Abstract Since bull fertility prediction remains challenging, the identification of potential fertility markers is important considering the economic benefits to the livestock industry. The main goal of this study was to determine the Na/K-ATPase activity and expression in thawed sperm of high (HF)- and low-fertility (LF) Angus bulls. Samples from three different batches/bulls with HF (n = 4) and LF (n = 4) were used. The Na/K-ATPase activity was determined after thawing, whereas sperm kinematics, membrane integrity, and expression of Na/K-ATPase on sperm surface were evaluated immediately post-thaw and after 120 minutes of incubation. Within the same incubation time, there was no difference on sperm membrane integrity, kinematics, and the expression of Na/K-ATPase on the sperm surface between HF and LF bulls. Kinematic parameters of LIN and VCL were not influenced by incubation time in samples from HF and LF, respectively. A tendency (P = 0.06) of higher Na/K-ATPase enzymatic activity for sperm of HF bulls compared to LF bulls was observed (0.49 ± 0.07 and 0.32 ± 0.06, respectively). In conclusion, Na/K-ATPase activity and expression in thawed sperm from Angus bulls are not related to the fertility index after fixed-time artificial insemination. However, sperm kinematics related to hyperactivation might indicate higher sperm cryotolerance for HF bulls. challenging industry Na/KATPase NaKATPase Na/K ATPase Na K (HF) lowfertility low (LF batchesbulls batches n 4 used thawing integrity postthaw post thaw 12 respectively P 0.06 006 0 06 0.49 049 49 (0.4 007 07 0.0 032 32 0.3 . respectively) conclusion fixedtime fixed insemination However KATPase NaK (HF 1 00 0.4 04 (0. 0. 03 3 (0 (

Abstract Although equine blastocysts ≤ 300 µm in diameter can be successfully vitrified, larger equine blastocysts are not good candidates for cryopreservation. As Na+, K+-ATPase is involved in maintaining blastocyst expansion, perhaps inhibition of this enzyme would be a viable method of reducing blastocyst diameter prior to cryopreservation. Objectives were to evaluate effects of ouabain-induced inhibition of Na+, K+-ATPase in equine blastocysts. Sixteen mares were ultrasonographically monitored, given deslorelin acetate to induce ovulation, and inseminated. Embryos (D7 and D9) were harvested and Na+, K+-ATPase inhibited for 1 or 6 h by exposure to 10-6 M ouabain, either natural ouabain or conjugated to fluorescein (OuabainFL), during incubation at 37° C. Evaluations included morphometric characteristics (bright field microscopy) and viability (Hoescht 33342 + propidium iodide). Blastocysts incubated for 6 h in Holding medium + ouabain (n=3) had, on average, a 45.7% reduction in diameter, with adverse morphologic features and no re-expansion after subsequent incubation in Holding medium for 12 h. In subsequent studies, even a 1-h exposure to Ouabain or OuabainFL, caused similar reductions, namely 38.7 ± 6.7% (n=5) and 33.6 ± 3.3% (n=7) for D7 and D9 blastocysts, respectively. Ouabain binding was confirmed after OuabainFL exposition and all embryos (n=12) lost viability. We concluded that Na+, K+-ATPase inhibition with ouabain caused death of equine blastocysts and therefore was not a viable method of reducing blastocyst size prior to cryopreservation.

Abstract The objective of this study was to investigate the need of seminal plasma removal for short-term cooling of buck semen in soybean lecithin (SL) based extender. Each pool was divided equally, and one half was subjected to centrifugation to remove seminal plasma (SP-), while the other half remained with seminal plasma (SP+). Then, both SP+ and SP- samples were diluted in two SL extenders (extender A = 1% SL; extender B = 2% SL), cooled to 5ºC and stored for 48 hours. The sperm kinetics, evaluated by CASA, and plasma membrane integrity (PMI), acrosomal integrity (ACI) and high mitochondrial membrane potential (HMMP), evaluated by epifluorescence microscopy, were determined within five minutes after reaching 5°C (T0), as well as after 24 (T24) and 48 (T48) hours of storage. Interactions (seminal plasma vs. extender vs. time;) were observed for all variables assessed. Total and progressive motility and other variables of sperm kinetics decreased after 24 hours of cooling in the SP+ group, and after 48 hours of storage, these same variables were lower in SP+/B compared to SP-/B groups. Furthermore, SP+ reduced PMI (extender B, T48), HMMP (A and B extenders, T48) and ACI (extender A, T0) compared to SP- samples. The interactions between seminal plasma and soybean lecithin phospholipids seemed to occur in a time-dependent manner. It was concluded that the removal of seminal plasma improves the quality of goat semen that was cooled in a soybean lecithin-based extender, especially when using 2% soybean lecithin.

