Effect of peritoneal fluid from patients with minimal/mild endometriosis on progesterone release by human granulosa-lutein cells obtained from infertile patients without endometriosis: A pilot study

Objective: To evaluate the effect of peritoneal fluid (PF) from women without and with minimal/mild endometriosis on progesterone (P) release by cultured human granulosa-lutein cells obtained from infertile patients without endometriosis submitted to ovarian hyperstimulation for in vitro fertilization (IVF). Study design: A pilot study was performed. Human granulosa-lutein cells, obtained from 11 infertile patients without endometriosis (tubal or male factors of infertility) submitted to ovarian hyperstimulation for IVF, were cultured without PF (basal production) and with increasing volumes of steroid-extracted PF samples from 11 patients with endometriosis and 11 patients without endometriosis. Progesterone (P) levels in the media after 72 h culture were measured by chemoluminescence assay. The non-parametric Mann-Whitney-test was used for statistical analysis. Results: PF from patients without endometriosis stimulated P release in a dose-dependent manner up to the dose of 100 mu l/ml (10% concentration) when compared with basal production (without adding PF). P release was similar in cultures stimulated with PF from patients with or without endometriosis at 1% (10 mu l/ml) and 5% (50 ml/ml) concentrations. At 10% concentration, there was a non-statistically significant reduction in progesterone release by granulosa cells stimulated with PF from patients with endometriosis. PF from patients with endometriosis significantly reduced P release at 30% concentration (300 mu l/ml). Conclusions: PF stimulates P release by human granulosa-lutein cells in a dose-dependent manner. However, higher concentrations of PF from patients with minimal/mild endometriosis reduce P release, suggesting it contains factors that may compromise ovarian steroidogenesis. (c) 2007 Elsevier Ireland Ltd. All rights reserved.

Addressing overtreatment in patients with refractory epilepsy at a tertiary referral centre in Brazil

Background. Patients with refractory epilepsy often have impaired quality of life (QOL) as a consequence of seizures and adverse effects of antiepileptic drugs. We assessed the impact of adverse effects on QOL and the utility of a structured instrument to help the physician manage adverse effects in patients with refractory epilepsy. Methods. Clinical characteristics, drug treatment and adverse effects were evaluated in 102 patients with refractory epilepsy at a single tertiary referral centre. The Adverse Events Profile (AEP) and Quality of Life in Epilepsy-31 (QOLIE-31) questionnaires were completed at baseline and after six months. At baseline, patients with a high burden of adverse effects (AEP scores >= 45) were randomized to an intervention or control group. AEP scores in the intervention group were available to the physician as an instrument to help to reduce adverse effects. Results. Ninety-five patients (93.1%) were on polytherapy. Sixty-six completed the questionnaires and, of these, 43 (65.1%) had a high AE burden and were randomized to the intervention and control group. QOLIE-31 scores were inversely correlated with AEP scores at both visits. Among randomized patients, AEP scores tended to decrease between the baseline and the final visit without significant differences between groups (intervention group: 54.1 +/- 6.1 vs 51.1 +/- 9.1; control group: 55.8 +/- 5.8 vs 50.5 +/- 12.2). QOLIE-31 scores did not change substantially between visits (intervention group: 45.9 +/- 17.4 vs 48.4 +/- 14; control group: 47.5 +/- 15.7 vs 45.2 +/- 18.9). Conclusion. A significant proportion of patients had a high toxicity burden which had an impact on their QOL. Reduction of over-treatment is a difficult challenge which cannot be addressed solely by providing a structured assessment of adverse effects, but requires a more comprehensive approach aimed at optimizing the many components of the management strategy.

