Uterine secretory activity and histology of prepubertal female dogs in an animal model of partial ablation of endometrial glands development / Atividade secretória uterina e histologia de cadelas pré-púberes em um modelo animal de ablação parcial do desenvolvimento das glândulas uterinas

Inadequate exposure of the female reproductive system to steroids in uterine developmental periods can partially inhibit the development of endometrial glands in dogs. However, the effects of steroids on the formed glands functionality remain unknown, as well as the possible occurrence of endometrial fibrosis. This study aimed to evaluate the secretory activity of endometrial glands in prebubertal female dogs submitted to a protocol of partial ablation of the uterine adenogenesis. Sixteen females of non-specific breed were distributed into two groups; MPA (n=8), females that received applications of medroxyprogesterone acetate every 3 weeks; and C (n=8) untreated control females. Ovariohysterectomy was performed in all animals at the age of 6 months and evaluated the uterine horns by histological and histochemistry exams. The secretion intensity (degrees 1-4) was evaluated using periodic acid-Schiff (PAS) and alcian blue (AB) pH 2.5. Histological evaluation was performed using Masson's trichrome and toluidine blue. Only degree 1 and 2 marks for PAS were observed in both groups, with no difference of uterine secretion intensity between the groups regarding the degrees found. However, the MPA group revealed higher intensity of uterine secretion compared to group C (p<0.05). Staining with AB pH 2.5 also revealed only degree 1 and 2 marks in both groups, with no statistically significance between them. Masson's trichrome staining revealed no marks in the periglandular region in both groups. A higher among of mast cells was observed in the myometrial region of the uterus in both groups. Prepubertal female dogs with partial ablation of the uterine adenogenesis present minimal uterine secretory activity, absence of periglandular fibrosis and increased presence of mast cells in the myometrium compared to endometrium.(AU)

A exposição inadequada do sistema reprodutor feminino a esteróides em períodos do desenvolvimento uterino pode inibir parcialmente o desenvolvimento das glândulas endometriais em cães. Entretanto, não se conhece os efeitos dos esteróides sobre a funcionalidade das glândulas formadas, bem como a possível ocorrência de fibrose endometrial. Objetivou-se avaliar a atividade secretória das glândulas endometriais de cadelas pré-púberes submetidas a protocolo de ablação parcial da adenogênese uterina. Foram utilizadas 16 fêmeas, sem-raça-definida, distribuídas nos grupos MPA (n=8), fêmeas que receberam aplicações de acetato de medroxiprogesterona a cada 3 semanas, e C (n=8), fêmeas controle não tratadas. Aos seis meses de idade, foi realizada ovariohisterectomia em todos os animais, e avaliados os cornos uterinos pelo exame histológico e de histoquímica. Para avaliar a intensidade de secreção (graus 1-4), foram utilizadas periodic acid-Schiff e alcian blue (AB) pH 2,5. Para a avaliação histológica foram utilizados tricrômico de Masson e azul de toluidina. Apenas marcações graus 1 e 2 foram observadas para PAS em ambos os grupos, sem diferença na intensidade de secreção uterina entre grupos com relação aos graus encontrados. Entretanto, o grupo MPA apresentou maior intensidade de secreção uterina em relação ao grupo C (p<0,05). Com relação ao AB pH 2,5, em ambos os grupos também foram encontradas apenas marcações de graus 1 e 2, sem diferença estatística entre grupos. Não foram observadas marcações para a coloração de tricrômico de Masson na região periglandular, em ambos os grupos. Foi observada maior quantidade de mastócitos presentes no útero na região do miométrio, em ambos os grupos. Conclui-se que cadelas pré-púberes com ablação parcial da adenogênese uterina apresentam mínima atividade secretória uterina, ausência de fibrose periglandular e maior presença de mastócitos no miométrio em relação ao endométrio.(AU)

Uterine secretory activity and histology of prepubertal female dogs in an animal model of partial ablation of endometrial glands development / Atividade secretória uterina e histologia de cadelas pré-púberes em um modelo animal de ablação parcial do desenvolvimento das glândulas uterinas

