Regulation of progesterone during follicular development by FSH and LH in sheep

Progesterone (P4) can participate in the development of female mammalian antral follicles through nuclear receptor (PGR). In this experiment, the differences of P4 synthesis and PGR expression in different developmental stages of sheep antral follicles (large > 5mm, medium 2-5mm, small < 2mm) were detected by enzyme-linked immunosorbent assay, immunohistochemistry, qRT-PCR and Western blotting. Secondly, sheep follicular granulosa cells were cultured in vitro. The effects of different concentrations of FSH and LH on P4 synthesis and PGR expression were studied. The results showed that acute steroid regulatory protein (StAR), cholesterol side chain lyase (P450scc) and 3β Hydroxysteroid dehydrogenase (3β-HSD) and PGR were expressed in antral follicles, and with the development of antral follicles in sheep, StAR, P450scc and the expression of 3β-HSD and PGR increased significantly. In vitro experiments showed that FSH and LH alone or together treatment could regulate P4 secretion and PGR expression in sheep follicular granulosa cells to varying degrees, hint P4 and PGR by FSH and LH, and LH was the main factor. Our results supplement the effects of FSH and LH on the regulation of P4 synthesis during follicular development, which provides new data for further study of steroid synthesis and function in follicular development.(AU)

Comparative analysis of the hormone production and gene expression profiles in ovine uterus tissue during oestrus cycle synchronized using medroxyprogesterone acetate plus eCG and prostaglandin analogue / Análise comparativa da produção de hormônios e perfis de expressão gênica em tecido uterino ovino durante o ciclo estral sincronizado usando acetato de medroxiprogesterona mais eCG e análogo de prostaglandina

The combination of medroxyprogesterone acetate (MPA) and gonadotropin chorionic (eCG) has been widely used to synchronize oestrus cycle in sheep, but their effects on the gene expression in uterine tissue are yet to be elucidated. To evaluate the effect of MPA + eCG or prostaglandin analogue (PA) treatments on the rate of oestrus cycle synchronization, as well as further hormone production and gene expression profiles in uterine tissue, 14 Santa Inês ewes were randomly selected. The MPA + eCG group (n=7) received intravaginal insertion of MPA-impregnated sponges for 14 days and was administered 350 IU eCG on the day of sponge withdrawal. The PA group (n=7) was administered two doses of 100 µg of PA separated by 12 days. The ewes were assessed for the rate of oestrus cycle synchronization and the serum concentrations of progesterone (P4) and estradiol (E2). Additionally, the expression of estrogen receptor (ERα), progesterone receptor (P4R), and immunolocalization of interferon receptor (IFNAR1) in the uterine tissue samples collected 15th day post-mating were examined. The rate of oestrus cycle synchronization was 100% (n=7/7) and 57.14% (n=4/7) in the MPA + eCG and PA groups, respectively. Moreover, the MPA + eCG group exhibited higher serum concentration of P4 than the PA group (p < 0.05). However, the E2 serum concentration did not differ between the two groups (p > 0.05). The relative expression of P4R and ERα mRNA analyzed using real-time PCR and immunodetection of IFNAR1 were similar between the two groups tested (p > 0.05). Conclusively, MPA + eCG treatment improved the rate of oestrus cycle synchronization and endogenous P4 production; however, it did not affect the expression of sex steroid receptors and IFNAR1 in uterine ovine tissue.(AU)

A combinação de acetato de medroxiprogesterona (MPA) com gonadotrofina coriônica (eCG) é amplamente utilizada para sincronizar o estro de ovelhas, mas seus possíveis efeitos sobre a expressão gênica em tecido uterino não foram elucidados. Para avaliar o efeito dos protocolos MPA + eCG ou análogo de prostaglandina (PA) sobre a taxa de sincronização do estro, bem como na futura produção hormonal e expressão gênica em tecido uterino, 14 ovelhas Santa Inês foram selecionadas. O grupo MPA + eCG (n=7) recebeu a inserção de esponjas impregnadas de MPA por via intravaginal por 14 dias e 350 UI de eCG no dia da retirada da esponja. O grupo PA recebeu duas doses de 100 µg de PA administradas com 12 dias de intervalo. As ovelhas foram avaliadas quanto à taxa de sincronização do estro, concentração sérica de progesterona (P4) e estradiol (E2). Além disso, foram examinadas a expressão do receptor de estradiol (ERα), receptor de progesterona (P4R) e localização do receptor de interferon (IFNAR1), a partir de amostras de tecido uterino coletadas 15 dias após o acasalamento. A taxa de sincronização do estro foi 100% (n = 7/7) e 57,14% (n = 4/7) nos grupos MPA + eCG e PA, respectivamente. Além disso, o grupo MPA + eCG apresentou maior concentração sérica de P4 em comparação com o grupo PA (P < 0,05). No entanto, a concentração sérica de E2 não diferiu entre os grupos testados (P > 0,05). A expressão relativa de RNAm para P4R e ERα analisado por PCR em tempo real e imunodetecção de IFNAR1 foi semelhante entre os grupos testados (P > 0,05). Em conclusão, o tratamento com MPA + eCG melhora a taxa de sincronização do estro e a produção de progesterona endógena; contudo, não afeta a regulação da expressão de receptores de esteroides sexuais e IFNAR1 no tecido uterino de ovinos.(AU)

Progesterone and estrogen receptor expression by canine cutaneous soft tissue sarcomas / Expressão de receptores para progesterona e estrógeno em sarcomas de tecidos moles cutâneos em cães

