Type I interferon receptors and interferon-τ-stimulated genes in peripheral blood mononuclear cells and polymorphonuclear leucocytes during early pregnancy in beef heifers.

This study characterised the expression of interferon (IFN)-τ-stimulated genes (ISGs) and Type I IFN receptors in circulating polymorphonuclear cells (PMNs) of beef heifers and compared it with expression in peripheral blood mononuclear cells (PBMCs) up to Day 20 of gestation. Nelore heifers (n=26) were subjected to fixed-time AI (FTAI) on Day 0. PMNs and PBMCs were isolated on Days 0, 10, 14, 16, 18 and 20 after FTAI. The abundance of target transcripts (ubiquitin-like protein (ISG15), 2'-5'-oligoadenylate synthetase 1 (OAS1), myxovirus resistance 1 (MX1), myxovirus resistance 2 (MX2), IFN receptor I (IFNAR1) and IFN receptor 2 (IFNAR2)) was determined using real-time quantitative polymerase chain reaction and compared between pregnant (n=8) and non-pregnant (n=9) females. In both PBMCs and PMNs, ISG15 and OAS1 expression was greater in pregnant than non-pregnant heifers on Days 18 and 20. There were no significant differences in the expression of ISGs between PBMCs and PMNs. A time effect on expression was found for IFNAR1 in PBMCs and IFNAR2 in PMNs, with decreased expression of both genes on Days 18 and 20. When the expression of these genes was compared between cell types only in pregnant heifers, IFNAR2 expression in PMNs had an earlier decrease when compared to its expression in PBMCs, starting from Day 18. In conclusion, PMNs do not respond earlier to the conceptus stimulus, and ISG15 and OAS1 expression in both PMNs and PBMCs can be used as a suitable marker for pregnancy diagnosis on Days 18 and 20. In addition, gestational status did not affect IFNAR1 and IFNAR2 expression, but IFNAR2 showed a distinct response between PMNs and PBMCs of pregnant heifers.

Role of extracellular vesicles during oocyte maturation and early embryo development.

The follicle is a dynamic microenvironment in the ovary where the oocyte develops. Intercellular communication between somatic cells and the oocyte inside the follicle is essential to generate a competent gamete. Extracellular vesicles are nanoparticles secreted by cells that mediate cell-to-cell communication in the follicle microenvironment and can be obtained from the follicular fluid. These extracellular vesicles have been studied as biomarkers and supplementation tools to mimic physiological conditions during assisted reproductive techniques because they are vehicles of bioactive molecules. Therefore, this paper reviews the importance of changes in the ovarian follicle and the effects of extracellular vesicles from follicular fluid during oocyte maturation and early embryo development. Finally, we propose that is important to consider the source of the extracellular vesicles to improve diagnostic methods and to increase invitro embryo production.

Deciphering the oviductal extracellular vesicles content across the estrous cycle: implications for the gametes-oviduct interactions and the environment of the potential embryo.

BACKGROUND: The success of early reproductive events depends on an appropriate communication between gametes/embryos and the oviduct. Extracellular vesicles (EVs) contained in oviductal secretions have been suggested as new players in mediating this crucial cross-talk by transferring their cargo (proteins, mRNA and small ncRNA) from cell to cell. However, little is known about the oviductal EVs (oEVS) composition and their implications in the reproductive success. The aim of the study was to determine the oEVs content at protein, mRNA and small RNA level and to examine whether the oEVs content is under the hormonal influence of the estrous cycle. RESULTS: We identified the presence of oEVs, exosomes and microvesicles, in the bovine oviductal fluid at different stages of the estrous cycle (postovulatory-stage, early luteal phase, late luteal phase and pre-ovulatory stage) and demonstrated that their composition is under hormonal regulation. RNA-sequencing identified 903 differentially expressed transcripts (FDR < 0.001) in oEVs across the estrous cycle. Moreover, small RNA-Seq identified the presence of different types of ncRNAs (miRNAs, rRNA fragments, tRNA fragments, snRNA, snoRNA, and other ncRNAs), which were partially also under hormonal influence. Major differences were found between post-ovulatory and the rest of the stages analyzed for mRNAs. Interesting miRNAs identified in oEVs and showing differential abundance among stages, miR-34c and miR-449a, have been associated with defective cilia in the oviduct and infertility. Furthermore, functional annotation of the differentially abundant mRNAs identified functions related to exosome/vesicles, cilia expression, embryo development and many transcripts encoding ribosomal proteins. Moreover, the analysis of oEVs protein content also revealed changes across the estrous cycle. Mass spectrometry identified 336 clusters of proteins in oEVs, of which 170 were differentially abundant across the estrous cycle (p-value< 0.05, ratio < 0.5 or ratio > 2). Our data revealed proteins related to early embryo development and gamete-oviduct interactions as well as numerous ribosomal proteins. CONCLUSIONS: Our study provides with the first molecular signature of oEVs across the bovine estrous cycle, revealing marked differences between post- and pre-ovulatory stages. Our findings contribute to a better understanding of the potential role of oEVs as modulators of gamete/embryo-maternal interactions and their implications for the reproductive success.

