Aspectos da biologia reprodutiva de Boa constrictor constrictor: um estudo histológico dos testículos nos períodos reprodutivos de quiescência e máxima atividade / Boa constrictor constrictor reproductive biology aspects: the testes histological study in the reproductive periods of quiescence and maximal activity

As serpentes pertencem ao segundo maior grupo dentro dos répteis, podendo apresentar sazonalidade quanto à espermatogênese, com produção descontínua ou contínua. O presente trabalho tem como objetivo caracterizar aspectos da biologia reprodutiva de Boa constrictor constrictor com base nos achados histológicos dos testículos nos períodos de máxima atividade (período de gametogênese) e quiescência. Os testículos de dois espécimes de Boa c. constrictor (7767 e 11752) foram cortados a uma espessura de 3µm em micrótomo, corados com azul de toluidina 1%, fotodocumentados e descritos. A presença de espermatozoides na luz do túbulo seminífero no indivíduo 7767 indica um período de máxima gametogênese, enquanto o lúmen dos túbulos seminíferos pouco evidentes, sem a presença de espermatozoides e de células gaméticas em divisão, caracteriza o indivíduo 11752 em período quiescente. Mediante os achados histológicos descritos no presente estudo, concluiu-se que Boa c. constrictor apresenta sazonalidade em relação à gametogênese, sendo esse padrão de sazonalidade associado ao período de cópulas relatado em literatura característico de serpentes com padrão pré-nupcial.(AU)

These snakes belong to the second largest group within the reptiles, being able to present seasonality regarding spermatogenesis, with discontinuous or continuous production. The present study aims to characterize Boa constrictor constrictor reproductive biology aspects from histological findings in testicles during periods of maximum activity (period of gametogenesis) and quiescence. The testicles of two specimens of Boa c. constrictor (7767 and 11752) were cut to a thickness of 3µm in microtome, stained with 1% toluidine blue, photodocumented and described. The spermatozoa presence in the seminiferous tubule lumen in individual 7767 indicates a period of maximum gametogenesis, whereas the seminiferous tubules lumen is not very evident without spermatozoa and the absence of dividing gametic cells characterizes individual 11752 in the quiescent period. Through the histological findings we concluded that Boa c. constrictor presents seasonality in relation to gametogenesis, and the pattern of reproductive seasonality observed along with the period of copulas reported in the literature resembles the pre-nuptial pattern.(AU)

Aspectos da biologia reprodutiva de Boa constrictor constrictor: um estudo histológico dos testículos nos períodos reprodutivos de quiescência e máxima atividade / Boa constrictor constrictor reproductive biology aspects: the testes histological study in the reproductive periods of quiescence and maximal activity

As serpentes pertencem ao segundo maior grupo dentro dos répteis, podendo apresentar sazonalidade quanto à espermatogênese, com produção descontínua ou contínua. O presente trabalho tem como objetivo caracterizar aspectos da biologia reprodutiva de Boa constrictor constrictor com base nos achados histológicos dos testículos nos períodos de máxima atividade (período de gametogênese) e quiescência. Os testículos de dois espécimes de Boa c. constrictor (7767 e 11752) foram cortados a uma espessura de 3µm em micrótomo, corados com azul de toluidina 1%, fotodocumentados e descritos. A presença de espermatozoides na luz do túbulo seminífero no indivíduo 7767 indica um período de máxima gametogênese, enquanto o lúmen dos túbulos seminíferos pouco evidentes, sem a presença de espermatozoides e de células gaméticas em divisão, caracteriza o indivíduo 11752 em período quiescente. Mediante os achados histológicos descritos no presente estudo, concluiu-se que Boa c. constrictor apresenta sazonalidade em relação à gametogênese, sendo esse padrão de sazonalidade associado ao período de cópulas relatado em literatura característico de serpentes com padrão pré-nupcial.(AU)