Canine soft tissue sarcomas (STS) comprise a heterogeneous group of malignancies that share similar histopathological features, a low to moderate recurrence rate and low metastatic potential. In human medicine, the expression of estrogen receptors (ER) and progesterone receptors (PR) in sarcomas has been studied to search for prognostic factors and new treatment targets. Similar studies have yet to be conducted in veterinary medicine. The objective of this study was therefore to investigate by immunohistochemistry (IHC) the ER and PR expression in a series of 80 cutaneous and subcutaneous sarcomas in dogs with histopathological features of peripheral nerve sheath tumor (PNST) and perivascular wall tumor (PWT). All cases were positive for PR and negative for ER. Tumors of high malignancy grade (grade III) exhibited higher PR expression than low-grade tumors (grade I). Tumors with mitotic activity greater than 9 mitotic figures/10 high power fields also exhibited higher PR expression. In addition, there was a positive correlation between cell proliferation (Ki67) and PR expression. Therefore, it is possible that progesterone plays a greater role than estrogen in the pathogenesis of these tumors. Future studies should explore the potential for selective progesterone receptor modulators as therapeutic agents in canine STS, as well as evaluating PR expression as a predictor of prognosis.(AU)

Sarcomas de tecidos moles (STM) caninos compreendem um grupo heterogêneo de neoplasias malignas, que apresentam alterações histopatológicas similares, baixa a moderada taxa de recorrência e baixo potencial metastático. Em medicina humana, a expressão de receptor para estrógeno (RE) e receptor para progesterona (RP) nos sarcomas tem sido estudada, visando a busca por fatores prognósticos e novos alvos para tratamentos. Na medicina veterinária, ainda não foram realizados estudos similares. O objetivo deste trabalho foi investigar por imuno-histoquímica a expressão de RE e RP em uma série de 80 sarcomas cutâneos e subcutâneos de cães, com características histopatológicas de tumor de bainha de nervo periférico e tumor de parede perivascular. Todos os casos foram positivos para RP e negativos para RE. Tumores de alto grau de malignidade (grau III) exibiram maior expressão deste receptor que os tumores de baixo grau (grau I). Tumores com atividade mitótica maior que 9 figuras mitóticas/10 campos de grande aumento também exibiram maior expressão do RP. Em adição, houve correlação positiva entre o índice de proliferação celular (Ki67) e a expressão de RP. Assim, é possível que a progesterona desempenhe maior papel que o estrógeno na patogênese desses tumores. Futuros trabalhos poderão explorar o potencial dos moduladores seletivos de RP como agente terapêutico em STM caninos, bem como avaliar a expressão de RP como preditiva de prognóstico.(AU)

Progesterone and estrogen receptor expression by canine cutaneous soft tissue sarcomas / Expressão de receptores para progesterona e estrógeno em sarcomas de tecidos moles cutâneos em cães

Canine soft tissue sarcomas (STS) comprise a heterogeneous group of malignancies that share similar histopathological features, a low to moderate recurrence rate and low metastatic potential. In human medicine, the expression of estrogen receptors (ER) and progesterone receptors (PR) in sarcomas has been studied to search for prognostic factors and new treatment targets. Similar studies have yet to be conducted in veterinary medicine. The objective of this study was therefore to investigate by immunohistochemistry (IHC) the ER and PR expression in a series of 80 cutaneous and subcutaneous sarcomas in dogs with histopathological features of peripheral nerve sheath tumor (PNST) and perivascular wall tumor (PWT). All cases were positive for PR and negative for ER. Tumors of high malignancy grade (grade III) exhibited higher PR expression than low-grade tumors (grade I). Tumors with mitotic activity greater than 9 mitotic figures/10 high power fields also exhibited higher PR expression. In addition, there was a positive correlation between cell proliferation (Ki67) and PR expression. Therefore, it is possible that progesterone plays a greater role than estrogen in the pathogenesis of these tumors. Future studies should explore the potential for selective progesterone receptor modulators as therapeutic agents in canine STS, as well as evaluating PR expression as a predictor of prognosis.(AU)

Sarcomas de tecidos moles (STM) caninos compreendem um grupo heterogêneo de neoplasias malignas, que apresentam alterações histopatológicas similares, baixa a moderada taxa de recorrência e baixo potencial metastático. Em medicina humana, a expressão de receptor para estrógeno (RE) e receptor para progesterona (RP) nos sarcomas tem sido estudada, visando a busca por fatores prognósticos e novos alvos para tratamentos. Na medicina veterinária, ainda não foram realizados estudos similares. O objetivo deste trabalho foi investigar por imuno-histoquímica a expressão de RE e RP em uma série de 80 sarcomas cutâneos e subcutâneos de cães, com características histopatológicas de tumor de bainha de nervo periférico e tumor de parede perivascular. Todos os casos foram positivos para RP e negativos para RE. Tumores de alto grau de malignidade (grau III) exibiram maior expressão deste receptor que os tumores de baixo grau (grau I). Tumores com atividade mitótica maior que 9 figuras mitóticas/10 campos de grande aumento também exibiram maior expressão do RP. Em adição, houve correlação positiva entre o índice de proliferação celular (Ki67) e a expressão de RP. Assim, é possível que a progesterona desempenhe maior papel que o estrógeno na patogênese desses tumores. Futuros trabalhos poderão explorar o potencial dos moduladores seletivos de RP como agente terapêutico em STM caninos, bem como avaliar a expressão de RP como preditiva de prognóstico.(AU)

Immunohistochemical localization of progesterone receptors alpha (PRA) in ovary of the pseudopregnant rabbit

Progesterone plays an important role in the reproductive function and follicular development in mammals. The aim of the present study was to examine the localization of progesterone receptor alpha (PRA) in ovary of pseudopregnant rabbit by immunohistochemical methods. Samples were collected from 14 h. to 18 days of pseudopregnancy. At the first stage of pseudopregnancy (14 h.), the rabbit ovary showed moderate immunostaining of PRA in the granulosa cells and theca interna cells of preovulatory follicle and in the stroma cells. At the middle stage of pseudopregnancy (3-7 days), the rabbit ovary showed strong immunostaining of PRA in ovarian surface epithelial cells, follicular cells of the primary follicle, granulosa cells and theca interna cells of the growing and antral follicles. Moderate immunoexpression of PRA were observed in the large lutein cells and endothelial cells of the corpus haemorrhagicum and corpus luteum and in the stroma cells. At the end of pseudopregnancy (18 days) strong PRA reactions were detected in the small lutein cells of the regressed corpus luteum. Moderate to strong PRA immuno-expression were observed in the proliferated theca interna cells of the atretic antral follicles. The atretic large lutein cells of the regressed corpus luteum showed negative immunostaining for PRA. This study showed that the PRA positive small lutein cells of the regressed corpus luteum and the PRA positive proliferated theca interna cells of the atretic antral follicles were transformed into PRA positive interstitial gland cells. In conclusion, the present study had described the distribution of PRA in the ovary of pseudopregnant rabbit, which is not discussed before in the available literature. It also gives more information about follicular dynamic, formation and origin of interstitial glands, mechanism of ovulation, formation and regression of the corpus luteum.

