Acute endometritis induced by Escherichia coli in mares evaluated through color doppler ultrasonography / [Endometrite aguda induzida por Escherichia coli em éguas avaliadas por ultrassonografia com Doppler colorido]

The objectives of this study were to characterize the endometritis induced in mares using color Doppler ultrasonography and traditional exams. Experiment 1. Mares (n=20) were submitted to intrauterine inoculation with Escherichia coli. Uterine evaluation was performed at M0 and M1. Experiment 2. Animals were divided into two groups: control group (n=10), and treated group (n=10) using phytotherapeutic solution. In both groups, the uterine evaluation was performed at time T1, T2, and T3. Experiment 3: Uterine evaluation was compared after antibiotic therapy, phytotherapy, and M0. For statistical analysis, the Tukey test, t Student, and Anova test were applied. Experiment 1. The mean values of vascularization at M1 were significantly higher than those obtained at M0 (P<0.05). Bacterial growth was observed in all samples collected. Experiment 2. The mean value of vascularization at time T1 in both groups was significantly higher (P<0.05) compared to M2 and M3. Experiment 3. After antibiotic therapy, the vascularization of the body and uterine horns was not equivalent to the vascularization presented at M0. We can conclude that it was not possible to correlate results obtained by color Doppler ultrasonography with the traditional findings for the diagnosis of endometritis.(AU)

Os objetivos deste estudo foram caracterizar a endometrite induzida em éguas utilizando-se a ultrassonografia com Doppler colorido e exames tradicionais. Experimento 1: as éguas (n=20) foram submetidas à inoculação intrauterina com Escherichia coli. A avaliação uterina foi realizada em M0 e M1. Experimento 2: os animais foram divididos em dois grupos: grupo controle (n=10) e grupo tratado (n=10), sendo usada solução fitoterápica. Nos dois grupos, a avaliação uterina ocorreu nos momentos T1, T2 e T3. Experimento 3: a avaliação uterina foi comparada após antibioticoterapia, fitoterapia e M0. Para análise estatística, foram aplicados os testes de Tukey, t de Student e ANOVA. Experimento 1: os valores médios de vascularização em M1 foram significativamente maiores que os obtidos no M0 (P<0,05). Houve crescimento bacteriano em todas as amostras coletadas. Experimento 2: o valor médio da vascularização no tempo T1 nos dois grupos foi significativamente maior (P<0,05) do que o obtido em M2 e M3. Experimento 3: após antibioticoterapia, a vascularização do corpo e dos cornos uterinos não era equivalente à vascularização apresentada em M0. Pode-se concluir que não foi possível correlacionar os resultados obtidos pela ultrassonografia com Doppler colorido com os achados tradicionais para o diagnóstico de endometrite.(AU)

Ultrassonografia testicular no modo color Doppler∕ espectral e avaliação andrológica de garanhões tratados com gonadotrofina coriônica humana (hCG) em diferentes estações do ano / [Testicular ultrasonography in the doppler / spectral color mode and andrological evaluation of stallions treated with human corionic gonadotrofin (hCG) in different seasons of the year]

Utilizaram-se quatro garanhões nos meses de janeiro, abril, julho e outubro de 2016, em dois protocolos, em que: GI (n=4; 5mL de solução salina, i.v.) e GII (n=4; 5000UI de hCG, i.v.) e subdividiram-se esses protocolos em ciclos (C1 e C2), seguindo o esquema crossover, sendo: CI=animais 1 (GI) e 2 (GII), avaliados nos dias D1, D3 e D5, e animais 3 (GI) e 4 (GII), em D2, D4 e D6; CII= animais 1 (GII) e 2 (GI), em D1, D3, D5, e animais 3 (GII) e 4 (GI), em D2, D4 e D6. Realizou-se o tratamento em D1 e D2 de cada ciclo e a ultrassonografia testicular no modo color Doppler e Doppler espectral, uma hora antes de cada coleta de sêmen e imediatamente após. Avaliou-se: número de reflexo de Flehmen, de montas sem ereção, início da monta, tempo de reação à ereção e total da monta e análises de qualidade seminal. Estatisticamente foram utilizados os testes de qui-quadrado e ANOVA. Não houve diferenças estatísticas (P>0,05) entre os parâmetros analisados. Concluiu-se que uma única dose de 5000UI de hCG em garanhões não causou alterações significativas nos parâmetros avaliados em diferentes estações do ano.(AU)