Inadequate exposure of the female reproductive system to steroids in uterine developmental periods can partially inhibit the development of endometrial glands in dogs. However, the effects of steroids on the formed glands functionality remain unknown, as well as the possible occurrence of endometrial fibrosis. This study aimed to evaluate the secretory activity of endometrial glands in prebubertal female dogs submitted to a protocol of partial ablation of the uterine adenogenesis. Sixteen females of non-specific breed were distributed into two groups; MPA (n=8), females that received applications of medroxyprogesterone acetate every 3 weeks; and C (n=8) untreated control females. Ovariohysterectomy was performed in all animals at the age of 6 months and evaluated the uterine horns by histological and histochemistry exams. The secretion intensity (degrees 1-4) was evaluated using periodic acid-Schiff (PAS) and alcian blue (AB) pH 2.5. Histological evaluation was performed using Masson's trichrome and toluidine blue. Only degree 1 and 2 marks for PAS were observed in both groups, with no difference of uterine secretion intensity between the groups regarding the degrees found. However, the MPA group revealed higher intensity of uterine secretion compared to group C (p<0.05). Staining with AB pH 2.5 also revealed only degree 1 and 2 marks in both groups, with no statistically significance between them. Masson's trichrome staining revealed no marks in the periglandular region in both groups. A higher among of mast cells was observed in the myometrial region of the uterus in both groups. Prepubertal female dogs with partial ablation of the uterine adenogenesis present minimal uterine secretory activity, absence of periglandular fibrosis and increased presence of mast cells in the myometrium compared to endometrium.(AU)

A exposição inadequada do sistema reprodutor feminino a esteróides em períodos do desenvolvimento uterino pode inibir parcialmente o desenvolvimento das glândulas endometriais em cães. Entretanto, não se conhece os efeitos dos esteróides sobre a funcionalidade das glândulas formadas, bem como a possível ocorrência de fibrose endometrial. Objetivou-se avaliar a atividade secretória das glândulas endometriais de cadelas pré-púberes submetidas a protocolo de ablação parcial da adenogênese uterina. Foram utilizadas 16 fêmeas, sem-raça-definida, distribuídas nos grupos MPA (n=8), fêmeas que receberam aplicações de acetato de medroxiprogesterona a cada 3 semanas, e C (n=8), fêmeas controle não tratadas. Aos seis meses de idade, foi realizada ovariohisterectomia em todos os animais, e avaliados os cornos uterinos pelo exame histológico e de histoquímica. Para avaliar a intensidade de secreção (graus 1-4), foram utilizadas periodic acid-Schiff e alcian blue (AB) pH 2,5. Para a avaliação histológica foram utilizados tricrômico de Masson e azul de toluidina. Apenas marcações graus 1 e 2 foram observadas para PAS em ambos os grupos, sem diferença na intensidade de secreção uterina entre grupos com relação aos graus encontrados. Entretanto, o grupo MPA apresentou maior intensidade de secreção uterina em relação ao grupo C (p<0,05). Com relação ao AB pH 2,5, em ambos os grupos também foram encontradas apenas marcações de graus 1 e 2, sem diferença estatística entre grupos. Não foram observadas marcações para a coloração de tricrômico de Masson na região periglandular, em ambos os grupos. Foi observada maior quantidade de mastócitos presentes no útero na região do miométrio, em ambos os grupos. Conclui-se que cadelas pré-púberes com ablação parcial da adenogênese uterina apresentam mínima atividade secretória uterina, ausência de fibrose periglandular e maior presença de mastócitos no miométrio em relação ao endométrio.(AU)

Colony aging affects the reproductive performance of Swiss Webster females used as recipients for embryo transfer

The objective was to evaluate the influence of colony aging in a Swiss Webster (SW) outbred stock used as recipients for embryo transfer. In the first study, a retrospective analysis was performed throughout several generations during a 38-month period in 2,398 embryos transferred to 108 SW recipients. A decrease in the percentage of live pups from transferred embryos was found at the end of the period. Impairment occurred due to the incidence of maternal cannibalism that increased from 0% to 67-100% (P 0.05), while pregnancy rate (pregnant/transferred recipients) and number of pups per delivered female were not affected throughout the period (P=NS). A following study was carried out to compare the reproductive performance of SW stock vs. B6D2F1 hybrid females in a 5-year interval. The study was conducted on a total of 893 embryos transferred to 40 females (20 SW and 20 B6D2F1) in Year /1, and 514 embryos transferred to 30 females (15 SW and 15 B6D2F1) in Year /5. No cases of maternal cannibalism were found on Year /1 in any of the strains (0/10 and 0/10). However, an incidence of 44,4% (4/9) was seen on Year /5 for SW, while for B6D2F1 the incidence was 0% (0/12) (P 0.05). Further examination of the uterus showed endometrial cysts and abnormal implantation sites in SW on Year /5 but not in B6D2F1 females. In conclusion, this study reports an impairment of the reproductive performance of an early aged SW outbred stock colony mainly due to the occurrence of maternal cannibalism. This finding has important implications for embryo transfer programs conducted in mouse facilities.