ABSTRACT: Owing to the lack of information in the literature about the ideal feeding frequency adopted by feedlot systems, this study was designed to evaluate the effect of different feeding frequencies on performance, carcass traits, feeding behavior and rumen morphometrics of Nellore cattle. Forty-eight yearling bulls were housed in individual pens with an average initial body weight of 358.2 kg. The experiment was designed as a completely randomized block and the treatments were the feeding of cattle one, two, three, or four times daily (n = 12 per treatment). Orthogonal contrasts were used to assess linear, quadratic and cubic relationship between feeding frequency and the dependent variable. As feeding frequency increased, final body weight (p = 0.03) and average daily gain (p = 0.01; one: 0.98 kg d–1; two: 0.96 kg d–1; three: 1.18 kg d–1; four: 1.14 kg d–1) increased linearly and the feed:gain ratio decreased (p = 0.02). Also, as feeding frequency increased, hot carcass weight was affected cubically (p = 0.01; one: 246.9 kg; two: 243.0 kg; three: 263.1 kg; four: 258.1 kg). During the adaptation period dry matter intake variation was affected cubically (p ≤ 0.05; one: 16 %; two: 23 %; three: 14 %; four: 16 %). Increases in feeding frequency affected papillae height (p < 0.01) and papillae surface area (p < 0.01) cubically, as well as producing increases in the mitotic index (p = 0.05; one: 9 %; two: 11 %; three: 13 %; four: 13 %) and cells in proliferation (p = 0.05) linearly. Thus, Nellore yearling bulls should be fed three times daily in feedlot operations.

RESUMO OBJETIVO: Avaliar o impacto da embolização arterial de miomas (EAM) sobre o volume uterino (VU), na função ovariana. MÉTODOS: Trinta pacientes com leiomioma se submeteram à EAM. Foram realizados exames de USPTV e FSH antes e três meses após a EAM. Foram analisados o VU em cm³, o diâmetro do mioma dominante (DMD) em cm e o FSH em UI/mL, expressos por média desvio padrão (DP) e submetidos a análise estatística pelo teste não paramétrico de Mann-Whitney. RESULTADOS: Foram incluidos na análise 29 casos. A média do VU pré-EAM foi 402,4 165,9 cm³, DMD pré-EAM 5,9 2,1 cm. O VU pós-EAM foi 258,9 118,6 cm³, DMD pós-EAM foi 4,6 1,8 cm. A média da dosagem de FSH pré-EAM foi 4,9 3,5 UI/mL e pós-EAM foi 5,5 4,7 UI/mL com p=0,5. Houve redução de 35% do VU, de 22% no DMD e a EAM não alterou significativamente os valores de FSH após três meses. CONCLUSÃO: O procedimento diminui significativamente o VU e DMD e, não há aumento significativo dos níveis séricos de FSH, não havendo, portanto, alterações na função ovariana.

PURPOSE: To evaluate the impact of uterine artery embolization (UAE) on uterine volume (UV), greater myoma diameter (GMD) and ovarian function three months after the procedure, by transvaginal pelvic ultrasonography (TVPUS) and by the determination of follicle-stimulating hormone (FSH). METHODS: Thirty patients with leiomyomas were submitted to UAE. TVPUS and FSH determination were performed before and three months after UAE. UV was determined in cm³, GMD in cm and FSH in IU/mL. Data are reported as as mean standard deviation (SD) and were analyzed statistically by the nonparametric Mann-Whitney test. RESULTS: Twenty-nine patients were analyzed. Before UAE, mean UV was 402.4 165.9 cm³ and GMD was 5.9 2.1 cm. After UAE, mean UV was 258.9 118.6 cm³ and GMD was 4.6 1.8 cm. Mean FSH concentration was 4.9 3.5 IU/mL before UAE and 5.5 4.7 IU/mL after UAE, with p=0.5. There was a 35% reduction of UV and a 22% reduction of GMD, with no changes in FSH values after three months. CONCLUSION: The procedure significantly reduced UV and GMD but did not cause a significant increase in FSH levels, thus causing no changes in ovarian function.

OBJETIVO: analisar as repercussões histomorfométricas da embolização das artérias uterinas (EAU) no tecido uterino, especialmente mediante quantificação de tecido colágeno, através de biópsia uterina antes e após tratamento de leiomioma uterino. MÉTODOS: participaram do estudo 15 pacientes portadoras de leiomiomas sintomáticos e/ou com infertilidade, submetidas à EAU após ciência do termo de consentimento livre e esclarecido, obedecendo aos critérios de exclusão do estudo. Foi realizada biópsia uterina na fase secretória do ciclo menstrual antes e três meses após o procedimento, para avaliação do colágeno. Após o processamento histológico do material, foram feitos cortes de 3µ, sendo alguns corados pela hematoxilina-eosina (HE), e outros pela coloração específica para fibras colágenas (Picrosirius red). Em seguida, foi realizada a leitura e interpretação das lâminas e a quantificação do colágeno. Sua quantificação foi calculada como o percentual da área composta por colágeno, e o resultado expresso em média±desvio padrão (DP). Os dados foram então submetidos à análise estatística pelo teste t de Student pareado (p<0,05). RESULTADOS: nas biópsias realizadas antes do tratamento, foi notada a presença de células musculares lisas, rodeadas por rica trama de fibras colágenas que compõem o tumor, vasos sanguíneos e núcleos de fibroblastos. Nas lâminas das biópsias realizadas após o tratamento, foi observada a presença de necrose de coagulação difusa, trombose vascular, áreas de calcificação e de infiltração linfoplasmocitária e nítida diminuição do componente colágeno. A porcentagem de fibras colágenas foi maior no grupo pré-EAU (84,07±1,41) do que no grupo pós-EAU, (81,05±1,50), com p<0,0001, e intervalo de confiança de 95% (IC95%) entre 2,080 e 3,827. CONCLUSÃO: a redução quantitativa e qualitativa do colágeno evidencia que o tratamento proposto é eficaz em reduzir a massa tumoral, composta principalmente por fibras colágenas de permeio às células musculares lisas neoplásicas. Todavia, são necessários estudos complementares a fim de se investigar a repercussão funcional e biológica dessas alterações histológicas.