5-HT2C receptor regulation of defensive responses in the rat dorsal periaqueductal gray

Activation of 5-HT2C receptors in limbic structures such as the amygdala and hippocampus increases anxiety. Indirect evidence obtained with non-selective 5-HT2C-interacting drugs suggests that the same may occur in the dPAG, a brainstem region consistently implicated in the genesis/regulation of panic attacks. In this study we used more selective agonists and antagonists to unveil the role played by dPAG 5-HT2C receptors in the regulation of anxiety- and panic-related defensive behaviors. Our results showed that intra-dPAG microinjection of the endogenous agonist 5-HT (20 nmol) or the 5-HT2C receptor agonists MK-212 (1 and 10 nmol) and RO-600175 (40 nmol) significantly increased inhibitory avoidance acquisition in rats tested in the elevated T-maze, suggesting an anxiogenic effect. 5-HT, but not the two 5-HT2C receptor agonists, inhibited escape performance. In the elevated T-maze, inhibitory avoidance and escape responses have been related to generalized anxiety and panic attacks, respectively. The behavioral effects caused by 5-HT and MK-212 were fully blocked by previous local microinjection of the 5-HT2C receptor antagonist SB-242084. Intra-dPAG injection of MK-212 also failed to affect escape expression in another test relating this behavior to panic, the electrical stimulation of the dPAG. Overall, the results indicate that 5-HT2C receptors in the dPAG are preferentially involved in the regulation of defensive behaviors related to anxiety, but not panic. This finding extends to the dPAG the prominent role that has been attributed to 5-HT2C receptors in anxiety generation. (C) 2010 Elsevier Ltd. All rights reserved.

A DNA vaccine candidate encoding the structural prM/E proteins elicits a strong immune response and protects mice against dengue-4 virus infection

A DNA vaccine expressing dengue-4 virus premembrane (prM) and envelope (E) genes was produced by inserting these genes into a mammalian expression plasmid (pCI). Following a thorough screening, including confirmation of protein expression in vitro, a recombinant clone expressing these genes was selected and used to immunize BALB/c mice. After 3 immunizations all the animals produced detectable levels of neutralizing antibodies against dengue-4 virus. The cytokines levels and T cell proliferation, detected ex vivo from the spleen of the immunized mice, showed that our construction induced substantial immune stimulation after three doses. Even though the antibody levels, induced by our DNA vaccine, were lower than those obtained in mice immunized with dengue-4 virus the levels of protection were high with this vaccine. This observation is further supported by the fact that 80% of the vaccine immunized group was protected against lethal challenge. In conclusion, we developed a DNA vaccine employing the genes of the prM and E proteins from dengue-4 virus that protects mice against this virus. (C) 2010 Elsevier Ltd. All rights reserved.

Age-related deregulation of Aire and peripheral tissue antigen genes in the thymic stroma of non-obese diabetic (NOD) mice is associated with autoimmune type 1 diabetes mellitus (DM-1)

Gene expression of peripheral tissue antigens (PTAs) in stromal medullary thymic epithelial cells (mTECs) is a key process to the negative selection of autoreactive thymocytes. This phenomenon was termed ""promiscuous gene expression"" (PGE), which is partially controlled by the Aire gene. Nevertheless, reasons for the correlation of Aire and PTAs with the emergence of autoimmune diseases are largely unknown, though it may be a result of a chronological effect. Although the effect of Aire mutations in pathogenic autoimmunity is well know, it could not be a unique cause for autoimmunity. Independently of mutations, temporal deregulation of Aire expression may imbalance Aire-dependent PTAs and/or wide PGE. This deregulation may be an early warning sign for autoimmune diseases as it guarantees autoantigen representation in the thymus. To assess this hypothesis, we studied the expression levels of Aire, Aire-dependent (Ins2) and Aire-independent (Gad67 and Col2a1) PTAs using real-time-PCR of the thymic stromal cells of NOD mice during the development of autoimmune type 1 diabetes mellitus (DM-1). Wide PGE was studied by microarrays in which the PTA genes were identified through parallel CD80(+) mTEC 3.10 cell line expression profiling. The results show that Aire gene was down-regulated in young pre-autoimmune (pre-diabetic) NOD mice. PGE and specific PTA genes were down-regulated in adult autoimmune diabetic animals. These findings represent evidence indicating that chronological deregulation of genes important to negative selection may be associated with the development of an autoimmune disease (DM-1) in mice.

