectations (lower) and final (higher) outcomes for aesthetics (P = 0.017) and function (P = 0.003). There was no correlation between expectations and post-treatment completion ratings according to the patients' age. There was also no correlation between the patients' gender and expectation scores. Patients presented higher expectations regarding their dentures than dental professionals. The dentist believed that dentures would bring fewer benefits than patients did, but his perception of denture benefits post-treatment was significantly higher than his expectations.

Norfluoxetine and venlafaxine in zebrafish larvae: Single and combined toxicity of two pharmaceutical products relevant for risk assessment.

Antidepressant metabolites are found in natural and waste waters. However, investigation of their toxic effects on aquatic animals, single or in mixture with other occurring psychoactive drugs, has been neglected. Here, effects of 80hpf exposure to norfluoxetine (0.64-400 ng/L), venlafaxine (16-10000 ng/L) or their combination (3.2 ng/L +2000 ng/L, respectively) were investigated in embryos and zebrafish larvae. Mortality, embryonic malformations, sensorymotor reflexes and the expression of 34 genes involved in the toxicants mode-of-action (MoA) and metabolism were evaluated (i.e. monoamine receptors and transporters, nuclear receptors, and detoxification transporters and enzymes). Compared to controls, norfluoxetine treatments only caused depigmentation of embryos and larvae. Venlafaxine-exposed larvae exhibited depigmentation and spinal deformities, impaired sensorymotor reflexes, alterations in the expression of genes belonging to the serotonergic, noradrenergic and dopaminergic pathways, as well as nuclear receptors related to lipid and drug metabolism. The mixture elicited distinct interaction effects, depending on the level of biological organisation analysed and the neurotransmitter pathways affected; synergism (lethality), no interaction (sensorymotor reflexes), antagonism and inverse agonism (gene expression). The results call for investigation of the toxicity of pharmaceutical metabolites single and in mixture, as well as their risk assessment in approaches accounting for possible interactions with other endocrine-disrupting compounds.

Patients' expectations and satisfaction of complete denture therapy and correlation with locus of control.

Considering the importance of patients' satisfaction with complete denture therapy and the influence of psychosocial aspects and expectations regarding satisfaction with their dentures, the present study aimed to test for a correlation between locus of control profiles and expectations before and satisfaction after complete denture therapy. Sixty-four patients rated their expectations for aesthetic and functional results of complete denture therapy on a 10-cm visual analogue scale (VAS), using scores from 0 (worst results) to 10 (best results). A similar questionnaire was used regarding satisfaction after final denture adjustments. Patients also answered a locus of control questionnaire in the initial interview. ANOVA and a chi-squared test were used to test for correlations among ratings, patient gender and locus of control profiles. The significance level adopted was 0.05. The expectation ratings before treatment were significantly lower than the post-treatment completion ratings, both for aesthetics (P < 0.001) and function (P = 0.004). There was no correlation between locus of control profiles and patients' ratings or between gender and VAS scores. No correlation was found between locus of control profiles and scores for expectations before and satisfaction after complete dentures.

Fluoxetine modulates the transcription of genes involved in serotonin, dopamine and adrenergic signalling in zebrafish embryos.

Neurotransmitters pathways in fish and mammals are phylogenetically conserved. Therefore, the environmental presence of psychopharmaceuticals, such as fluoxetine (FLU), are likely to interact with fish serotonergic, dopaminergic and adrenergic systems, affecting their response and associated biological functions. Hence, the present work aimed at evaluating the effects of FLU in the transcription of genes involved in serotonin, dopamine and adrenergic transporters and receptors signalling in early stages of Danio rerio development. Embryos (1 hpf) were exposed for 80 h to different concentrations of FLU (0.0015, 0.05, 0.1, 0.5 and 0.8 µM) and mRNA levels of sert, 5-ht1a, 5-ht2c, dat, drd1b, drd2b, net, adra2a, adra2b, adra2c, vmat and mao were evaluated. A sensorimotor reflex assay was also performed demonstrating a significant decrease in tail reflex at 0.1 and 0.5 µM. The transcription levels of serotonergic and dopaminergic transporters (sert and dat) and vmat were down-regulated at environmentally relevant concentration (0.0015 µM). Receptors 5-ht2c, drd2b adra2b and adra2c mRNA levels also displayed a down regulation pattern after FLU exposure. In conclusion, this study demonstrated the interaction of FLU with the neurotransmission system at environmentally relevant concentrations by changing transcription patterns. Therefore, given the importance of these signalling pathways it is possible that their disruption can ultimately disturb the escape behaviour and biological functions in fish. Hence, evaluating the presence of this psychopharmaceutical in the aquatic environment should be implemented in future monitoring programmes.

Danio rerio embryos on Prozac - Effects on the detoxification mechanism and embryo development.