Immunohistochemical localization of progesterone receptors alpha (PRA) in ovary of the pseudopregnant rabbit

Progesterone plays an important role in the reproductive function and follicular development in mammals. The aim of the present study was to examine the localization of progesterone receptor alpha (PRA) in ovary of pseudopregnant rabbit by immunohistochemical methods. Samples were collected from 14 h. to 18 days of pseudopregnancy. At the first stage of pseudopregnancy (14 h.), the rabbit ovary showed moderate immunostaining of PRA in the granulosa cells and theca interna cells of preovulatory follicle and in the stroma cells. At the middle stage of pseudopregnancy (3-7 days), the rabbit ovary showed strong immunostaining of PRA in ovarian surface epithelial cells, follicular cells of the primary follicle, granulosa cells and theca interna cells of the growing and antral follicles. Moderate immunoexpression of PRA were observed in the large lutein cells and endothelial cells of the corpus haemorrhagicum and corpus luteum and in the stroma cells. At the end of pseudopregnancy (18 days) strong PRA reactions were detected in the small lutein cells of the regressed corpus luteum. Moderate to strong PRA immuno-expression were observed in the proliferated theca interna cells of the atretic antral follicles. The atretic large lutein cells of the regressed corpus luteum showed negative immunostaining for PRA. This study showed that the PRA positive small lutein cells of the regressed corpus luteum and the PRA positive proliferated theca interna cells of the atretic antral follicles were transformed into PRA positive interstitial gland cells. In conclusion, the present study had described the distribution of PRA in the ovary of pseudopregnant rabbit, which is not discussed before in the available literature. It also gives more information about follicular dynamic, formation and origin of interstitial glands, mechanism of ovulation, formation and regression of the corpus luteum.(AU)

Participação de membros da superfamília TGF beta na foliculogênese, luteinização e luteólise / Role of TGF-beta superfamily members during folliculogenesis, luteinization and luteolysis

Fatores produzidos no ovário como os membros da família dos fatores de crescimento transformantes beta (TGFβ) e seus receptores, são essenciais durante o desenvolvimento folicular. Membros desta superfamília desempenham papel chave na fertilidade e diferenças espécie-específicas na regulação desses fatores têm sido descritas, envolvendo as funções ovarianas em condições fisiológicas ou patológicas. A proteína morfogenética óssea 15 (BMP15) e o fator de crescimento e diferenciação 9 (GDF9) destacam-se, pois desempenham papéis importantes na regulação do crescimento e da diferenciação folicular. Ainda, há evidências de que outras BMPs, ativinas, inibinas e seus receptores também possam estar envolvidos no controle da foliculogênese, ovulação/luteinização e luteólise. A maioria dos dados demostram que os TGFs atuam regulando negativamente a síntese de progesterona, o que sugere envolvimento na inibição da luteinização e promoção da luteólise. O avanço no entendimento das funções destes fatores locais poderá possibilitar o desenvolvimento tanto de novas estratégias contraceptivas, como também para controle do ciclo estral ou menstrual.(AU)

Factors produced in the ovary, such as transforming growth factors beta members (TGFβ) and their receptors, play a key role during follicular development. Members from this family have an important role in female fertility and species-specific differences in their regulation have been described, being involved in ovarian function regulation under both physiological and pathological conditions. Bone morphogenetic protein 15 (BMP15) and growth and differentiation factor 9 (GDF9) are the most studied factors due to their involvement in the regulation of follicular development and differentiation. Besides BMP15 and GDF9, other BMPs, activins, inhibins and their receptors may be involved in the control of folliculogenesis, ovulation/luteinization and luteolysis. Most studies demonstrate that TGFβ members negatively regulate progesterone synthesis, suggesting an involvement in luteolysis. The advance in the knowledge of the function of these local factors may allow the development of new contraceptive strategies as well as new approaches to control the estrous or menstrual cycle.(AU)

Participação de membros da superfamília TGF beta na foliculogênese, luteinização e luteólise / Role of TGF-beta superfamily members during folliculogenesis, luteinization and luteolysis

Fatores produzidos no ovário como os membros da família dos fatores de crescimento transformantes beta (TGFβ) e seus receptores, são essenciais durante o desenvolvimento folicular. Membros desta superfamília desempenham papel chave na fertilidade e diferenças espécie-específicas na regulação desses fatores têm sido descritas, envolvendo as funções ovarianas em condições fisiológicas ou patológicas. A proteína morfogenética óssea 15 (BMP15) e o fator de crescimento e diferenciação 9 (GDF9) destacam-se, pois desempenham papéis importantes na regulação do crescimento e da diferenciação folicular. Ainda, há evidências de que outras BMPs, ativinas, inibinas e seus receptores também possam estar envolvidos no controle da foliculogênese, ovulação/luteinização e luteólise. A maioria dos dados demostram que os TGFs atuam regulando negativamente a síntese de progesterona, o que sugere envolvimento na inibição da luteinização e promoção da luteólise. O avanço no entendimento das funções destes fatores locais poderá possibilitar o desenvolvimento tanto de novas estratégias contraceptivas, como também para controle do ciclo estral ou menstrual.