Differential expression of HDAC3, HDAC7 and HDAC9 is associated with prognosis and survival in childhood acute lymphoblastic leukaemia

Altered expression of histone deacetylases (HDACs) is a common feature in several human malignancies and may represent an interesting target for cancer treatment, including haematological malignancies. We evaluated the mRNA gene expression profile of 12 HDAC genes by quantitative real-time polymerase chain reaction in 94 consecutive childhood acute lymphoblastic leukaemia (ALL) samples and its association with clinical/biological features and survival. ALL samples showed higher expression levels of HDAC2, HDAC3, HDAC8, HDAC6 and HDAC7 when compared to normal bone marrow samples. HDAC1 and HDAC4 showed high expression in T-ALL and HDAC5 was highly expressed in B-lineage ALL. Higher than median expression levels of HDAC3 were associated with a significantly lower 5-year event-free survival (EFS) in the overall group of patients (P = 0.03) and in T-ALL patients (P = 0.01). HDAC7 and HADC9 expression levels higher than median were associated with a lower 5-year EFS in the overall group (P = 0.04 and P = 0.003, respectively) and in B-lineage CD10-positive patients (P = 0.009 and P = 0.005, respectively). Our data suggest that higher expression of HDAC7 and HDAC9 is associated with poor prognosis in childhood ALL and could be promising therapeutic targets for the treatment of refractory childhood ALL.

Identification and characterization of a new member of snake venom thrombin inhibitors from Bothrops insularis using a proteomic approach

Snake venom C-type lectin-like proteins (CLPs) are ubiquitously found in Viperidae snake venoms and differ from the C-type lectins as they display different biological activities but no carbohydrate-binding activity. Previous analysis of the transcriptome obtained from the Bothrops insularis venom gland showed the presence of two clusters homologous to bothrojaracin (BJC) chains a and P. In an effort to identify a new BJC-like molecule, we used an approach associated with proteomic technologies to identify the presence of the expressed protein and then to purify and characterize a new thrombin inhibitor from B. insularis venom. We also constructed homology models of this protein and BJC, which were compared with other C-type lectin-like family members and revealed several conserved features of this intriguing snake venom toxin family. (C)0 2007 Elsevier Ltd. All rights reserved.

The use of different doses of metamizol for post-operative analgesia in dogs

Objective To evaluate the post-operative analgesic effect of metamizol (dipyrone) administered intravenously at three different doses (15 mg kg(-1), 25 mg kg(-1) and 35 mg kg(-1)) compared to placebo in dogs undergoing ovariohysterectomy. Study design Prospective, comparative, randomized. blinded trial. Animals Forty healthy bitches, aged 1-6 years, weighing 10-35 kg Methods The animals were randomly divided into four groups and received their respective treatments immediately after surgery: placebo group (0.9% saline solution), D15 group (metamizol 15 mg kg(-1) IV), D25 group (metamizol 25 mg kg(-1) IV), D35 group (metamizol 35 mg kg(-1) IV). The following variables were measured: sedation, pulse rate (PR). respiratory rate (f(R)). arterial blood pressure (ABP), plasma catecholamines. serum cortisol, blood urea nitrogen (BUN) and creatinine metabolites. albumin, alanine aminotransferase (ALT), alkaline phosphatase (ALP). hemogram. platelet counts and level of analgesia which was assessed by visual analog (VAS). descriptive and behavioral scales. Patients were monitored for 48 hours after the administration of the analgesic agent. Rescue analgesia (tramadol, 2 mg kg(-1), intramuscularly) was provided for animals with pain scores >= 4, as determined by the VAS or descriptive scale. Results The D25 and D35 groups showed equivalent post-operative analgesia, as shown by decreased pain scores, according to the three different pain scales, and fewer animals that required rescue analgesia. Significantly lower serum cortisol concentrations were observed in the D25 and D35 groups when compared to the placebo and D15 groups. No hematologic, renal, hepatic or clinical adverse effects were observed during the treatment. Conclusions and clinical relevance Metamizol administered intravenously at 25 or 35 mg kg(-1) can provide adequate post-operative analgesia in bitches undergoing ovariohysterectomy.