In the past decade the presence of psychopharmaceuticals, including fluoxetine (FLU), in the aquatic environment has been associated with the increasing trend in human consumption of these substances. Aquatic organisms are usually exposed to chronic low doses and, therefore, risk assessments should evaluate the effects of these compounds in non-target organisms. Teleost fish possess an array of active defence mechanisms to cope with the deleterious effects of xenobiotics. These include ABC transporters, phase I and II of cellular detoxification and oxidative stress enzymes. Hence, the present study aimed at characterising the effect of FLU on embryo development of the model teleost zebrafish (Danio rerio) concomitantly with changes in the detoxification mechanisms during early developmental phases. Embryos were exposed to different concentrations of FLU (0.0015, 0.05, 0.1, 0.5 and 0.8µM) for 80hours post fertilization. Development was screened and the impact in the transcription of key genes, i.e., abcb4, abcc1, abcc2, abcg2, cyp1a, cyp3a65, gst, sod, cat, ahr, pxr, pparα, pparß, pparγ, rxraa, rxrab, rxrbb, rxrga, rxrgb, raraa, rarab, rarga evaluated. In addition, accumulation assays were performed to measure the activity of ABC proteins and antioxidant enzymes (CAT and Cu/ZnSOD) after exposure to FLU. Embryo development was disrupted at the lowest FLU concentration tested (0.0015µM), which is in the range of concentrations found in WWTP effluents. Embryos exposed to higher concentrations of FLU decreased Cu/Zn SOD, and increased CAT (0.0015 and 0.5µM) enzymatic activity. Exposure to higher concentrations of FLU decreased the expression of most genes belonging to the detoxification system and upregulated cat at 0.0015µM of FLU. Most of the tested concentrations downregulated pparα, pparß, pparγ, and raraa, rxraa, rxrab, rxrbb rxrgb and ahr gene expression while pxr was significantly up regulated at all tested concentrations. In conclusion, this study shows that FLU can impact zebrafish embryo development, at concentrations found in effluents of WWTPs, concomitantly with changes in antioxidant enzymes, and the transcription of key genes involved in detoxification and development. These finding raises additional concerns supporting the need to monitor the presence of this compound in aquatic reservoirs.

Simvastatin effects on detoxification mechanisms in Danio rerio embryos.

The transcription and protein activity of defence mechanisms such as ABC transporters, phase I and II of cellular detoxification and antioxidant enzymes can be altered in the presence of emerging contaminants such as pharmaceuticals impacting the overall detoxification mechanism. The present work aimed to characterise the effects of simvastatin on the detoxification mechanisms of embryonic stages of Danio rerio. In a first approach, constitutive transcription of key genes involved in detoxification was determined. Embryos were collected at different developmental stages, and transcription patterns of genes coding for ABC transporters, phase I and II and oxidative stress were analysed. With exception of abcc2, all genes seem to be from maternal transfer (0-2 hpf). Embryos were then exposed to different concentrations of simvastatin (5 and 50 µg/L), verapamil and MK571 (10 µM; ABC protein inhibitors) and a combination of simvastatin and ABC inhibitors. mRNA expression levels of abcb4, abcc1, abcc2, abcg2, cyp1a, cyp3a65, gst, sod, cat was evaluated. Accumulation assays to measure ABC proteins activity and activity of EROD, GST, CAT and Cu/ZnSOD, were also undertaken. Simvastatin acted as a weak inhibitor of ABC proteins and increased EROD and GST activity, whereas Cu/ZnSOD and CAT activity were decreased. Simvastatin up-regulated abcb4 and cyp3a65 transcription (both concentrations), as well as abcc1 and abcc2 at 50 µg/L, and down-regulated gst, sod, cat at 5 µg/L. In conclusion, our data revealed the interaction of simvastatin with detoxification mechanisms highlighting the importance of monitoring the presence of this emerging contaminant in aquatic environments.

Grass pollen sensitization in dogs in Paraná, Brazil: comparison between percutaneous and intradermal testing / [Sensibilização a polens em cães no Paraná, Brasil: comparação entre o teste percutâneo e o teste intradérmico]

Canine atopic dermatitis is an inflammatory, genetic, pruritic and chronic dermatosis that affects between 10 and 30% of dogs and one of the most important allergens is grass pollen. The objective of this study was to evaluate the sensitization to grass pollen allergens in dogs with canine atopic dermatitis and to compare intradermal skin test (IDT) with percutaneous test (PT). For this study, ten healthy dogs and 39 dogs with atopic dermatitis were tested. Dogs were submitted to IDT and PT for Lolium multiflorum, Cynodon dactylon and Paspalum notatum. The IDT and PT tests were compared using the Proportion Test. All healthy dogs were negative to both tests. Ten atopic dogs (25.6%) responded positively to the PT and none were positive in IDT. C. dactylon, L. multiflorum and P. notatum were responsible for positive reactions in 70%, 70% and 30% of positive dogs, respectively. The number of positive reactions in PT were statistically higher than IDT (P<0.05). In conclusion, grass pollen can be important source of allergens for dogs in Paraná state (Brazil) and the PT showed higher sensitization to grass pollen in dogs with atopic dermatitis than IDT.(AU)