Colony aging affects the reproductive performance of Swiss Webster females used as recipients for embryo transfer

The objective was to evaluate the influence of colony aging in a Swiss Webster (SW) outbred stock used as recipients for embryo transfer. In the first study, a retrospective analysis was performed throughout several generations during a 38-month period in 2,398 embryos transferred to 108 SW recipients. A decrease in the percentage of live pups from transferred embryos was found at the end of the period. Impairment occurred due to the incidence of maternal cannibalism that increased from 0% to 67-100% (P 0.05), while pregnancy rate (pregnant/transferred recipients) and number of pups per delivered female were not affected throughout the period (P=NS). A following study was carried out to compare the reproductive performance of SW stock vs. B6D2F1 hybrid females in a 5-year interval. The study was conducted on a total of 893 embryos transferred to 40 females (20 SW and 20 B6D2F1) in Year /1, and 514 embryos transferred to 30 females (15 SW and 15 B6D2F1) in Year /5. No cases of maternal cannibalism were found on Year /1 in any of the strains (0/10 and 0/10). However, an incidence of 44,4% (4/9) was seen on Year /5 for SW, while for B6D2F1 the incidence was 0% (0/12) (P 0.05). Further examination of the uterus showed endometrial cysts and abnormal implantation sites in SW on Year /5 but not in B6D2F1 females. In conclusion, this study reports an impairment of the reproductive performance of an early aged SW outbred stock colony mainly due to the occurrence of maternal cannibalism. This finding has important implications for embryo transfer programs conducted in mouse facilities.(AU)

Quantification of collagen fibers in canine uteri treated with medroxyprogesterone acetate

Collagen plays essential roles in remodeling uterine tissue during decidualization, implantation, pregnancy and involution. To understand whether the progestational agent medroxyprogesterone acetate (MPA) can modify the organization and deposit of collagen in the uteri of normal bitches (Canis Tlupus familiaris), we assessed uterine tissues by histochemistry. Uteri were grouped as: nulliparous (n=11), multiparous (n=11) and treated with MPA (n=11; nulliparous with two treatments; 5mg/kg; i.m.). The amount, location and birefringence of interstitial collagen types I and III in the fold and base of the endometrial stroma and the myometrial muscular layers were studied on sections stained with Picrosirius Red by polarized light microscopy and evaluated by ANOVA. No differences were observed in the endometrium. In the myometrium, differences were observed in collagen type I between MPA-treated and nulliparous uteri vs. multiparous (p<0.05), and differences in collagen type III between nulliparous and multiparous uteri vs. MPA-treated (p=0.0001). In conclusion, two doses of MPA had no significant effect on the investigated collagens in the extracellular matrix.(AU)

Quantification of collagen fibers in canine uteri treated with medroxyprogesterone acetate

Collagen plays essential roles in remodeling uterine tissue during decidualization, implantation, pregnancy and involution. To understand whether the progestational agent medroxyprogesterone acetate (MPA) can modify the organization and deposit of collagen in the uteri of normal bitches (Canis Tlupus familiaris), we assessed uterine tissues by histochemistry. Uteri were grouped as: nulliparous (n=11), multiparous (n=11) and treated with MPA (n=11; nulliparous with two treatments; 5mg/kg; i.m.). The amount, location and birefringence of interstitial collagen types I and III in the fold and base of the endometrial stroma and the myometrial muscular layers were studied on sections stained with Picrosirius Red by polarized light microscopy and evaluated by ANOVA. No differences were observed in the endometrium. In the myometrium, differences were observed in collagen type I between MPA-treated and nulliparous uteri vs. multiparous (p<0.05), and differences in collagen type III between nulliparous and multiparous uteri vs. MPA-treated (p=0.0001). In conclusion, two doses of MPA had no significant effect on the investigated collagens in the extracellular matrix.(AU)

The infectious disease epidemiologic triangle of bovine uterine diseases

Postpartum uterine diseases are important for animal welfare and economic reasons, causing cow discomfort, elimination from the herd and impaired reproductive performance. Metritis is characterized as an abnormally enlarged uterus and a fetid, watery, red-brown uterine discharge within 21 days after parturition. Endometritis is defined as inflammation of the endometrium after 21 days postpartum without systemic signs of illness, and can be considered the chronic stage of uterine inflammation. It has been reported that the metritis affects 10 to 20% of cows, and endometritis affects 5.3 to 52.6% of cows. Metritis affects the cow systemically, and has a negative impact on milk production and reproductive performance. Cows affected with endometritis are not systemically ill, and do not have their milk production altered; however, they have impaired reproductive performance. Metritis and endometritis are complex multifactorial diseases, and a wide range of factors contributes to their occurrence. They are often associated with mixed bacterial infection of the uterus, and the major pathogens associated with uterine diseases are Escherichia coli, Trueperella pyogenes and Fusobacterium necrophorum. Events during the transition period related to negative energy balance and metabolic imbalance, mineral deficiencies, leading to immunosuppression are of great important during establishment of intrauterine bacterial infections. This, combined with endometrium trauma events during parturition (such as calving related problems), and environmental factors (poor hygiene at calving, housing type and calving season), increases the risk of metritis and endometritis.(AU)