Subclinical mastitis interferes with ovulation, oocyte and granulosa cell quality in dairy cows.

The objective was to evaluate the effect of mastitis by somatic cell count (SCC) on follicular growth, ovulation, oocytes and cumulus cells quality and on the concentration and size of exosomes in follicular fluid of dairy cows. In the study, crossbred cows (Bos taurus - Holstein x Bos indicus - Gir) were classified for analysis as Control (SCC<200.000 cells/mL) and Mastitis (SCC>400.000 cells/mL) groups. In experiment 1 (follicular dynamics), cows (n = 57) were submitted to ultrasound evaluations every 24 h, from progesterone-releasing-intravaginal-device (PRID) removal (D8) until 48 h later (D10). Thereafter, evaluations were performed every 12 h, until ovulation or up to 96 h after PRID removal. In experiment 2 (oocyte, cumulus complexes, and follicular fluid evaluation), cows (n = 26) were submitted to follicular aspiration (OPU) for oocyte quality and cumulus cells transcript evaluation. The amount of cumulus complexes transcripts (BCL2, BAX, PI3K, PTEN, FOXO3) was determined by Real-Time Polymerase Chain Reaction. Moreover, seven days after the OPU session, the dominant follicle was aspirated. Exosomes were isolated from the follicular fluid for evaluation of particle size and concentration. Ovulation rate [Control 77.4% (24/31) and Mastitis 57.7% (15/26); P = 0.09] and viable oocytes rate [Control 59.1% (130/220) and Mastitis 41.9% (125/298); P = 0.01] were higher in Control animals. Additionally, there was a greater number of degenerate oocytes [Control 6.7 ±â€¯1.2 and Mastitis 13.3 ±â€¯5.5; (P = 0.001)] in subclinical mastitis cows. There was greater abundance (P = 0.003) of BAX cumulus cell transcripts and exosome mean (P = 0.03) and mode (P = 0.02) was smaller in subclinical mastitis cows. In conclusion, ovulation rate, oocyte quality, and exosome diameter were smaller in cows with SCC>400.000 cells/mL.

Low levels of exosomal-miRNAs in maternal blood are associated with early pregnancy loss in cloned cattle.

Nuclear reprogramming mediated by somatic cell nuclear transfer (SCNT) has many applications in medicine. However, animal clones show increased rates of abortion and reduced neonatal viability. Herein, we used exosomal-miRNA profiles as a non-invasive biomarker to identify pathological pregnancies. MiRNAs play important roles in cellular proliferation and differentiation during early mammalian development. Thus, the aim of this study was to identify exosomal-miRNAs in maternal blood at 21 days of gestation that could be used for diagnosis and prognosis during early clone pregnancies in cattle. Out of 40 bovine-specific miRNAs, 27 (67.5%) were with low abundance in the C-EPL (Clone - Early pregnancy loss) group compared with the C-LTP (Clone - Late pregnancy) and AI-LTP (Artificial Insemination - Late pregnancy) groups, which had similar miRNAs levels. Bioinformatics analysis of the predicted target genes demonstrated signaling pathways and functional annotation clusters associated with critical biological processes including cell proliferation, differentiation, apoptosis, angiogenesis and embryonic development. In conclusion, our results demonstrate decreased exosomal-miRNAs in maternal blood at 21 days of gestation in cloned cattle pregnancies that failed to reach term. Furthermore, the predicted target genes regulated by these 27 miRNAs are strongly associated with pregnancy establishment and in utero embryonic development.

Cellular and extracellular vesicular origins of miRNAs within the bovine ovarian follicle.