Four stallions were used in January, April, July and October 2016 in two protocols: GI (n=4; 5ml saline, iv) and GII (n=4; 5000 hCG, iv), and these protocols were subdivided into cycles (C1 and C2) following the cross over scheme, as follows: CI=animal 1 (GI) and 2 (GII) evaluated on days D1, D3 and D5 and animal 3 (GI) and 4 (GII) at D 2, D 4 and D 6; CII=animal 1 (GII) and 2 (GI) at D1, D3, D5 and animal 3 (GII) and 4 (GI) at D2, D4 and D6. Treatment was performed on D1 and D2 of each cycle and testicular ultrasound in color Doppler and spectral Doppler mode, one hour before each semen collection and immediately after. We evaluated: Flehmen's reflex number, mounts without erection, start of the mount, reaction time to erection and total mount and seminal quality analyzes. Statistically, the Chi-square and ANOVA tests were used. There were no statistical differences (P>0.05) between the analyzed parameters. It was concluded that a single dose of 5000IU hCG in stallions did not cause significant changes in the parameters evaluated in different seasons of the year.(AU)

Deficiency in proliferative, angiogenic, and LH receptors in the follicle wall: implications of season toward the anovulatory condition.

This study aimed to gain insight on the effect of different seasons of the year on the expression pattern of growth factor and hormone receptors involved in follicle development. A novel follicle wall biopsy technique was used to collect in vivo follicle wall layers (ie, granulosa, theca interna, and theca externa) and follicular fluid samples from growing dominant follicles, simultaneously and repeatedly, using the same mares during the spring anovulatory (SAN), spring ovulatory (SOV), summer (SU), and fall ovulatory (FOV) seasons. The immunofluorescent expression patterns of epidermal growth factor receptor (EGFR), Ki-67, vascular endothelial growth factor receptor (VEGFR), and LH receptor (LHR) were evaluated in each follicle wall layer, in addition to intrafollicular estradiol and nitric oxide (NO). Proliferative proteins (EGFR and Ki-67) were highly (P < 0.05-P < 0.001) expressed during the SOV season compared with the SAN and FOV seasons. Lower (P < 0.05-P < 0.001) expression of both proteins was observed during SU compared with the SOV season. The expression of VEGFR was greater (P < 0.05-P < 0.01) in the theca interna of dominant follicles during the SOV season compared with the SAN and SU seasons. Similarly, in the overall quantification, the VEGFR expression was greater (P < 0.001) during the SOV season compared with the SU and FOV seasons. A higher (P < 0.05) LHR expression was detected in the theca interna during the SOV season than the SAN season. Furthermore, a higher (P < 0.05-P < 0.001) expression of LHR was observed in the granulosa, theca interna, and in the overall quantification during the SOV season compared with the SU and FOV seasons. Intrafollicular NO concentration did not differ (P > 0.05) among different seasons of the year. The intrafollicular estradiol concentration was higher (P < 0.05) during the SU compared with the SAN season and higher (P < 0.05) during the FOV season compared with the SAN and SOV seasons. In conclusion, the synergistic effect of lower expression of proliferative protein, angiogenic, and LH receptors in at least some of the layers of the follicle wall seems to trigger dominant follicles toward the anovulation process during the spring and fall transitional seasons.

Seasonal variation in equine follicular fluid proteome.

BACKGROUND: Proteomic studies of follicular fluid (FF) exist for several species, including the horse; however, the seasonal influence on FF proteome has not been explored in livestock. The application of high-throughput proteomics of FF in horse has the potential to identify seasonal variations of proteins involved in follicle and oocyte growth. METHODS: This study (i) profiles the proteomes of equine FF collected from dominant growing follicles during the spring anovulatory season (SAN), and spring (SOV), summer (SUM), and fall (FOV) ovulatory seasons; and (ii) identifies season-dependent regulatory networks and associated key proteins. RESULTS: Regardless of season, a total of 90 proteins were identified in FF, corresponding to 63, 72, 69, and 78 proteins detected in the SAN, SOV, SUM, and FOV seasons, respectively. Fifty-two proteins were common to all seasons, a total of 13 were unique to either season, and 25 were shared between two seasons or more. Protein-to-protein interaction (PPI) analysis indicated the likely critical roles of plasminogen in the SAN season, the prothrombin/plasminogen combination in SUM, and plasminogen/complement C3 in both SOV and FOV seasons. The apolipoprotein A1 appeared crucial in all seasons. The present findings show that FF proteome of SUM differs from other seasons, with FF having high fluidity (low viscosity). CONCLUSIONS: The balance between the FF contents in prothrombin, plasminogen, and coagulation factor XII proteins favoring FF fluidity may be crucial at the peak of the ovulatory season (SUM) and may explain the reported lower incidence of hemorrhagic anovulatory follicles during the SUM season.

In vivo antral follicle wall biopsy: a new research technique to study ovarian function at the cellular and molecular levels.