PURPOSE: to analyze histomorphometric consequences of the uterine arteries embolization (UAE) in the uterine tissue, especially by collagen tissue quantification through uterine biopsy, before and after treatment of uterine leiomyoma. METHODS: 15 patients with symptomatic leyomioma and/or infertility, submitted to UAE, participated in the study according to the study exclusion criteria, after having signed an informed consent. Uterine biopsy was performed in the secretory phase of the menstrual cycle, before and three months after the procedure, to evaluate the collagen. After the histological processing of the material, 3 µ slices were prepared, some of them dyed with hematoxiline-eosin (HE) and others with the specific dye for collagen fibers (Picrosirius red). Then, the slides were examined and interpreted, and the collagen quantified. The amount was calculated as the percent of the area composed by collagen, and the result expressed in mean±standard deviation (SD). Data has then been submitted to statistical analysis by Student's paired t test (p<0.05). RESULTS: the presence of smooth muscle cells was observed in the biopsies performed before the treatment, surrounded by a rich network of collagen fibers, which are part of the tumor, blood vessels and fibroblast nuclei. On the slides of biopsies performed after the treatment, it was observed the presence of widespread coagulation necrosis, vascular thrombosis, calcification and lymphoplasmocitary infiltration areas and clear reduction of the collagen component. The percentage of collagen fibers was higher in the pre-UAE group (84.07±1.41), than in the post-UAE (81.05±1.50) group, with p<0.0001, and 95% confidence interval (CI95%) from 2.080 to 3.827. CONCLUSION: the quantitative and qualitative collagen reduction clearly shows that the proposed treatment is efficient in reducing the tumoral mass, composed mainly by collagen fibers intermingled with neoplasic smooth muscle cells. Nevertheless, complementary studies are needed to investigate the functional and biological consequences of these histological changes.

O objetivo deste estudo foi identificar as proteínas do plasma seminal de caprinos da raça Alpina Americana criados na região Nordeste do Brasil que estão relacionadas ao índice pluviométrico e à qualidade do sêmen. O sêmen foi obtido pelo método de vagina artificial a partir de três reprodutores e foi avaliado quanto aos parâmetros macroscópicos e microscópicos. O perfil de proteínas do plasma seminal foi realizado por eletroforese bidimensional. Os parâmetros volume do sêmen, integridade do acrossoma e proteínas totais evidenciaram diferença significativa (P<0,05) entre os períodos de alto (1,7mL, 90,3% e 372µg mL-1, respectivamente) e baixo (1,2mL, 80,3% e 494µg mL-1, respectivamente) índice pluviométrico. Foram detectados, nos períodos de alto e baixo índice pluviométrico, 47 e 49 spots de proteínas com massa molecular relativa de 4 a 106kDa e de 15 a 97kDa e ponto isoelétrico de 3,00 a 8,96 e de 4,48 a 9,83, respectivamente. Apenas no período de alto índice pluviométrico foram observados os grupo de proteínas de 13kDa e 45kDa. Conclui-se que o sêmen de caprinos da raça Alpina Americana criados na região Nordeste do Brasil apresenta-se com melhor qualidade quando colhido no período de alto índice pluviométrico, o que pode ser atribuído à presença das proteínas de 13kDa e 45kDa.

The aim of this study was to identify proteins in seminal plasma of goats raised in the Northeast of Brazil related with precipitation index and semen quality. Semen was obtained from three bucks and evaluated to the microscopic and macroscopic parameters. The profile of seminal plasma proteins was performed by analysis of two-dimensional electrophoresis. Volume, acrosome integrity and total proteins had significant difference (P<0.05) between the periods of high (1.7mL, 90.3% and 372g mL-1, respectively) and low (1.2mL, 80.3% and 494µg mL-1, respectively) precipitation index. It was detected during high and low precipitation index, 47 and 49 spots of proteins with molecular weight of 4 to 106kDa and 15 to 97kDa, and isoelectric point of 3.00 to 8.96, and 4.48 to 9.83, respectively. Only in the period of high precipitation index were observed groups of proteins with 13kDa and 45kDa. It can be concluded that semen of Alpine American goats raised in the Northeast of Brazil has best quality when obtained in the period of high precipitation index, which can be attributed to the presence of protein with 13kDa and 45kDa.