Factors produced in the ovary, such as transforming growth factors beta members (TGFβ) and their receptors, play a key role during follicular development. Members from this family have an important role in female fertility and species-specific differences in their regulation have been described, being involved in ovarian function regulation under both physiological and pathological conditions. Bone morphogenetic protein 15 (BMP15) and growth and differentiation factor 9 (GDF9) are the most studied factors due to their involvement in the regulation of follicular development and differentiation. Besides BMP15 and GDF9, other BMPs, activins, inhibins and their receptors may be involved in the control of folliculogenesis, ovulation/luteinization and luteolysis. Most studies demonstrate that TGFβ members negatively regulate progesterone synthesis, suggesting an involvement in luteolysis. The advance in the knowledge of the function of these local factors may allow the development of new contraceptive strategies as well as new approaches to control the estrous or menstrual cycle.

Partial ablation of endometrial glands in dogs after exposure to progestin during the neonatal period

Bitches with uteri devoid of endometrial glands should be sterile, and consequently could contribute to the population control of dogs. Considering that an inadequate exposure of the female reproductive system to steroids can lead to the formation of the uterine gland knock-out (UGKO) phenotype in some species, the aim of this study was to evaluate the effect of serial applications of medroxyprogesterone acetate (MPA) from birth until the age of six months on the development of endometrial glands in bitches. For this purpose, 16 female mongrel dogs from different litters were distributed into either an MPA group (n = 8), animals treated with 10 mg kg sc (Promone-E®, Pfizer, Brasil) at 3-week intervals, from day one after birth until the age of six months, or a control group (n = 8), composed of animals that only received a 0.9% NaCl solution in place of MPA. At six months of age, ovariohysterectomy was performed and uterine horn samples were collected for histological and immunohistochemical examinations. The bitches from the MPA-treated group presented a 35% decrease in the number of endometrial glands, a larger diameter of the endometrial glands, a greater epithelial height, as well as a greater thickness of the uterine wall, endometrium, and myometrium. However, no significant differences were observed between the two groups in the expression of ER-α, ER-β, and PR on the surface epithelium and endometrial stroma. Therefore, the serial application of MPA from birth until the age of 6 months do not completely ablate the development of the endometrial glands in bitches, but impair it by 35%.

Partial ablation of endometrial glands in dogs after exposure to progestin during the neonatal period

Bitches with uteri devoid of endometrial glands should be sterile, and consequently could contribute to the population control of dogs. Considering that an inadequate exposure of the female reproductive system to steroids can lead to the formation of the uterine gland knock-out (UGKO) phenotype in some species, the aim of this study was to evaluate the effect of serial applications of medroxyprogesterone acetate (MPA) from birth until the age of six months on the development of endometrial glands in bitches. For this purpose, 16 female mongrel dogs from different litters were distributed into either an MPA group (n = 8), animals treated with 10 mg kg sc (Promone-E®, Pfizer, Brasil) at 3-week intervals, from day one after birth until the age of six months, or a control group (n = 8), composed of animals that only received a 0.9% NaCl solution in place of MPA. At six months of age, ovariohysterectomy was performed and uterine horn samples were collected for histological and immunohistochemical examinations. The bitches from the MPA-treated group presented a 35% decrease in the number of endometrial glands, a larger diameter of the endometrial glands, a greater epithelial height, as well as a greater thickness of the uterine wall, endometrium, and myometrium. However, no significant differences were observed between the two groups in the expression of ER-α, ER-β, and PR on the surface epithelium and endometrial stroma. Therefore, the serial application of MPA from birth until the age of 6 months do not completely ablate the development of the endometrial glands in bitches, but impair it by 35%.(AU)

Influência da neoplasia mamária na concentração sérica de hormônios e na expressão de receptores de estrógeno e progesterona em cadelas / Influence of mammary neoplasm on the serum concentration of hormones and on the expression of estrogen and progesterone receptors in bitches

O objetivo desse estudo foi avaliar as concentrações séricas de estradiol, progesterona e prolactina, bem como a expressão gênica dos receptores de estrógeno α e β e de progesterona em cadelas com neoplasias mamárias. Foram utilizadas 60 cadelas adultas, sem raça definida que foram distribuídas em dois grupos. O Grupo I constituído por 30 cadelas portadoras de neoplasias mamárias e o Grupo II constituído por 30 cadelas saudáveis, não portadoras de neoplasia. Para os tutores, foram aplicados questionários sobre fatores epidemiológicos da doença. Após avaliação dos exames pré-operatórios, as cadelas com neoplasia mamária foram submetidas à mastectomia, coletaram-se fragmentos das neoplasias e linfonodos regionais, os quais foram processados para análise histopatológica. Para as dosagens hormonais de estradiol, progesterona e prolactina foram colhidas amostras de sangue em tubos sem anticoagulante e os soros foram submetidos à técnica de eletroquimioluminescência. A expressão gênica dos receptores hormonais foi realizada por meio da técnica de Real-time PCR e para isso foram coletados fragmentos das neoplasias mamárias e extraído o RNA para obtenção do cDNA. A expressão do mRNA para os REα, REβ e RP foi avaliada a partir da amplificação desses genes utilizando primers específicos. Verificaram-se maiores níveis séricos de estradiol (média de 38,98±13,68pg/mL) em cadelas portadoras de neoplasias mamárias malignas quando comparadas as cadelas do grupo controle (p<0,05). Já os níveis séricos de prolactina foram maiores (média de 0,231±0,201ng/mL) nas cadelas que não possuíam neoplasias mamárias quando comparadas ao Grupo I (p<0,05). Para os níveis de progesterona não foram observadas diferença entre os diferentes grupos (p>0,05). Tanto os tumores malignos como os benignos expressaram REα, REβ e RP, não havendo diferença (p>0,05) na expressão entre tumores malignos ou benignos ou relacionada aos outros fatores prognósticos investigados (estadiamento clínico, presença de ulceração, vascularização e tempo de evolução do processo). Os níveis séricos de estradiol aumentaram significativamente com o estadiamento clínico da doença (p<0,05). Verificou-se moderada correlação negativa entre os níveis séricos de estradiol e prolactina. Dessa forma, conclui-se que as dosagens séricas de estradiol e PRL foram influenciadas pela malignidade do tumor e pelo estadiamento clínico das neoplasias. Os receptores hormonais foram expressos pelas neoplasias, independentemente do tipo tumoral e não estão associados aos outros fatores prognóstico clássicos, como presença de ulceração, vascularização ou estadiamento clínico.(AU)