The infectious disease epidemiologic triangle of bovine uterine diseases

Postpartum uterine diseases are important for animal welfare and economic reasons, causing cow discomfort, elimination from the herd and impaired reproductive performance. Metritis is characterized as an abnormally enlarged uterus and a fetid, watery, red-brown uterine discharge within 21 days after parturition. Endometritis is defined as inflammation of the endometrium after 21 days postpartum without systemic signs of illness, and can be considered the chronic stage of uterine inflammation. It has been reported that the metritis affects 10 to 20% of cows, and endometritis affects 5.3 to 52.6% of cows. Metritis affects the cow systemically, and has a negative impact on milk production and reproductive performance. Cows affected with endometritis are not systemically ill, and do not have their milk production altered; however, they have impaired reproductive performance. Metritis and endometritis are complex multifactorial diseases, and a wide range of factors contributes to their occurrence. They are often associated with mixed bacterial infection of the uterus, and the major pathogens associated with uterine diseases are Escherichia coli, Trueperella pyogenes and Fusobacterium necrophorum. Events during the transition period related to negative energy balance and metabolic imbalance, mineral deficiencies, leading to immunosuppression are of great important during establishment of intrauterine bacterial infections. This, combined with endometrium trauma events during parturition (such as calving related problems), and environmental factors (poor hygiene at calving, housing type and calving season), increases the risk of metritis and endometritis.

Changes in various endometrial proteins during cloprostenol-induced failure of early pregnancy in mares

Various major proteins in uterine secretions flushed from mares on day 20 of gestation have been identified and related to levels of gene expression in endometrial biopsies. Pregnancy-related changes were determined by comparison with samples from mares on day 20 after being given a luteolytic IM dose of cloprostenol on day 18, and with cycling mares that were not pregnant on days 17-20. Proteins in cell-free uterine flush samples were identified by LC-MS/MS of trypsin- digested peptides. Levels of endometrial expression were estimated from raw signal strength in Agilent 21322 E. caballus expression microarray. In this paper we describe a subset of pregnancy-related endometrial proteins with potential roles in embryonic development and/or mucosal innate immunity. Endometrial proteins that increased greatly during pregnancy included GM2 activator (GM2A), lipocalin 2 (LCN2), stanniocalcin 1 (STC1) and uterine serpin (SERPINA14). Endometrial proteins that decreased in normal pregnancy included secretory phospholipase A2 (sPLA2), secretoglobin 1A1 (SCGB1A1), and vanin 1 (VNN1). Cloprostenol-induced failure of pregnancy resulted in increased endometrial expression of SPLA2, cathepsin L1 (CTSL1), interleukin-1 receptor antagonist (IL1RN), SCGB1A1 epididymal secretory protein E1 (NPC2), connective tissue growth factor (CTGF), retinol binding protein 4 (RBP4), uteroferrin/TRAP5 (ACP5), SERPINA14, annexin A1 (ANXA1), annexin A3 (ANXA3), and vanin 3 (VNN3). Expression of P19 uterocalin (P19) was similarly high in pregnant and non-pregnant endometrium, and virtually undetectable in the yolk-sac wall. High levels of expression of ANXA2, GM2A, NPC2, CTSL1 and RBP4 in both endometrium and yolk-sac wall suggests that these have important roles on both sides of the maternal:conceptus interface. The properties and distribution of GM2A and NPC2 suggest that they are important in the transport of phospholipids and sterols. SPLA2, LCN2, SCGB1A1 and IL1RA have potential roles in mucosal innate immunity but their production is reduced during early pregnancy. These studies provide an inventory of many proteins in the uterine lumen during early pregnancy, and a subset for which local endometrial expression is altered when pregnancy is compromised by cloprostenol-induced luteolysis.

Changes in various endometrial proteins during cloprostenol-induced failure of early pregnancy in mares