These snakes belong to the second largest group within the reptiles, being able to present seasonality regarding spermatogenesis, with discontinuous or continuous production. The present study aims to characterize Boa constrictor constrictor reproductive biology aspects from histological findings in testicles during periods of maximum activity (period of gametogenesis) and quiescence. The testicles of two specimens of Boa c. constrictor (7767 and 11752) were cut to a thickness of 3µm in microtome, stained with 1% toluidine blue, photodocumented and described. The spermatozoa presence in the seminiferous tubule lumen in individual 7767 indicates a period of maximum gametogenesis, whereas the seminiferous tubules lumen is not very evident without spermatozoa and the absence of dividing gametic cells characterizes individual 11752 in the quiescent period. Through the histological findings we concluded that Boa c. constrictor presents seasonality in relation to gametogenesis, and the pattern of reproductive seasonality observed along with the period of copulas reported in the literature resembles the pre-nuptial pattern.(AU)

Saccharum angustifolium (Nees) Trin. (Poales: Poaceae) as hibernacle for stink bugs during the soybean and corn off-season in Southern Brazil / Saccharum angustifolium (Nees) Trin. (Poales: Poaceae) como hibernáculo para percevejos durante a entressafra de soja e milho no sul do Brasil

Host plants are key factors in the survival of true bugs during unfavorable periods. The objective of this study was to evaluate the composition, diversity and abundance of pentatomid bugs in Saccharum angustifolium (Nees) Trin. (Poales: Poaceae). Plants were evaluated during the soybean and corn off--season for five years. A total of 250 plants were sampled, the true bug population was counted and used for statistical and faunistic analysis. We observed the occurrence of the species: Euschistus heros (F.), Dichelops furcatus (F.), Dichelops melacanthus (Dallas), Edessa meditabunda (F.), and Edessa ruformaginata (De Geer). Among the species, the highest population density was verified for E. meditabunda, E. heros, and D. furcatus. We also observed a direct effect of the clump diameter on the population density of E. heros, D. furcatus, and E. meditabunda. Finally, we conclude that S. angustifolium plants serve as adequate hibernacles for the survival of true bugs of economic importance during the soybean and corn off-season.(AU)

Plantas hospedeiras são fatores fundamentais na sobrevivência de percevejos durante períodos desfavoráveis. O objetivo deste trabalho foi avaliar a composição, diversidade e abundância de espécies de percevejos pentatomídeos ocorrentes em plantas de Saccharum angustifolium (Nees) Trin (Poales: Poaceae). As plantas foram avaliadas durante a entressafra de soja e milho por cinco anos. Foram amostradas 250 plantas, sendo a população de percevejos contabilizada e utilizada para a análise estatística e faunística. Observamos a ocorrência das espécies: Euschistus heros (F.), Dichelops furcatus (F.), Dichelops melacanthus (Dallas), Edessa meditabunda (F.) e Edessa ruformaginata (De Geer). Entre as espécies, foi verificada uma maior densidade populacional para E. meditabunda, E. heros e D. furcatus. Observamos, também, o efeito direto do diâmetro de touceira sobre a densidade populacional de E. heros, D. furcatus e E. meditabunda. Por fim, concluímos que plantas de S. angustifolium servem como hibernáculos adequados para a sobrevivência de percevejos de importância econômica durante a entressafra de soja e milho.(AU)

Saccharum angustifolium (Nees) Trin. (Poales: Poaceae) as hibernacle for stink bugs during the soybean and corn off-season in Southern Brazil / Saccharum angustifolium (Nees) Trin. (Poales: Poaceae) como hibernáculo para percevejos durante a entressafra de soja e milho no sul do Brasil

Host plants are key factors in the survival of true bugs during unfavorable periods. The objective of this study was to evaluate the composition, diversity and abundance of pentatomid bugs in Saccharum angustifolium (Nees) Trin. (Poales: Poaceae). Plants were evaluated during the soybean and corn off--season for five years. A total of 250 plants were sampled, the true bug population was counted and used for statistical and faunistic analysis. We observed the occurrence of the species: Euschistus heros (F.), Dichelops furcatus (F.), Dichelops melacanthus (Dallas), Edessa meditabunda (F.), and Edessa ruformaginata (De Geer). Among the species, the highest population density was verified for E. meditabunda, E. heros, and D. furcatus. We also observed a direct effect of the clump diameter on the population density of E. heros, D. furcatus, and E. meditabunda. Finally, we conclude that S. angustifolium plants serve as adequate hibernacles for the survival of true bugs of economic importance during the soybean and corn off-season.(AU)