The aim of this study was to evaluate the serum concentrations of estradiol, progesterone, prolactin, the gene expression of estrogen α and β and progesterone receptors in bitches with mammary neoplasms. Sixty adult crossbred bitches distributed in two groups were used. Group I consisted of 30 bitches with mammary neoplasms and Group II consisted of 30 healthy bitches without neoplasia. For the tutors, interviews were made about the disease epidemiology. After preoperative examinations, bitches with mammary neoplasia were submitted to mastectomy; fragments of the neoplasms and regional lymph nodes were collected and processed for histopathological analysis. Blood samples were collected in tubes without anticoagulant and the serum was analyzed by electrochemiluminescence to measure estradiol, progesterone and prolactin. The gene expression of the hormonal receptors was performed by means of the Real-time PCR technique, thus fragments of mammary neoplasms were collected and the RNA was extracted to obtain cDNA. Expression of the mRNA for ERα, ERβ and PR was assessed from the amplification of these genes using specific primers. Higher serum levels of estradiol (mean 38.98±13.68pg/mL) were observed in bitches with malignant neoplasms when compared to the control bitches (p<0.05). Serum prolactin levels were higher (mean of 0.231±0.201ng/mL) in bitches that did not have mammary neoplasms when compared to Group I (p<0.05). No difference was observed for related to the progesterone levels between the groups (p>0.05). Both malignant and benign tumors expressed ERα, ERβ and RP with no statistical difference (p>0.05) and there were no difference related to the other prognostic factors investigated (clinical staging, presence of ulceration, vascularization and aging of neoplasms). Serum estradiol levels increased significantly with the clinical staging of the disease (p<0.05). There was a moderate negative correlation between serum levels of estradiol and prolactin. It was concluded that serum levels of estradiol and PRL were influenced by tumor malignancy and clinical staging of neoplasms. Hormonal receptors were expressed by neoplasms, regardless of tumor type and are not associated with other classical prognostic factors, such as ulceration, vascularization or clinical staging.(AU)

Influência da neoplasia mamária na concentração sérica de hormônios e na expressão de receptores de estrógeno e progesterona em cadelas / Influence of mammary neoplasm on the serum concentration of hormones and on the expression of estrogen and progesterone receptors in bitches

O objetivo desse estudo foi avaliar as concentrações séricas de estradiol, progesterona e prolactina, bem como a expressão gênica dos receptores de estrógeno α e β e de progesterona em cadelas com neoplasias mamárias. Foram utilizadas 60 cadelas adultas, sem raça definida que foram distribuídas em dois grupos. O Grupo I constituído por 30 cadelas portadoras de neoplasias mamárias e o Grupo II constituído por 30 cadelas saudáveis, não portadoras de neoplasia. Para os tutores, foram aplicados questionários sobre fatores epidemiológicos da doença. Após avaliação dos exames pré-operatórios, as cadelas com neoplasia mamária foram submetidas à mastectomia, coletaram-se fragmentos das neoplasias e linfonodos regionais, os quais foram processados para análise histopatológica. Para as dosagens hormonais de estradiol, progesterona e prolactina foram colhidas amostras de sangue em tubos sem anticoagulante e os soros foram submetidos à técnica de eletroquimioluminescência. A expressão gênica dos receptores hormonais foi realizada por meio da técnica de Real-time PCR e para isso foram coletados fragmentos das neoplasias mamárias e extraído o RNA para obtenção do cDNA. A expressão do mRNA para os REα, REβ e RP foi avaliada a partir da amplificação desses genes utilizando primers específicos. Verificaram-se maiores níveis séricos de estradiol (média de 38,98±13,68pg/mL) em cadelas portadoras de neoplasias mamárias malignas quando comparadas as cadelas do grupo controle (p<0,05). Já os níveis séricos de prolactina foram maiores (média de 0,231±0,201ng/mL) nas cadelas que não possuíam neoplasias mamárias quando comparadas ao Grupo I (p<0,05). Para os níveis de progesterona não foram observadas diferença entre os diferentes grupos (p>0,05). Tanto os tumores malignos como os benignos expressaram REα, REβ e RP, não havendo diferença (p>0,05) na expressão...(AU)

The aim of this study was to evaluate the serum concentrations of estradiol, progesterone, prolactin, the gene expression of estrogen α and β and progesterone receptors in bitches with mammary neoplasms. Sixty adult crossbred bitches distributed in two groups were used. Group I consisted of 30 bitches with mammary neoplasms and Group II consisted of 30 healthy bitches without neoplasia. For the tutors, interviews were made about the disease epidemiology. After preoperative examinations, bitches with mammary neoplasia were submitted to mastectomy; fragments of the neoplasms and regional lymph nodes were collected and processed for histopathological analysis. Blood samples were collected in tubes without anticoagulant and the serum was analyzed by electrochemiluminescence to measure estradiol, progesterone and prolactin. The gene expression of the hormonal receptors was performed by means of the Real-time PCR technique, thus fragments of mammary neoplasms were collected and the RNA was extracted to obtain cDNA. Expression of the mRNA for ERα, ERβ and PR was assessed from the amplification of these genes using specific primers. Higher serum levels of estradiol (mean 38.98±13.68pg/mL) were observed in bitches with malignant neoplasms when compared to the control bitches (p<0.05). Serum prolactin levels were higher (mean of 0.231±0.201ng/mL) in bitches that did not have mammary neoplasms when compared to Group I (p<0.05). No difference was observed for related to the progesterone levels between the groups (p>0.05). Both malignant and benign tumors expressed ERα, ERβ and RP with no statistical difference (p>0.05) and there were no difference related to the other prognostic factors investigated (clinical staging, presence of ulceration, vascularization and aging of neoplasms). Serum estradiol levels increased significantly with the clinical staging of the...(AU)

Influência da neoplasia mamária na concentração sérica de hormônios e na expressão de receptores de estrógeno e progesterona em cadelas