Temporal evolution of epithelial, vascular and interstitial lung injury in an experimental model of idiopathic pulmonary fibrosis induced by butyl-hydroxytoluene

This study was undertaken to test whether the structural remodelling of pulmonary parenchyma can be sequentially altered in a model and method that demonstrate the progression of the disease and result in remodelling within the lungs that is typical of idiopathic pulmonary fibrosis. Three groups of mice were studied: (i) animals that received 3-5-di-tert-butyl-4-hydroxytoluene (BHT) and were killed after 2 weeks (early BHT = 9); (ii) animals that received BHT and were killed after 4 weeks (late BHT = 11); (iii) animals that received corn oil solution (control = 10). The mice were placed in a ventilated Plexiglas chamber with a mixture of pure humidified oxygen and compressed air. Lung histological sections underwent haematoxylin-eosin, immunohistochemistry (epithelial, endothelial and immune cells) and specific staining (collagen/elastic fibres) methods for morphometric analysis. When compared with the control group, early BHT and late BHT groups showed significant decrease of type II pneumocytes, lower vascular density in both and higher endothelial activity. CD4 was increased in late BHT compared with early and control groups, while CD8, macrophage and neutrophil cells were more prominent only in early BHT. The collagenous fibre density were significantly higher only in late BHT, whereas elastic fibre content in late BHT was lower than that in control group. We conclude that the BHT experimental model is pathologically very similar to human usual interstitial pneumonia. This feature is important in the identification of animal models of idiopathic pulmonary fibrosis that can accurately reflect the pathogenesis and progression of the human disease.

Effects of aeration on characteristics of sugarcane silage.

Effects of aeration on characteristics of sugarcane silage. This trial aimed at evaluating, the deleterious effects of aeration time on nutritive value and other fermentative characteristics of sugarcane silage. A completely randomized design was used with three treatments and four repetitions per treatment. Fresh chopped sugarcane was exposed to aeration for 0, 4 or 8 hours, and ensiled soon After exposure, the material was ensiled in 12 laboratory silos (plastic buckets). Silos were opened 85 after ensiling, when organic acids contents and chemical composition of silages were determined. Deviation of linearity (p < 0.05) was observed for aeration time on dry matter. A positive linear effect was observed (p < 0.05) on ADF, NDF and soluble carbohydrates content, but negative for ammoniacal nitrogen content and in vitro digestibility of dry matter. For organic acids content, deviation of linearity was observed on acetic acid, with the lowest content (1.5% of DM) observed after 8 hours of aeration, and a negative linear effect was observed for lactic and butyric acids, as well as for pH values. There were no effects on ethanol concentration, which remained very high (22% of DM), regardless of aeration time. Aerobic stability of silage worsened with the increase in aeration time.

Effects of the utilization of homeopathic elements in commercial diluent on swine sperm viability

It has been speculated that the homeopathic treatment of sperm cells in order to improve semen quality could be promising. However, few data is available and its use in spermatozoa requires investigation. It is well established that mitochondrial membrane potential is an important viability parameter of spermatozoa and it is intimately related to reproductive efficiency. In this manner, new technologies in order to improve the activity of sperm cells and, finally, the fecundity of swine herds are of extremely importance. Due to the lack of knowledge of homeopathic treatment effect on spermatozoa, the aim of the present study was to verify the effect of three different homeopathic treatments on viability of boar sperm cells. Three homeopathic treatments composed by Pulsatila CH6, Pulsatila and Avena CH6, Avena CH6 and one control treatment (sucrose) were added to diluted boar semen, which were cooled for 24 or 48 h. Interestingly, no positive effect of homeopathic treatments was observed over semen viability. However, it was demonstrated that the 24 h of cooling storage provided more viable sperm cells when compared to the 48-h period. This effect of storage period on sperm viability was assessed by intact plasmatic membrane, intact acrosome and mitochondrial membrane potential evaluation.