BACKGROUND: In vivo studies involving molecular markers of the follicle wall associated with follicular fluid (FF) milieu are crucial for a better understanding of follicle dynamics. The inability to obtain in vivo samples of antral follicle wall (granulosa and theca cells) without jeopardizing ovarian function has restricted advancement in knowledge of folliculogenesis in several species. The purpose of this study in mares was to develop and validate a novel, minimally invasive in vivo technique for simultaneous collection of follicle wall biopsy (FWB) and FF samples, and repeated collection from the same individual, during different stages of antral follicle development. We hypothesized that the in vivo FWB technique provides samples that maintain the normal histological tissue structure of the follicle wall layers, offers sufficient material for various cellular and molecular techniques, and allows simultaneous retrieval of FF. METHODS: In Experiment 1 (ex vivo), each follicle was sampled using two techniques: biopsy forceps and scalpel blade (control). In Experiment 2 (in vivo), FWB and FF samples from 10-, 20-, and 30-mm follicles were repeatedly and simultaneously obtained through transvaginal ultrasound-guided technique. RESULTS: In Experiment 1, the thickness of granulosa, theca interna, and theca externa layers was not influenced (P > 0.05) by the harvesting techniques. In Experiment 2, the overall recovery rates of FWB and FF samples were 97 and 100%, respectively. However, the success rate of obtaining samples with all layers of the follicle wall and clear FF varied according to follicle size. The expression of luteinizing hormone receptor (LHR) was mostly confined in the theca interna layer, with the estradiol-related receptor alpha (ERRα) in the granulosa and theca interna layers. The 30-mm follicle group had greater (P < 0.05) LHR expression in the theca interna and ERRα in the granulosa layer compared to the other groups. The overall expression of LHR and ERRα, and the intrafollicular estradiol were higher (P < 0.05 - P < 0.0001) in the 30-mm follicle group. CONCLUSION: The in vivo technique developed in this study can be repeatedly and simultaneously used to provide sufficient FWB and FF samples for various cellular and molecular studies without jeopardizing the ovarian function, and has the potential to be translated to other species, including humans.

Use of different pressures for transvaginal follicular aspiration in mares / Uso de diferentes pressões para aspiração folicular transvaginal em éguas

The success of transvaginal follicular aspiration in mares can be influenced by several factors, such as vacuum pump pressure levels. The present study aimed to investigate the effect of different negative pressures (150, 280 and 400mmHg) of the vacuum pump on the oocyte recovery in the mares. The mares (n=10) were undergoing follicular aspiration using three different negative pressures for three consecutive estrous cycles as follows: G150 = 150mmHg (n = 10); G280 = 280mmHg (n = 10); G400 = 400mmHg (n = 10). Every estrous cycle, the group that the mare would participate was drawn, and each animal participated once in each group. Only preovulatory follicle was used, about 30 to 36 hours after application of hCG. To compare the results, the chi-square test was used (5% significance) and Fisher exact test, when recommended. Thirty preovulatory follicles (diameter 36.1±1.80mm) were aspirated and ten oocytes were recovered (33.3%). There was no statistical difference between the experimental groups (p=0.59). Thus, accord to the results observed in this study, we could conclude that the negative pressure of the vacuum pump used was not efficient to increase oocyte recovery.(AU)

O sucesso da técnica de aspiração folicular transvaginal em éguas pode ser influenciado de maneira determinante por diversos fatores, tais como níveis de pressão da bomba de vácuo. Diante disso, o presente experimento visou investigar o efeito de diferentes pressões negativas (150, 280 e 400mmHg) da bomba de vácuo sobre a taxa de recuperação de oócitos em éguas. As éguas (n=10) foram submetidas à aspiração folicular utilizando-se três diferentes pressões negativas por três ciclos estrais consecutivos, da seguinte maneira: G150= 150mmHg (n=10); G280= 280mmHg (n=10); G400= 400mmHg (n=10). A cada ciclo estral, sorteava-se o grupo do qual a égua participaria, sendo que cada animal integrou um grupo somente uma vez. Foi puncionado somente folículo pré-ovulatório, em torno de 30 a 36 horas após a aplicação do hCG. Os resultados foram comparados utilizando-se o teste qui-quadrado (a 5% de significância) e o Fisher Exato, quando recomendados. Foram aspirados 30 folículos pré-ovulatórios (diâmetro 36,1±1,80mm) e recuperados 10 oócitos (33,3%). Não houve diferença estatística entre os grupos experimentais (P=0,59). Dessa forma, mediante os resultados obtidos no presente estudo, foi possível concluir que a pressão negativa da bomba de vácuo utilizada não se mostrou determinante para elevar a recuperação oocitária.(AU)