ABSTRACT: The aim of this study was to evaluate the serum concentrations of estradiol, progesterone, prolactin, the gene expression of estrogen and and progesterone receptors in bitches with mammary neoplasms. Sixty adult crossbred bitches distributed in two groups were used. Group I consisted of 30 bitches with mammary neoplasms and Group II consisted of 30 healthy bitches without neoplasia. For the tutors, interviews were made about the disease epidemiology. After preoperative examinations, bitches with mammary neoplasia were submitted to mastectomy; fragments of the neoplasms and regional lymph nodes were collected and processed for histopathological analysis. Blood samples were collected in tubes without anticoagulant and the serum was analyzed by electrochemiluminescence to measure estradiol, progesterone and prolactin. The gene expression of the hormonal receptors was performed by means of the Real-time PCR technique, thus fragments of mammary neoplasms were collected and the RNA was extracted to obtain cDNA. Expression of the mRNA for ER, ER and PR was assessed from the amplification of these genes using specific primers. Higher serum levels of estradiol (mean 38.98±13.68pg/mL) were observed in bitches with malignant neoplasms when compared to the control bitches (p 0.05). Serum prolactin levels were higher (mean of 0.231±0.201ng/mL) in bitches that did not have mammary neoplasms when compared to Group I (p 0.05). No difference was observed for related to the progesterone levels between the groups (p>0.05). Both malignant and benign tumors expressed ER, ER and RP with no statistical difference (p>0.05) and there were no difference related to the other prognostic factors investigated (clinical staging, presence of ulceration, vascularization and aging of neoplasms). Serum estradiol levels increased significantly with the clinical staging of the disease (p 0.05). There was a moderate negative correlation between serum levels of estradiol and prolactin. It was concluded that serum levels of estradiol and PRL were influenced by tumor malignancy and clinical staging of neoplasms. Hormonal receptors were expressed by neoplasms, regardless of tumor type and are not associated with other classical prognostic factors, such as ulceration, vascularization or clinical staging.

RESUMO: O objetivo desse estudo foi avaliar as concentrações séricas de estradiol, progesterona e prolactina, bem como a expressão gênica dos receptores de estrógeno e e de progesterona em cadelas com neoplasias mamárias. Foram utilizadas 60 cadelas adultas, sem raça definida que foram distribuídas em dois grupos. O Grupo I constituído por 30 cadelas portadoras de neoplasias mamárias e o Grupo II constituído por 30 cadelas saudáveis, não portadoras de neoplasia. Para os tutores, foram aplicados questionários sobre fatores epidemiológicos da doença. Após avaliação dos exames pré-operatórios, as cadelas com neoplasia mamária foram submetidas à mastectomia, coletaram-se fragmentos das neoplasias e linfonodos regionais, os quais foram processados para análise histopatológica. Para as dosagens hormonais de estradiol, progesterona e prolactina foram colhidas amostras de sangue em tubos sem anticoagulante e os soros foram submetidos à técnica de eletroquimioluminescência. A expressão gênica dos receptores hormonais foi realizada por meio da técnica de Real-time PCR e para isso foram coletados fragmentos das neoplasias mamárias e extraído o RNA para obtenção do cDNA. A expressão do mRNA para os RE, RE e RP foi avaliada a partir da amplificação desses genes utilizando primers específicos. Verificaram-se maiores níveis séricos de estradiol (média de 38,98±13,68pg/mL) em cadelas portadoras de neoplasias mamárias malignas quando comparadas as cadelas do grupo controle (p 0,05). Já os níveis séricos de prolactina foram maiores (média de 0,231±0,201ng/mL) nas cadelas que não possuíam neoplasias mamárias quando comparadas ao Grupo I (p 0,05). Para os níveis de progesterona não foram observadas diferença entre os diferentes grupos (p>0,05). Tanto os tumores malignos como os benignos expressaram RE, RE e RP, não havendo diferença (p>0,05) na expressão entre tumores malignos ou benignos ou relacionada aos outros fatores prognósticos investigados (estadiamento clínico, presença de ulceração, vascularização e tempo de evolução do processo). Os níveis séricos de estradiol aumentaram significativamente com o estadiamento clínico da doença (p 0,05). Verificou-se moderada correlação negativa entre os níveis séricos de estradiol e prolactina. Dessa forma, conclui-se que as dosagens séricas de estradiol e PRL foram influenciadas pela malignidade do tumor e pelo estadiamento clínico das neoplasias. Os receptores hormonais foram expressos pelas neoplasias, independentemente do tipo tumoral e não estão associados aos outros fatores prognóstico clássicos, como presença de ulceração, vascularização ou estadiamento clínico.

Effects of ovarian pathologies and uterine inflammations on adenomyosis in bitches

Background: Adenomyosis is a non-neoplastic condition, characterized by the presence of endometrial glands or cells in the myometrium. Adenomyosis is defined by post-operative microscopic examination of uterine tissue and a rarely seen infertility problem in bitches. The process of transformation of healthy uterus into adenomiotic uterus is still a mystery. We aimed to determine role of ovarian pathologies and uterine inflammations on pathogenesis of uterine adenomyosis in bitches.Cases: After ovariohysterectomy, the uteri were fixed and processed by routine methods. Sections were stained with HematoxylinEosin, Massons trichrome and Periodic-Acid-Schiff and immunohistochemically; with p53 protein, Ki-67, α-smooth-muscle actin, cytokeratin-CK 8, 14, 19 antibodies, estrogen and progesterone receptors by Avidin-Biotin-Complex-Peroxidase method. Histopathologically, 7 of the collected tissue samples (n = 38) were diagnosed as adenomyosis. There were unilateral (n = 6) or bilateral (n = 1) adenomyosis in which endometrial glands had broken through the myometrium, and in some cases there were fibromyoma, inflammation, papillary hyperplasia and cysts in glands. Vaginal fibroma, fibromyoma and benign mix tumor (n = 1), simple carcinoma (n = 3), carcinosarcoma (n = 3) in mammary gland were determinated histopathologically. In addition to these findings, follicular cysts (n = 4), parovarian cyst (n = 1), adenocarcinoma and Rete ovarii adenoma were diagnosed in some ovaries. The CK 19 was scored at endometrial glands (n = 2) positively and also, muscle fibers were stained positively with α-smooth-muscle actin antibody (n = 7).Discussion: Adenomyosis is an infertility problem in adult dogs and is hard to detect with routine gynecological examinations. Adenomyosis can be seen unaccompanied by other gynecological problems such as endometritis or leiomyomas, while mammary tumors with adenomyosis have been previously reported in women and in only one case of a bitch.[...](AU)