Various major proteins in uterine secretions flushed from mares on day 20 of gestation have been identified and related to levels of gene expression in endometrial biopsies. Pregnancy-related changes were determined by comparison with samples from mares on day 20 after being given a luteolytic IM dose of cloprostenol on day 18, and with cycling mares that were not pregnant on days 17-20. Proteins in cell-free uterine flush samples were identified by LC-MS/MS of trypsin- digested peptides. Levels of endometrial expression were estimated from raw signal strength in Agilent 21322 E. caballus expression microarray. In this paper we describe a subset of pregnancy-related endometrial proteins with potential roles in embryonic development and/or mucosal innate immunity. Endometrial proteins that increased greatly during pregnancy included GM2 activator (GM2A), lipocalin 2 (LCN2), stanniocalcin 1 (STC1) and uterine serpin (SERPINA14). Endometrial proteins that decreased in normal pregnancy included secretory phospholipase A2 (sPLA2), secretoglobin 1A1 (SCGB1A1), and vanin 1 (VNN1). Cloprostenol-induced failure of pregnancy resulted in increased endometrial expression of SPLA2, cathepsin L1 (CTSL1), interleukin-1 receptor antagonist (IL1RN), SCGB1A1 epididymal secretory protein E1 (NPC2), connective tissue growth factor (CTGF), retinol binding protein 4 (RBP4), uteroferrin/TRAP5 (ACP5), SERPINA14, annexin A1 (ANXA1), annexin A3 (ANXA3), and vanin 3 (VNN3). Expression of P19 uterocalin (P19) was similarly high in pregnant and non-pregnant endometrium, and virtually undetectable in the yolk-sac wall. High levels of expression of ANXA2, GM2A, NPC2, CTSL1 and RBP4 in both endometrium and yolk-sac wall suggests that these have important roles on both sides of the maternal:conceptus interface. The properties and distribution of GM2A and NPC2 suggest that they are important in the transport of phospholipids and sterols. SPLA2, LCN2, SCGB1A1 and IL1RA have potential roles in mucosal innate immunity but their production is reduced during early pregnancy. These studies provide an inventory of many proteins in the uterine lumen during early pregnancy, and a subset for which local endometrial expression is altered when pregnancy is compromised by cloprostenol-induced luteolysis.(AU)

Developmental aspects of early pregnancy in mares

In mares, the embryo mi grates to the uterus between days 5 and 6 postovulation beginning its mobility through all uterine segments, which is essential for the maternal recognition of pregnancy. During the mobility phase, the embryonic vesicle shows a linear growth rate until its fixation between days 15 and 17, when the orientation phenomenon occurs. From fixation to day 28 of pregnancy, the embryonic growth is less evident (plateau) by cross-section ultrasound examination. After this period the linear growth rate is reestablished until day 46. This plateau is attributed to the increased uterine tone that compresses the vesicle and to volume expansion, making it difficult to detect the conceptus growth only by the cross-section diameter. Around day 20, the embryo proper is visualized as an echogenic spot in the ventral aspect of the vesicle. Additionally, development of allantoic sac, embryonic heartbeat, yolk sac regression and posterior umbilical cord formation also can be visualized from days 20 to 40. An intimate interaction between uterus and conceptus is essential for the normal pregnancy development. Color- and spectral-Doppler ultrasonography can be useful for the evaluation of this interface. A gradual increase on uterine vascularity during the early pregnancy and transient changes in endometrial vascularity accompanying the vesicle location during the mobility phase have been described. Around day 38 of gestation, the formation of the endometrial cups begins and, consequently, the synthesis of the equine chorionic gonadotropin (eCG) induces the formation and development of supplementary corpora lutea, which are important to secrete progesterone and to maintain pregnancy until around day 120.

Developmental aspects of early pregnancy in mares

In mares, the embryo mi grates to the uterus between days 5 and 6 postovulation beginning its mobility through all uterine segments, which is essential for the maternal recognition of pregnancy. During the mobility phase, the embryonic vesicle shows a linear growth rate until its fixation between days 15 and 17, when the orientation phenomenon occurs. From fixation to day 28 of pregnancy, the embryonic growth is less evident (plateau) by cross-section ultrasound examination. After this period the linear growth rate is reestablished until day 46. This plateau is attributed to the increased uterine tone that compresses the vesicle and to volume expansion, making it difficult to detect the conceptus growth only by the cross-section diameter. Around day 20, the embryo proper is visualized as an echogenic spot in the ventral aspect of the vesicle. Additionally, development of allantoic sac, embryonic heartbeat, yolk sac regression and posterior umbilical cord formation also can be visualized from days 20 to 40. An intimate interaction between uterus and conceptus is essential for the normal pregnancy development. Color- and spectral-Doppler ultrasonography can be useful for the evaluation of this interface. A gradual increase on uterine vascularity during the early pregnancy and transient changes in endometrial vascularity accompanying the vesicle location during the mobility phase have been described. Around day 38 of gestation, the formation of the endometrial cups begins and, consequently, the synthesis of the equine chorionic gonadotropin (eCG) induces the formation and development of supplementary corpora lutea, which are important to secrete progesterone and to maintain pregnancy until around day 120.(AU)

Remodelamento dinâmico da matriz extracelular endometrial modula a receptividade em bovinos