The ovarian follicle components must provide an ideal environment to ensure the success of reproductive processes, and communication between follicular cells is crucial to support proper oocyte growth. Recently, it has been demonstrated that the presence of extracellular vesicles (EVs) carrying microRNAs (miRNAs) in follicular fluid represents an important autocrine and paracrine communication mechanism inside the ovarian follicle. In this study, we tested the hypothesis that the miRNA content of EVs isolated from ovarian follicular (granulosa and cumulus-oocyte complexes) cell-conditioned culture media is dependent upon cell type. We initially screened bovine granulosa cells (GCs) and cumulus-oocyte complexes (COCs), as well as their derived EVs for 348 miRNAs using real-time PCR, and detected 326 miRNAs in GCs and COCs cells and 62 miRNAs in GCs and COCs EVs. A bioinformatics analysis of the identified cell-specific and differentially expressed miRNAs predicted that they likely modulate important cellular processes, including signalling pathways such as the PI3K-Akt, MAPK and Wnt pathways. By investigating the origins of miRNAs within the follicular fluid, the results of this study provide novel insights into follicular miRNA content and intercellular communication that may be of invaluable use in the context of reproductive technologies, diagnostic of ovarian-related diseases and/or the identification of biomarkers for oocyte and embryo quality.

Omeprazole and ranitidine in long-term treatment of duodenal ulcer: a double-blind comparison of length of time in remission.

In most patients duodenal ulcer is a chronic relapsing disease. If no active maintenance treatment or eradication therapy is given after healing, around 70-100% of patients have a relapse during the first year. We conducted a double-blind multicenter study in 472 patients with duodenal ulcer. They were treated with omeprazole 20 mg every morning for four or eight weeks and when healed were randomly allocated to maintenance treatment with either omeprazole 20 mg every morning or ranitidine 150 mg at bedtime for up to six months. The patients were assessed by endoscopy at monthly intervals until healing occurred. Thereafter scheduled endoscopy was carried out after 1, 3, and 6 months of maintenance treatment or immediately in the event of a suspected relapse. Healing status (intention to treat approach) was 87% at four weeks and 93% at eight weeks. At six months the estimated remission rate was 90% for omeprazole and 82% for ranitidine (P = 0.03, 95% CI 1-15%). The incidence of adverse events was similar during the two maintenance treatments. Treatment with omeprazole 20 mg every morning maintained significantly more patients in remission than treatment with ranitidine 150 mg at bedtime.

The combination of alprazolam and alcohol on behavioral aggression.

Forty-eight moderate social drinkers were assigned to one of four treatments: alprazolam (1 mg) and alcohol (0.5 g/kg); alprazolam (1 mg) and placebo drink; placebo capsule and alcohol (0.5 g/kg); and placebo capsule and placebo drink. Breath alcohol concentrations and ratings of mood and intoxication were completed at 90, 150 and 210 minutes post-drug (45, 105 and 165 minutes post-alcohol). Subjects competed in a competitive reaction time task at 105 minutes post-drug during which psychophysiological measures were simultaneously monitored. Active treatments increased sedation and intoxication and the task increased feelings of hostility and anxiety in all subjects. Aggressive responding increased in all groups in response to provocation but some stress response dampening was shown after both alcohol and alprazolam on the psychophysiological measures and after alprazolam on subjective ratings of anxiety. The combination of alprazolam and alcohol increased behavioral aggression more than would have been predicted from the sum of the single effects, confirming clinical reports of behavioral dyscontrol.

The effects of alprazolam alone and combined with alcohol on central integrative activity.

The effects of alprazolam 1 mg both alone and in combination with 0.5 g/kg of alcohol were examined on self-ratings of intoxication and measures of central and peripheral activity such as EEG, auditory evoked response, tremor at 90, 150 and 210 min post drug. Forty-eight healthy volunteers were assigned randomly to 4 independent groups who received: alprazolam and placebo drink, alprazolam and alcohol, placebo capsule and alcohol, placebo capsule and placebo drink respectively. Alprazolam decreased the amplitudes of the 3 potentials of the evoked response, decreased activity in the 8-13 Hz and increased activity in the 13.5-26 Hz wavebands of the EEG and decreased the frequency at which fusion was perceived. Alcohol prolonged reaction time and increased tremor. The effects were not always additive and alprazolam was dominant in the combination.