A dermatite atópica canina é uma dermatose inflamatória, genética, prurítica e crônica que afeta entre 10% e 30% dos cães, e um dos alérgenos mais importantes são os polens de gramíneas. O objetivo deste estudo é avaliar a sensibilização a alérgenos de polens de gramíneas em cães com dermatite atópica e comparar o teste intradérmico (TID) com o teste percutâneo (TP). Para o estudo, 10 cães hígidos e 39 cães com dermatite atópica foram testados. Estes foram submetidos ao TID e ao TP para Lolium multiflorum, Cynodon dactylon e Paspalum notatum. TID e TP foram comparados usando-se o teste de proporção. Todos os cães hígidos foram negativos em ambos os testes. Dez cães atópicos (25,6%) responderam positivamente ao TP e nenhum ao TID. C. dactylon, L. multiflorum e P. notatum foram responsáveis por reações positivas de 70%, 70% e 30% dos cães positivos, respectivamente. O número de reações positivas no TP foi estatisticamente maior que no TID (P<0,05). Foi concluído que os polens de gramíneas podem ser importantes fontes de alérgenos para cães no estado do Paraná (Brasil) e que o TP mostrou maior sensibilização a polens em cães com dermatite atópica que o TID.(AU)

A Ca2+-stimulated, Mg2+-dependent ATPase activity in subcellular fractions from Schistosoma mansoni.

A Ca2+-stimulated, Mg2+-dependent ATPase activity was found in subcellular fractions from Schistosoma mansoni. Its specific and relative activities were higher in the heterogeneous cuticle fraction and in the microsomal fraction. The K0.5 for ATPase activation by free Ca2+ was 0.2-0.5 microM. This is the first description of an ATPase activity stimulated by Ca2+ in the micromolar range in S. mansoni.

A (Ca(2+)-Mg2+)ATPase from Schistosoma mansoni is coupled to an active transport of calcium.

The (Ca(2+)-Mg2+)ATPase activity in microsomes of Schistosoma mansoni is fully inhibited by vanadate (I50 = 2.5 microM). 45Ca2+ is accumulated within microsomal vesicles in an ATP-dependent process that is enhanced 5-fold in the presence of 40 mM phosphate. Accumulated 45Ca2+ is rapidly released by 5 microM of the Ca2+ ionophore A23187 (t1/2 less than or equal to 6 s). (Ca(2+)-Mg2+)ATPase activity and Ca2+ uptake share the same subcellular distribution pattern and similar Ca2+ sensitivities (K0.5 = 0.39 microM and 0.15 microM, respectively). The substrate selectivity is high for both ATPase activity and Ca2+ transport. These results indicate the presence of an active transport of Ca2+ coupled to the (Ca(2+)-Mg2+)ATPase activity previously described in this parasite. A plasma membrane localization and physiological role in calcium homeostasis are suggested.

Evidence for ryanodine receptors in Schistosoma mansoni.

The present study investigated the presence of ryanodine receptors in the trematode Schistosoma mansoni. [3H]Ryanodine specific binding sites were found in the four subcellular fractions of S. mansoni; however, more binding sites were recovered in the heterogeneous fraction P1 and the microsomal fraction P4, as was thapsigargin-sensitive (Ca2+-Mg2+)ATPase activity, marking the sarco/endoplasmic reticulum calcium ATPase (SERCA) pumps. This binding had an equilibrium dissociation constant (Kd) in the nanomolar range, an apparent maximal number of receptors (Bmax) of about 80 fmol/mg of protein, and was modulated by ions (Ca2+, Mg2+) and some pharmacological tools such as caffeine. Ryanodine was able to accelerate the rate of 45Ca2+ release from actively loaded vesicles, and also to induce a transient contraction of the whole worm. We conclude that ryanodine-sensitive Ca2+ release channels are present in S. mansoni, with properties very similar to the ones present in higher animals.

Praziquantel has no direct effect on (Na(+)+K+)-ATPases and (Ca2(+)-Mg2+)ATPases of Schistosoma mansoni.

Therapeutic concentrations of praziquantel produce a rapid and intense contraction of the human flatworm Schistosoma mansoni. As an action on ATPases responsible for calcium homeostasis arises as a possible explanation for the molecular mechanism of this effect, we tested here the effect of praziquantel on different preparations from male adult worms that were previously characterized for their content in (Na(+)+K+)-ATPase and (Ca2(+)-Mg2+)ATPase activities from different origins. Concentrations as high as 100 microM praziquantel did not inhibit (Na(+)+K+)-ATPase from tegument and carcass nor (Ca2(+)-Mg2+)ATPase from heterogeneous (P1) and microsomal (P4) fractions. As 100 microM praziquantel was also without effect on calcium permeability of microsomal vesicles actively loaded with 45Ca2+, the present results discard three hypotheses recently raised for the mechanism of praziquantel-induced contraction of S. mansoni.