Detection of Rabies Virus Antibodies in Brazilian Free-Ranging Wild Carnivores

Rabies virus is a pathogen of major concern in free-ranging wild carnivores in several regions of the world, but little is known about its circulation in Brazilian wild carnivores. Sera from 211 free-ranging wild carnivores, captured from 2000 to 2006 in four locations of two Brazilian biomes (Pantanal and Cerrado), were tested for rabies antibodies. Twenty-six individuals (12.3%) had neutralizing antibody titers >= 0.10 IU/ml. The four sampled locations had antibody-positive animals, suggesting that Rabies virus circulates in all of these regions. Results underscore the risk posed by rabies for conservation of Brazilian carnivores and the possibility of the animals acting as reservoirs for the Rabies virus.

Long-Term Maternal Separation Differentially Alters Serum Corticosterone Levels and Blood Neutrophil Activity in A/J and C57BL/6 Mouse Offspring

Objectives: In this work, we searched for maternal separation effects on serum corticosterone levels and blood neutrophil activity in adult male A/J and C57BL/6 mouse offspring. Methods: 40 male A/J mice and 40 male C57BL/6 mice were divided within each strain into two groups. Mice in the maternal separation group were separated from their mothers (1 h/day) on postnatal days 0-13. Mice in the control group were left undisturbed. On postnatal day 45, blood was drawn from all mice and used to assess neutrophil activity by flow cytometry and serum corticosterone levels by radioimmunoassay. Results: The results showed that each mouse strain responded differently to maternal separation, but in both cases, serum corticosterone levels were affected. In both strains, adult mice that experienced maternal separation showed lower serum corticosterone levels than control mice. In relation to control mice kept together with their mothers, the levels of serum corticosterone were 72.7 and 36.36% lower in A/J and C57BL/6 mice submitted to maternal separation, respectively. The current findings showed that maternal separation increased neutrophil activity in mice after reaching adulthood. The observed effects, although in the same direction, differed between A/J and C57BL/6 mice. Maternal separation increased both the percentage and intensity of phagocytosis in C57BL/6 mice, but had no effects on A/J mice. Furthermore, maternal separation increased basal and propidium iodide-labeled Staphylococcus aureus-induced oxidative burst in A/J mice but did not affect oxidative burst in C57BL/6 mice. Finally, phorbol myristate acetate-induced oxidative burst increased in both strains. Conclusion: These results indicate that early maternal separation increases innate immunity, most likely by modifying hypothalamus-pituitary-adrenal axis activity. This suggests that maternal separation is a good model for stress which produces long-term neuroimmune changes whatever the animal species and strain used. Copyright (C) 2011 S. Karger AG, Basel

Heat stress impairs performance parameters, induces intestinal injury, and decreases macrophage activity in broiler chickens

Studies on environmental consequences of stress on animal production have grown substantially in the last few years for economic and animal welfare reasons. Physiological, hormonal, and immunological deficits as well as increases in animals` susceptibility to diseases have been reported after different stressors in broiler chickens. The aim of the current experiment is to describe the effects of 2 different heat stressors (31 +/- 1 and 36 +/- 1 degrees C/10 h per d) applied to broiler chickens from d 35 to 42 of life on the corticosterone serum levels, performance parameters, intestinal histology, and peritoneal macrophage activity, correlating and discussing the obtained data under a neuroimmune perspective. In our study, we demonstrated that heat stress (31 +/- 1 and 36 +/- 1 degrees C) increased the corticosterone serum levels and decreased BW gain and food intake. Only chickens submitted to 36 +/- 1 degrees C, however, presented a decrease in feed conversion and increased mortality. We also showed a decrease of bursa of Fabricius (31 +/- 1 and 36 +/- 1 degrees C), thymus (36 +/- 1 degrees C), and spleen (36 +/- 1 degrees C) relative weights and of macrophage basal (31 +/- 1 and 36 +/- 1 degrees C) and Staphylococcus aureus-induced oxidative burst (31 +/- 1 degrees C). Finally, mild multifocal acute enteritis characterized by an increased presence of lymphocytes and plasmocytes within the jejunum`s lamina propria was also observed. The stress-induced hypothalamic-pituitary-adrenal axis activation was taken as responsible for the negative effects observed on the chickens` performance and immune function and also the changes of the intestinal mucosa. The present obtained data corroborate with others in the field of neuroimmunomodulation and open new avenues for the improvement of broiler chicken welfare and production performance.