Plantas hospedeiras são fatores fundamentais na sobrevivência de percevejos durante períodos desfavoráveis. O objetivo deste trabalho foi avaliar a composição, diversidade e abundância de espécies de percevejos pentatomídeos ocorrentes em plantas de Saccharum angustifolium (Nees) Trin (Poales: Poaceae). As plantas foram avaliadas durante a entressafra de soja e milho por cinco anos. Foram amostradas 250 plantas, sendo a população de percevejos contabilizada e utilizada para a análise estatística e faunística. Observamos a ocorrência das espécies: Euschistus heros (F.), Dichelops furcatus (F.), Dichelops melacanthus (Dallas), Edessa meditabunda (F.) e Edessa ruformaginata (De Geer). Entre as espécies, foi verificada uma maior densidade populacional para E. meditabunda, E. heros e D. furcatus. Observamos, também, o efeito direto do diâmetro de touceira sobre a densidade populacional de E. heros, D. furcatus e E. meditabunda. Por fim, concluímos que plantas de S. angustifolium servem como hibernáculos adequados para a sobrevivência de percevejos de importância econômica durante a entressafra de soja e milho.(AU)

Hormonal and ultrasonographic characterization of the seasonal reproductive cycle of male and female Crotalus durissus terrificus

Research concerning to characterize seasonal reproductive cycle in males and females of Crotalus durissus terrificus by ultrasound and hormonal measurement. Reproductive aspects (follicular and testicular cycles, and pregnancy) from 28 adult snakes (14 males and 14 females) during different months of the years were studied. Snakes housed individually in cages in an environment with controlled luminosity and humidity, and fed monthly. Females were examined by ultrasound during quiescence and active follicular phase, and pregnancy for follicular and embryo/fetal development. Males were evaluated to testicular echotexture and measurements during reproductive and non-reproductive season. The blood samples were collected from males and females for serum testosterone and progesterone determination, respectively. In 77% males the testes were identified by ultrasound and found increased size during summer, decreased serum testosterone in winter, and positive correlation between serum testosterone and testes size. There was no change in testicular echotexture in according to season. Testosterone concentration was decreased during winter and it was positively correlated with testes size. In 71% females, were observed follicular development (vitellogenesis) and gestation since winter to spring by ultrasonography. Parturition occurred mainly in summer. Pregnancy length was 123.0 ± 11.4 days, with mean 6.9 ± 1.5 newborns/female, and there was gradual increase of serum progesterone during this period. There was no variation in progesterone concentration in nongravid females. Males and females Tropical Rattlesnake show seasonal variation of reproductive cycle and was clear a biennial cycle in female. The ultrasonography can be considered an essential tool to accomplish the follicular development, pregnancy and testicular alterations in Tropical Rattlesnake.(AU)

Hormonal and ultrasonographic characterization of the seasonal reproductive cycle of male and female Crotalus durissus terrificus

Research concerning to characterize seasonal reproductive cycle in males and females of Crotalus durissus terrificus by ultrasound and hormonal measurement. Reproductive aspects (follicular and testicular cycles, and pregnancy) from 28 adult snakes (14 males and 14 females) during different months of the years were studied. Snakes housed individually in cages in an environment with controlled luminosity and humidity, and fed monthly. Females were examined by ultrasound during quiescence and active follicular phase, and pregnancy for follicular and embryo/fetal development. Males were evaluated to testicular echotexture and measurements during reproductive and non-reproductive season. The blood samples were collected from males and females for serum testosterone and progesterone determination, respectively. In 77% males the testes were identified by ultrasound and found increased size during summer, decreased serum testosterone in winter, and positive correlation between serum testosterone and testes size. There was no change in testicular echotexture in according to season. Testosterone concentration was decreased during winter and it was positively correlated with testes size. In 71% females, were observed follicular development (vitellogenesis) and gestation since winter to spring by ultrasonography. Parturition occurred mainly in summer. Pregnancy length was 123.0 ± 11.4 days, with mean 6.9 ± 1.5 newborns/female, and there was gradual increase of serum progesterone during this period. There was no variation in progesterone concentration in nongravid females. Males and females Tropical Rattlesnake show seasonal variation of reproductive cycle and was clear a biennial cycle in female. The ultrasonography can be considered an essential tool to accomplish the follicular development, pregnancy and testicular alterations in Tropical Rattlesnake.