A matriz extracelular do endométrio (ECM) é constituída por moléculas secretadas que compõem o microambiente celular e são ativadas ou suprimidas principalmente pelos hormônios esteróides ovarianos, estradiol (E2) e progesterona (P4) durante o ciclo estral. A identificação de genes envolvidos no remodelamento e receptividade pode levar à descoberta de importantes processos biológicos ligados ao sucesso gestacional. Os objetivos foram: 1. identificar a relação de differentes tamanhos de folículos pré-ovulatórios (FPO) e corpo lúteo (CL) e de seus respectivos hormônios E2 e P4 com a expressão endometrial de genes associados com o remodelamento da ECM durante o período de pré-implantação; e 2. analisar a relação entre a expressão gênica de determinados componentes da ECM avaliada no dia 6 após inseminação artificial (IA) com sucesso gestacional. Para tal, dois experimentos foram realizados. No experimento 1, estudo 1 e estudo 2, 42 e 74 vacas Nelore (Bos indicus) adultas, respectivamente foram sincronizadas obtendo-se ao final dois grupos com distintos tamanhos FPO e CL consequentemente, distintas concentrações de E2 no proestro e P4 no diestro. Os grupos foram: Folículo Grande-CL Grande (FG-CLG; estudo 1, n=20; estudo 2, n=35) e Folículo Pequeno-CL Pequeno (FP-CLP; estudo 1, n=22; estudo 2, n=39). Amostras de tecido endometrial foram coletadas por biópsia no D0 (estro) e pós-mortem no D4 (estudo 1) e D7 (estudo 2). Concentrações de E2 e P4 foram mensuradas por radioimunoensaio (RIA) obtendo-se menores concentrações no grupo FP-CLP. No experimento 2, vacas adultas, Nelore (Bos indicus; n=33) foram sincronizadas utilizando um protocol a base de prostaglandina 2 (PGF2) e observação de estro. As vacas foram inseminadas artificialmente (IA) e seis dias após, uma biópsia endometrial coletada. O diagnóstico de gestação foi realizado após 30 dias por meio de ultrasonografia (US) e então as vacas foram divididas em grupo Prenhe e Não-Prenhe (P e NP) para análise retrospectiva. Abundância de transcritos foi avaliada por sequenciamento (RNAseq) assim como qPCR em amostras de ambos experimentos. Realizaram-se também exames histológicos em amostras do D4 e D7 (estudo 2) para avaliação de colágeno total assim como espessura de fibras colágenas. Resultados determinaram uma maior abundância de transcritos relacionados ao remodelamento de MEC, em destaque TGF, MMPs, TIMPs e colágenos em vacas pertencentes aos grupos NP e FP-CLP. O mesmo foi observado para abundância de colágeno. No entanto, não observou-se diferença na relação entre fibras grossas e finas entre os tratamentos. Análises de correlação e regressão indicaram que folículos pré-ovulatórios de maior tamanho geram CL maiores e assim maiores concentrações de P4, a qual está negativamente associada à abundância de colágenos. Assim, de acordo com resultados aqui descritos, assume-se que a alteração da homeostase da MEC devido ao incremento na abundância de colágeno pode ser prejudicial à gestação em bovinos.

The endometrial extracellular matrix (ECM) é made up of secretory molecules that make up the cellular microenvironment and suffer activation or suppression mainly by the ovarian steroid hormones, estradiol (E2) and progesterone (P4) during the estrous cycle. The identification of genes that are involved in endometrial remodeling and receptivity may reveal important biological processes linked to gestational success. The objectives were: 1. identify the relationship preovulatory follicle (POF) size and corpus luteum (CL) and its respective hormones, E2 and P4 on the endometrial expression of genes related to extracellular matrix remodeling during the pre-implantation period; and 2. analyze the relationship between endometrial ECM gene expression evaluated on day 6 post artificial insemination (AI) and its relationship to pregnancy success. For such, two experiments were carried on. On experiment 1, study 1 and study 2, 42 and 74, respectively, adult Nelore (Bos indicus) cows were synchronized aiming to manipulate the peri-ovulatory endocrine environment, obtaining at the end of the protocol, two groups with distinct pre-ovulatory follicle (POF) and corpus luteum (CL) sizes, leading to groups with distinct E2 and P4 concentrations. The groups were: Large Follicle/CL (LF/CL; study 1, n=20, study 2, n=35) and Small Follicle/Cl (SF/CL; study 1, n=22; study 2, n=39). Endometrial samples were collected by biopsy on D0 (Estrus) and post-mortem on D4 and on study 2 post-mortem on D7. P4 and E2 concentrations were measured by RIA with a significative difference between the groups, being lower hormonal concentrations in the SF-SCL group and higher concentrations in the LF-LCL group . In experiment 2, adult Nelore (Bos indicus) cows (n=33) were synchronized using a prostaglandin 2 (PGF2) and heat detection based protocol. The cows were AI and six days after an endometrial biopsy was collected. Pregnancy diagnosis was performed on day 30 by ultrasound (US) examination and cows were divided into pregnant and non-pregnant (P vs. NP) groups for a retrospective analysis. Histology was performed on D4 and D7 samples for total collagen abundance as well as fiber thickness. Correlation and regression analysis indicate that larger preovulatory follicles as well as higher P4 concentrations have a negative effect on collagen content. RNA-Seq analysis and confirmation by qPRC was performed on selected samples from experiment 1, study 2 and experiment 2. Comparison of mRNA levels of ECM components samples revealed higher levels of transcripts envolved in ECM remodeling, highlighting TGF, MMPs, TIMPs and collagnes in NP cows when compared with P cows as well as in the SF-SCL compared to the LF-LCL group. The same was observed for collagen abundance. However, there was no difference between thin and thick collagen fibers between treatments. Correlation and regression analysis indicate that larger POF lead to larger CL and hence, higher P4 concentrations, which has a negative effects on collagen abundance. Therefore, according to the results presented here, we can imply that an alteration in ECM homeostasis due to increased collagen abundance may be harmful to pregnancy in cows.