Effects of ovarian pathologies and uterine inflammations on adenomyosis in bitches

Background: Adenomyosis is a non-neoplastic condition, characterized by the presence of endometrial glands or cells in the myometrium. Adenomyosis is defined by post-operative microscopic examination of uterine tissue and a rarely seen infertility problem in bitches. The process of transformation of healthy uterus into adenomiotic uterus is still a mystery. We aimed to determine role of ovarian pathologies and uterine inflammations on pathogenesis of uterine adenomyosis in bitches.Cases: After ovariohysterectomy, the uteri were fixed and processed by routine methods. Sections were stained with HematoxylinEosin, Masson’s trichrome and Periodic-Acid-Schiff and immunohistochemically; with p53 protein, Ki-67, α-smooth-muscle actin, cytokeratin-CK 8, 14, 19 antibodies, estrogen and progesterone receptors by Avidin-Biotin-Complex-Peroxidase method. Histopathologically, 7 of the collected tissue samples (n = 38) were diagnosed as adenomyosis. There were unilateral (n = 6) or bilateral (n = 1) adenomyosis in which endometrial glands had broken through the myometrium, and in some cases there were fibromyoma, inflammation, papillary hyperplasia and cysts in glands. Vaginal fibroma, fibromyoma and benign mix tumor (n = 1), simple carcinoma (n = 3), carcinosarcoma (n = 3) in mammary gland were determinated histopathologically. In addition to these findings, follicular cysts (n = 4), parovarian cyst (n = 1), adenocarcinoma and Rete ovarii adenoma were diagnosed in some ovaries. The CK 19 was scored at endometrial glands (n = 2) positively and also, muscle fibers were stained positively with α-smooth-muscle actin antibody (n = 7).Discussion: Adenomyosis is an infertility problem in adult dogs and is hard to detect with routine gynecological examinations. Adenomyosis can be seen unaccompanied by other gynecological problems such as endometritis or leiomyomas, while mammary tumors with adenomyosis have been previously reported in women and in only one case of a bitch.[...]

Estrogen and progesterone receptors immunolabeling in mammary solid carcinoma in an African Hedgehog (Atelerix albiventris) with concurrent uterine fibrosarcoma

The African Hedgehog (Atelerix albiventris) has gained popularity as a pet in several parts of the world. This is a report of a concomitant finding of spontaneous mammary and uterine tumors in an adult female African Hedgehog. The animal presented abdominal masses and died suddenly. Upon necropsy, not only the mammary tumor mass, but also a uterine neoplasm was found. Formalin fixed and paraffin embedded tissues were submitted for histopathological examination. The mammary tumor was diagnosed as a solid carcinoma, and the uterine tumor as fibrosarcoma. By immunohistochemistry, the mammary tumor showed positivity for pancytokeratins and estrogen and progesterone receptors, while the uterine neoplasm exhibited positivity for vimentin. There was high cross-immunoreactivity between anti-human antibodies. The positivity for hormonal receptors antigens may represent a relationship between estrogen and progesterone levels and the development of mammary tumors, as in other mammals.

Estrogen and progesterone receptors immunolabeling in mammary solid carcinoma in an African Hedgehog (Atelerix albiventris) with concurrent uterine fibrosarcoma

The African Hedgehog (Atelerix albiventris) has gained popularity as a pet in several parts of the world. This is a report of a concomitant finding of spontaneous mammary and uterine tumors in an adult female African Hedgehog. The animal presented abdominal masses and died suddenly. Upon necropsy, not only the mammary tumor mass, but also a uterine neoplasm was found. Formalin fixed and paraffin embedded tissues were submitted for histopathological examination. The mammary tumor was diagnosed as a solid carcinoma, and the uterine tumor as fibrosarcoma. By immunohistochemistry, the mammary tumor showed positivity for pancytokeratins and estrogen and progesterone receptors, while the uterine neoplasm exhibited positivity for vimentin. There was high cross-immunoreactivity between anti-human antibodies. The positivity for hormonal receptors antigens may represent a relationship between estrogen and progesterone levels and the development of mammary tumors, as in other mammals.(AU)

Cloprostenol administration in the first week postpartum reduces expression of oxytocin receptors in the endometrium in Holstein-Zebu cows / Administração de cloprostenol na primeira semana pós-parto pode reduzir a expressão de receptores de ocitocina no endométrio de vacas mestiças Holandês-Zebu

The present study investigated the hormonal profile and expression of prostaglandin F2α (PGF2α), oxytocin and estrogen receptors in uterine tissues of postpartum cows treated with cloprostenol. Twenty Holstein-Zebu crossbred cows were treated with saline solution (treatment CONT) or cloprostenol (treatment CLO), both administered two and five days postpartum. Blood samples were collected on days two, seven, 14, 21 and 28 postpartum for progesterone, PGF2α metabolite (PGFM) and estradiol determination, and endometrial biopsy was performed in order to quantify the expression of oxytocin receptor (OXTR), prostaglandin F receptor (PTGFR) and estrogen receptor 1 (ERS1) genes. In the CLO treatment, expression of OXTR was reduced (P<0.05) but no difference (P>0.05) between treatments was found for PTGFR and ERS1 expression. Estrogen concentrations increased progressively until day 14 (P<0.05) and the highest OXTR expression and lowest PTGFR expression were observed on day 14 (P<0.05) in both treatments. Serum PGFM concentrations were high throughout the experiment. In conclusion, cloprostenol administration at days two and five of postpartum seems to reduce OXTR expression in the endometrium in crossbred cows.(AU)