Control of growth and development of preantral follicle: insights from in vitro culture

The regulation of folliculogenesis involves a complex interaction among endocrine, paracrine and autocrine factors. The mechanisms involved in the initiation of the growth of the primordial follicle, i.e., follicular activation and the further growth of primary follicles up to the pre-ovulatory stage, are not well understood at this time. The present review focuses on the regulation and development of early stage (primordial, primary, and secondary) folliculogenesis highlighting the mechanisms of primordial follicle activation, growth of primary and secondary follicles and finally transition from secondary to tertiary follicles. We also discuss the importance of in vitro follicle culture for the understanding of folliculogenesis during the preantral phase. Studies suggest that follicular development from primordial to early antral stages is primarily controlled by intra-ovarian ligands but it can also be influenced by many extra-ovarian factors. The control of early folliculogenesis is, therefore, extremely complex because several ligands act through distinct signaling pathways that form sophisticated information networks responding to multiple, often opposing, stimuli. The balance among different stimuli determines follicular survival or death as well as quiescence or activation (growth). The distribution of the ligands and their corresponding receptors varies among follicular compartments and species, and significant changes in gene expression pattern among follicular categories have been reported. Knowing that follicular requirements during early folliculogenesis can be stage-specific and speciesspecific, in vitro culture studies offer an alternative to evaluate single and combined factors during a specific period of follicular development. Herewith we summarize the main findings obtained in vitro together with the mechanisms regulating folliculogenesis.(AU)

Control of growth and development of preantral follicle: insights from in vitro culture

The regulation of folliculogenesis involves a complex interaction among endocrine, paracrine and autocrine factors. The mechanisms involved in the initiation of the growth of the primordial follicle, i.e., follicular activation and the further growth of primary follicles up to the pre-ovulatory stage, are not well understood at this time. The present review focuses on the regulation and development of early stage (primordial, primary, and secondary) folliculogenesis highlighting the mechanisms of primordial follicle activation, growth of primary and secondary follicles and finally transition from secondary to tertiary follicles. We also discuss the importance of in vitro follicle culture for the understanding of folliculogenesis during the preantral phase. Studies suggest that follicular development from primordial to early antral stages is primarily controlled by intra-ovarian ligands but it can also be influenced by many extra-ovarian factors. The control of early folliculogenesis is, therefore, extremely complex because several ligands act through distinct signaling pathways that form sophisticated information networks responding to multiple, often opposing, stimuli. The balance among different stimuli determines follicular survival or death as well as quiescence or activation (growth). The distribution of the ligands and their corresponding receptors varies among follicular compartments and species, and significant changes in gene expression pattern among follicular categories have been reported. Knowing that follicular requirements during early folliculogenesis can be stage-specific and speciesspecific, in vitro culture studies offer an alternative to evaluate single and combined factors during a specific period of follicular development. Herewith we summarize the main findings obtained in vitro together with the mechanisms regulating folliculogenesis.