ANALISE MORFOLÓGICA E HISTOQUIMICA DA ENDOMETRIOSE INDUZIDA EM RATAS APOS O TRATAMENTO COM EXTRATO ETANÓLICO DE Bumelia sartorum Mart

A endometriose é um processo inflamatório, caracterizado pela presença de implantes de endométrio fora da cavidade uterina. Atualmente, as opções terapêuticas para a endometriose, incluem terapia médica ou cirurgia. Contudo, grande parte da população mundial ainda utiliza plantas medicinais, como por exemplo a Bumelia sartorum Mart, popularmente conhecida como quixaba, para algumas infecções do aparelho reprodutor feminino. Assim, resolveu-se testar a hipótese se o tratamento de ratas induzidas a endometriose com o extrato etanólico da casca de B. sartorum causa uma regressão dos implantes endometriais e se provoca alterações nos níveis hormonais de estrógeno, progesterona e prolactina. Foram utilizadas 25 ratas albinas, com 90 dias de idade, divididas em cinco grupos: GI ratas induzidas a endometriose (Controle); GII ratas induzidas a endometriose, tratadas com placebo (álcool 20%) por 15 dias; GIII ratas induzidas a endometriose, tratadas com extrato da quixaba por 05 dias; GIV ratas induzidas a endometriose, tratadas com extrato da quixaba por 10 dias; GV ratas induzidas a endometriose, tratadas com extrato da quixaba por 15 dias. O extrato foi administrado por gavagem, numa concentração de 32 mg/mL/rata. Os resultados mostraram que os implantes do GI, GII e GIII apresentaram grandes cistos, infiltrado leucocitário e presença intensa de fibras colágenas, que são características da endometriose. Entretanto, nos grupos GIV e GV houve redução dos cistos, do infiltrado leucocitário e presença de macrófagos. A análise do extrato etanólico por CLAE/EM, Infravermelho e UV/Vis mostrou a presença de 11% do monômero de catequina/epicatequina, 60% dos seus dímeros e 3% dos seus trímeros. Houve redução dos níveis de estrógeno e aumento da progesterona apenas nas ratas do grupo GV e os níveis de prolactina não diferiram entre os grupos experimentais. Os grupos GIII, GIV e GV apresentaram os maiores índices apoptóticos, enquanto que apenas os grupos GIV e GV mostraram redução de glândulas e vasos sanguíneos nos implantes. Assim, conclui-se que o extrato etanólico bruto de B. sartorum é rico em compostos fenólicos, que pode promover uma regressão dos implantes endometriais e controla os níveis hormonais, quando utilizado por 15 dias na concentração de 32 mg/mL. Entretanto, experimentos adicionais com os fracionados do extrato etanólico bruto devem ser feitos para esclarecer melhor os mecanismos de ação envolvidos na atividade antiinflamatória na endometriose