Edpidemia de fiebre aftosa en Bage, Rs, Brasil, 1980. Evaluacion de dos sistemas de vacunacion / Epidemic of foot-and-mouth disease in Bage, Rs, Brazil, 1980. Evaluation of two systems of vaccination

El programa de control de la fiebre aftosa en Rio Grande do Sul, Brasil, se inicio en 1965. Desde 1979 gran parte del municipio de Bage, en este estado, cuenta con un programa demonstrativo de vacunacion con vacuna antiaftosa inactivada y con adyuvante oleoso producida por el Centro Panamericano de Fiebre AFtosa (CPFA). Durante 1980 el citado municipio fue afectado por una epidemia de febre aftosa producida por el virus O1. El presente trabajo analiza las consecuencias morbidas de la enfermedad en el rebaño sometido a este programa de vacunacion con relacion al resto del rebano bovino del municipio que es vacunado con vacuna de hidroxido de aluminio-saponina, inactivada en velhiculo acuoso (denominado sistema "comercial"). La enfermedad se registro en 73 rebaños de los 1.813 sometidos al sistema "comercial" y en 20 de los 620 pertenecientes al programa de vacuna oleosa, lo que dio tasas de rebaños afectados, con ajuste al tamano, de 5 y 2% respectivamente. Las tasas de morbilidad pra la poblacion bovina del municipio fueron de 45 y 6 por mil, respectivamente, para ambos sistemas de vacunacion. Separando las poblaciones que habian recibido solamente una vacunacion con vacuna oleosa al principio de la epidemia de aquellas que recibieron 2 o mas, las diferencias de morbi-mortalidad entre los grupos fueron mas acentuadas, siendo las tasas de morbilidad y mortalidad por mil bovinos respectivamente de 323 y 9,7 para los sometidos a solamente una vacunacion con vacuna oleosa, y de 29 y 0,08 para los que recibieron dos o mas vacunaciones con vacuna oleosa. Se concluye que la vacunacion con adyuvante oleoso aplicada en forma oficial dio resultados altamente satisfactorios teniendo en cuenta la alta exposicion viral a la que estuvo sometida la poblacion del municipio.

O leucograma nos estados de choque. / Leukogram in shock

Os autores fazem um estudo retrospectivo do leucograma de 125 casos de pacientes admitidos em estado de choque no Centro de Tratamento Intensivo do Hospital das Clinicas da UFMG, em periodo de oito anos.Desses pacientes, 70 eram portadores de choque septicemico, 34 de cardiogenico e 2l de hipovolemico. A analise estatistica dos dados observados sugere que, de modo analogo ao que se observa na apendicite aguda, o leucograma do choque septicemico possa ser utilizado como elemento auxiliar do seu diagnostico

Hypokalemic respiratory muscle paralysis following Strongyloides stercoralis hyperinfection: a case report.

Strongyloides stercoralis hyperinfection in a malnourished 6-year-old boy was characterized by severe diarrhea, dehydration and marked hypokalemia, followed by acute respiratory failure due to respiratory muscle paralysis, and cardiac arrest. He was resuscitated and maintained with positive pressure ventilation and intravenous infusion of potassium and gluco-saline solution. These measures produced recovery from the respiratory muscle paralysis. Stool examination revealed eggs and numerous larvae of S. stercoralis, and thiabendazole was initiated. He continued to have severe diarrhea and again developed marked hypokalemia with respiratory muscle paralysis, abdominal distention, and cardiac arrhythmias with cardiac arrest episodes. In spite of fluid and electrolyte replacement the patient died.

Tratamento intensivo do estado de choque. Determinantes clinicos do prognostico. / Intensive treatment of shock state. Clinical determinants of the prognosis

Neste trabalho sao apresentados os resultados do tratamento, em Centro de Tratamento Intensivo, de 356 pacientes em estado de choque. A correlacao entre os dados clinicos e a evolucao foi feita com base numa analise computadorizada dos dados obtidos de 274 tabulacoes; aquelas consideradas mais importantes sao descritas. A mortalidade foi de 63,5% sendo mais elevada (p < 0,01) na presenca de cardiopatia (72,9%), insuficiencia respiratoria (81,4%), acidose metabolica (75,5%) e quando o choque se instalou apos a admissao no CTI (82,6%). As intervencoes terapeuticas nao destinadas ao tratamento do mecanismo primario desencadeante do choque e a presenca de complicacoes do choque (insuficiencia renal aguda, arritmias cardiacas, parada cardiaca e disturbios da consciencia) estiveram relacionadas com elevada mortalidade (p < 0,01). A mortalidade foi tambem influenciada pela etiologia primaria do estado de choque: hipovolemico (45,7%), septicemico (54,8%), cardiogenico (74,7%), misto (75,0%) e indeterminado (81,6%) - (p < 0,01)