Na/K-ATPase and Regulation of Sperm Function

A standard bull breeding soundness evaluation (BBSE) identifies bulls with semen that is grossly abnormal. Nonetheless, semen samples classified as satisfactory based on these traditional approaches differ in fertility; perhaps there are submicroscopic differences in sperm characteristics affecting fertility. Therefore, a better understanding of molecular regulation of sperm function could promote development of novel, evidence-based approaches to predict male fertility. Recently the α4 isoform of Na/K-ATPase (ATP1A4) has received considerable attention, due to its testisspecific expression in post-meiotic germ cells and mature sperm, in addition to its regulation of sperm motility and capacitation. Using fresh bull sperm, we determined that ATP1A4 resided in specialized microdomains (raft and non-raft) of the sperm plasma membrane and activated specific signaling (caveolin-1, EGFR, Src, ERK1/2) molecules during sperm capacitation. Furthermore, ATP1A4 was the predominant isoform responsible for total Na/K-ATPase activity in capacitated sperm. Despite the widely accepted dogma of transcriptional/translational quiescence, bovine sperm translated ATP1A4 mRNA on mitochondrial or mitochondrial-type ribosomes, increasing their content and activity during capacitation. Proteomic analysis of raft and non-raft fractions revealed a significant interaction between ATP1A4 and plakoglobin, a member of the β-catenin family of proteins involved in cell adhesion, in the equatorial segment of capacitated sperm, suggesting a potential role in sperm-oolemma fusion. In frozen-thawed sperm, ATP1A4 content and activity was greater in high- versus low-fertility bulls. Additionally, ATP1A4-induced increases in ROS, calcium, actin polymerization and tyrosine phosphorylation were also involved in regulating post-thaw sperm function in these bulls. Overall, results demonstrated that ATP1A4 had unique roles in controlling several aspects of sperm physiology, acting through well-established enzyme activity and signaling functions. Consequently, isoforms of Na/K-ATPase are potential biomarkers for male fertility.

Na/K-ATPase and Regulation of Sperm Function

A standard bull breeding soundness evaluation (BBSE) identifies bulls with semen that is grossly abnormal. Nonetheless, semen samples classified as satisfactory based on these traditional approaches differ in fertility; perhaps there are submicroscopic differences in sperm characteristics affecting fertility. Therefore, a better understanding of molecular regulation of sperm function could promote development of novel, evidence-based approaches to predict male fertility. Recently the α4 isoform of Na/K-ATPase (ATP1A4) has received considerable attention, due to its testisspecific expression in post-meiotic germ cells and mature sperm, in addition to its regulation of sperm motility and capacitation. Using fresh bull sperm, we determined that ATP1A4 resided in specialized microdomains (raft and non-raft) of the sperm plasma membrane and activated specific signaling (caveolin-1, EGFR, Src, ERK1/2) molecules during sperm capacitation. Furthermore, ATP1A4 was the predominant isoform responsible for total Na/K-ATPase activity in capacitated sperm. Despite the widely accepted dogma of transcriptional/translational quiescence, bovine sperm translated ATP1A4 mRNA on mitochondrial or mitochondrial-type ribosomes, increasing their content and activity during capacitation. Proteomic analysis of raft and non-raft fractions revealed a significant interaction between ATP1A4 and plakoglobin, a member of the β-catenin family of proteins involved in cell adhesion, in the equatorial segment of capacitated sperm, suggesting a potential role in sperm-oolemma fusion. In frozen-thawed sperm, ATP1A4 content and activity was greater in high- versus low-fertility bulls. Additionally, ATP1A4-induced increases in ROS, calcium, actin polymerization and tyrosine phosphorylation were also involved in regulating post-thaw sperm function in these bulls. Overall, results demonstrated that ATP1A4 had unique roles in controlling several aspects of sperm physiology, acting through well-established enzyme activity and signaling functions. Consequently, isoforms of Na/K-ATPase are potential biomarkers for male fertility.(AU)

Envolvimento de uma glicogênio sintase na morfogênese em embriões de Aedes aegypti