Endometriosis is an inflammatory process, characterized by the presence of endometrial implants outside the uterine cavity. Currently, treatment options for endometriosis include medical therapy or surgery. However, much of the world's population still use medicinal plants, such as Bumelia sartorum Mart, popularly known as quixaba for some infections of the female reproductive system. So we decided to test the hypothesis that the treatment of rats induced endometriosis with the ethanol extract of the bark of B. sartorum causes a regression of endometrial implants and causes changes in hormone levels of estrogen, progesterone and prolactin. A total of 25 albino rats, 90 days old, were divided into five groups: GI- endometriosis-induced mice (control); GII- endometriosis induced famale rats treated with placebo (20% ethanol) for 15 days; GIII- famale rats induced endometriosis treated with quixaba extract for 05 days; GIV- famale rats induced endometriosis treated with quixaba extract for 10 days; GV- famale rats induced endometriosis treated with quixaba extract for 15 days. The extract was administered by gavage at a concentration of 32 mg / ml / rat. The results showed that the implants of GI, GII and GIII showed large cysts, leukocyte infiltration and intense presence of collagen fibers, which are characteristics of endometriosis. However, in the GIV and GV groups there was a reduction of the cysts, leukocyte infiltration and presence of macrophages. The analysis of the ethanol extract by HPLC / MS, IR and UV / Vis showed the presence of 11% of the monomer of catechin / epicatechin, 60% of dimers and trimers of their 3%. There was a reduction in estrogen levels and increased progesterone only in rats GV group, and prolactin levels did not differ between experimental groups. The GIII, GIV, and GV groups had the highest apoptotic indices, whereas only GIV, and GV groups showed a reduction glands and blood vessels in the implants. Thus, it is concluded that the ethanol extract of B. sartorum is rich in phenolic compounds, which can promote regression of endometrial implants and controls hormone levels when used for 15 days at a concentration of 32 mg / ml. However, further experiments with the fractionated ethanol extract should be made to clarify the mechanisms involved in the anti-inflammatory activity in endometriosis.

Aspectos histopatológicos da adenomiose em úteros bovinos nas diferentes fases do ciclo estral / Histophatological aspects of adenomyosis in bovine uteri in different phases of the estrous cycle

Relacionaram-se as características da adenomiose com as fases do ciclo estral em 61 peças de úteros de bovinos colhidas em matadouros. A adenomiose foi classificada em superficial e profunda. A fase do ciclo estral foi estimada pela morfologia, pela coloração e pela vascularização do corpo lúteo e presença ou não de folículos ovarianos maiores que 8mm. Os animais que estavam em anestro (n=11) apresentaram a menor ocorrência de adenomiose (8,2 por cento), e os que estavam na fase lútea média (n=21), a maior (31,0 por cento). Nas fases lútea inicial (n=13) e folicular (n=16) as ocorrências foram semelhantes, 18,0 e 22,9 por cento, respectivamente. A maior porcentagem de adenomiose profunda ocorreu nas fases lútea inicial e média, 45,0 e 47,4 por cento, respectivamente, e durante o anestro e a fase folicular foram de 20,0 e 14,3 por cento, respectivamente. Os resultados sugerem que a fase do ciclo estral influencia na ocorrência de adenomiose e no grau de infiltração miometrial das glândulas endometriais(AU)

The relationship of the adenomyosis characteristics and the phases of the estrus cycle in 61 cows bovine's uteruses collected in slaughterhouses was studied. The adenomyoses were classified as superficial and deep. The morphology, staining and vascularization of the corpus luteum and the presence or not of larger ovarian follicles than 8mm helped to estimate tthe estrus cycle. The cows in anestrus (n=11) showed the least occurrence of adenomyosis (8.2 percent) and the animals in the medium luteal phase (n=21) the largest one(31,0 percent). In the initial luteal phase (n=13) and the follicular phase (n=16) the occurrences of adenomyosis were similar and equals to 18.0 and 22.9 percent, respectively. The largests percentage of deep adenomyosis were found in the initial and in the medium luteal phases, 45.0 and 47.4 percent, respectively, and during the anestrus and the follicular phase they were 20.0 and 14.3 percent, respectively. The data suggest that the cycle phase influences in adenomyosis occurrence and in the degree of miometrial infiltration of the endometrial glands(AU)

Experimental model for endometriosis. Comparative histological study between the ectopic and eutopic endometrium

Thyrty-six rats werw implanted with endometrial squares (2 X 2mm) to the left parietal peritoneum and the right uterine horn was tied. The rats were mated for 20 days and then sacrificed. Topic and ectopic endometrial histological pattern were compared. There was a correlation between the histological pattern of the autografted endometrium and the eutopic endometrium im most rats, pregnant or not. This suggests that both responded in a similar way to the same endocrine stimuli. The experimental model for endometriosis admiting the histological correlation between the uterine endometrium and the ectopic is factible and it is easy to be repetead.

Trinta e seis ratas foram submetidas a implante cirúrgico de retalho endometrial quadrangular (2 x 2mm) no peritônio parietal esquerdo e ligadura do corno uterino direito. As ratas foram acasaladas por 20 dias e logo após sacrificadas. Foram comparados o padrão histológico do endométrio eutópico e do ectópico.Houve correspondência entre o padrão histológico do endométrio auto-transplantado e o endométrio eutópico na maioria das ratas, prenhes ou não, sugerindo que ambos responderam de forma semelhante aos mesmo estímulos endócrinos. O modelo experimental de endometriose na rata, permitindo a correlação histológica entre o endométrio uterino e o ectópico, é viável e de fácil repetição.