O presente estudo avaliou o perfil hormonal e a expressão gênica de receptores de prostaglandina F2α (PGF2α), ocitocina e estrógeno no endométrio de vacas pós-parto tratadas com cloprostenol. Vinte vacas mestiças Holandês-Zebu foram tratadas com solução salina (tratamento CONT, n = 10) ou cloprostenol (tratamento CLO, n = 10), ambos administrados dois e cinco dias após o parto. Amostras de sangue foram coletadas nos dias dois, sete, 14, 21 e 28 pós-parto para mensuração de progesterona, de metabólito de PGF2α (PGFM) e de estradiol, e foram obtidas biópsias endometriais para quantificar a expressão de PTGFR, OXTR e ESR1. No tratamento CLO, a expressão gênica de receptores de ocitocina foi menor (P<0,05). As concentrações de estrógeno aumentaram progressivamente até o dia 14 (P<0,05). A maior expressão de OXTR foi observada no dia 14 (P<0,05). A expressão de ESR1 foi semelhante entre os tratamentos (P>0,05). Os níveis de PGFM foram altos durante todo o estudo. Conclui-se que a administração de cloprostenol nos dias dois e cinco pós-parto parece diminuir a expressão de OXTR no endométrio em vacas mestiças.(AU)

Cloprostenol administration in the first week postpartum reduces expression of oxytocin receptors in the endometrium in Holstein-Zebu cows / Administração de cloprostenol na primeira semana pós-parto pode reduzir a expressão de receptores de ocitocina no endométrio de vacas mestiças Holandês-Zebu

The present study investigated the hormonal profile and expression of prostaglandin F2α (PGF2α), oxytocin and estrogen receptors in uterine tissues of postpartum cows treated with cloprostenol. Twenty Holstein-Zebu crossbred cows were treated with saline solution (treatment CONT) or cloprostenol (treatment CLO), both administered two and five days postpartum. Blood samples were collected on days two, seven, 14, 21 and 28 postpartum for progesterone, PGF2α metabolite (PGFM) and estradiol determination, and endometrial biopsy was performed in order to quantify the expression of oxytocin receptor (OXTR), prostaglandin F receptor (PTGFR) and estrogen receptor 1 (ERS1) genes. In the CLO treatment, expression of OXTR was reduced (P<0.05) but no difference (P>0.05) between treatments was found for PTGFR and ERS1 expression. Estrogen concentrations increased progressively until day 14 (P<0.05) and the highest OXTR expression and lowest PTGFR expression were observed on day 14 (P<0.05) in both treatments. Serum PGFM concentrations were high throughout the experiment. In conclusion, cloprostenol administration at days two and five of postpartum seems to reduce OXTR expression in the endometrium in crossbred cows.(AU)

O presente estudo avaliou o perfil hormonal e a expressão gênica de receptores de prostaglandina F2α (PGF2α), ocitocina e estrógeno no endométrio de vacas pós-parto tratadas com cloprostenol. Vinte vacas mestiças Holandês-Zebu foram tratadas com solução salina (tratamento CONT, n = 10) ou cloprostenol (tratamento CLO, n = 10), ambos administrados dois e cinco dias após o parto. Amostras de sangue foram coletadas nos dias dois, sete, 14, 21 e 28 pós-parto para mensuração de progesterona, de metabólito de PGF2α (PGFM) e de estradiol, e foram obtidas biópsias endometriais para quantificar a expressão de PTGFR, OXTR e ESR1. No tratamento CLO, a expressão gênica de receptores de ocitocina foi menor (P<0,05). As concentrações de estrógeno aumentaram progressivamente até o dia 14 (P<0,05). A maior expressão de OXTR foi observada no dia 14 (P<0,05). A expressão de ESR1 foi semelhante entre os tratamentos (P>0,05). Os níveis de PGFM foram altos durante todo o estudo. Conclui-se que a administração de cloprostenol nos dias dois e cinco pós-parto parece diminuir a expressão de OXTR no endométrio em vacas mestiças.(AU)

Mammary fibroadenomatoid hyperplasia in a heifer / Hiperplasia fibroadenomatoide mamária em uma novilha

This manuscript described the anatomopathological and immunohistochemical findings in a rare case of mammary fibroadenomatoid hyperplasia in a 12-month-old Holstein heifer. A yellow, multilobulated, firm 20cm x 9cm x 6.5cm mass affecting the right quarters of the udder was observed. Total mastectomy was performed. Microscopic evaluation revealed severe hyperplasia of the mammary epithelium and numerous well-differentiated and mildly pleomorphic acini. Additionally, moderate proliferation of the fibrous connective tissue and the myoepithelial cells near the proliferating acini was evident. About 50% of the proliferating epithelial cells showed positive nuclear labeling for estrogen and progesterone receptors, and approximately one-third were positive for Ki-67. In addition, the myoepithelial cells exhibited diffuse nuclear immunoreactivity for p63. Based on the clinical, morphologic, and immunohistochemical findings, a diagnosis of mammary fibroadenomatoid hyperplasia with probable influence of ovarian steroids was made.

Este artigo descreve os achados anatomopatológicos e imuno-histoquímicos de um caso raro de hiperplasia fibroadenomatoide mamária em uma novilha Holandesa de 12 meses. Observou-se uma massa amarelada, firme, multilobulada, medindo 20 x 9 x 6,5 cm e afetando os quartos direitos do úbere. Mastectomia total foi realizada. Microscopicamente, notou-se hiperplasia acentuada do epitélio mamário e numerosos ácinos levemente pleomórficos e bem diferenciados. Proliferação moderada do tecido conjuntivo fibroso e das células mioepiteliais próximos aos ácinos proliferados era também evidente. Aproximadamente 50% das células epiteliais proliferadas mostraram marcação nuclear positiva para receptores de estrógeno e progesterona e cerca de um terço dessas células era positivo para Ki-67. Além disso, as células mioepiteliais exibiram reação nuclear e difusa para p63. Com base nos achados clínicos, morfológicos e imuno-histoquímicos, firmou-se o diagnóstico de hiperplasia fibroadenomatoide mamária com provável influência de esteroides ovarianos.