A principal forma de combate ao mosquito Aedes aegypti é o controle populacional deste vetor. Entretanto, o uso indiscriminado de inseticidas tem levado o aumento de casos de resistência, este fenômeno levou a busca de novos conhecimentos para o desenvolvimento de fármacos e estratégias de controle populacionais tornam-se necessários para isso novos estudos que auxiliem na maior compreensão da fisiologia e do metabolismo deste inseto. Pela sua importância e dinâmica de eventos que ocorrem nesta etapa, os ovos desta espécie apresentam particularidades físico-químicas que permitem desde a resistência ao ressecamento até a quiescência. Neste trabalho buscou-se mostrar correlações existentes entre o metabolismo energético e os processos de morfogênese do mosquito Aedes aegypti, através do silenciamento gênico de uma enzima descrita como exclusiva do metabolismo de carboidratos, a Glicogênio Sintase (GS). Para isso foi estabelecido um protocolo de permeabilização de ovos para utilização de dupla fita de RNA para silenciamento gênico e avaliação de parâmetros como esclerotização dos ovos, taxa de eclosão e morfologia dos embriões. Para a permeabilização dos ovos para silenciamento foi utilizada uma solução de clorito de sódio 1%, sendo verificada a permeabilização destes ovos através de marcadores como o Laranja de Acridina, que foi capaz de promover a marcação em ovos e larvas após ao tratamento com clorito. Verificamos também uma taxa de silenciamento próxima de 60%, além disso, nos embriões silenciados foi observado reduções nas taxas de esclerotização dos ovos e eclosão das larvas. Quando foi analisado o teor de glicogênio presente também se observou uma redução de 60% do glicogênio presente nas larvas. Nas análises morfológicas foi observada falta de compartimentalização do embrião, ausência de produção de vitelo e falta de polarização do embrião. Utilizando um protocolo de microinjeção para silenciamento gênico nas fêmeas adultas observou-se fenômenos morfológicos semelhantes. A partir destes dados mostram que o metabolismo de carboidratos atue além de uma importante fonte de energia, como um importante fator para a organogênese e desenvolvimento embrionário do mosquito Aedes aegypti.

The main form of Aedes aegypti mosquito control is the population control of this vector. However, the indiscriminate use of insecticides has led to increased cases of resistance, this phenomenon led to the search for new knowledge for the development of drugs and population control strategies become necessary for this new studies that help in the better understanding of physiology and metabolism of this insect. Because of their importance and the dynamics of events that occur in this stage, the eggs of this species present physiology and chemical peculiarities that allow from resistance to dryness to quiescence. This work aimed to show correlations between energy metabolism and the morphogenesis processes of the Aedes aegypti mosquito, through the gene silencing of an enzyme described as exclusive to carbohydrate metabolism, Glycogen Synthase (GS). For this, an egg permeability protocol was established for the use of double strand of RNA for gene silencing and evaluation of parameters such as esclerotization of eggs, hatching rate and embryo morphology. For the permeabilization of the eggs for silencing, a solution of 1% sodium chlorite was used, and the permeabilization of these eggs was verified through markers such as Acridine Orange, which was able to promote labeling in eggs and larvae after treatment with chlorite. We also observed a silencing rate close to 60%, in addition, in the silenced embryos we observed reductions in the rates of esclerotization of eggs and hatching of the larvae. When the present glycogen content was analyzed, a 60% reduction of the glycogen present in the larvae was observed. The morphological analysis showed a lack of compartmentalization of the embryo, absence of calf production and lack of embryo polarization. Using a microinjection protocol for gene silencing in adult females, similar morphological phenomena were observed. From these data show that carbohydrate metabolism acts in addition to an important source of energy, as an important factor for the organogenesis and embryonic development of the Aedes aegypti mosquito.

Sazonalidade reprodutiva em machos de tartaruga (Kinosternon scorpioides) de vida livre no Nordeste brasileiro evidenciado por imunolocalização de enzimas esteroidogênicas no testículo e epidídimo

No Estado do Maranhão, na região da Baixada Maranhense, presenta na fauna silvestre o réptil Kinosternon scorpioides, um quelônio de água doce popularmente conhecido como jurará e que possui valor social, econômico e ambiental para os ribeirinhos da cidade de São Bento. Este estudo contempla suas características biológicas reprodutivas baseadas em seu ambiente natural, com o intuito de permitir a preservação e o estabelecimento de planos de manejo reprodutivo e de uso sustentável da espécie. Recentemente poucos estudos em todo o mundo tratam sobre os aspectos do ciclo reprodutivo de tartarugas em face das características endócrinas, na América do Sul estudos desse tipo são recentes e escassos, sendo assim este é o primeiro estudo, que se tem conhecimento, que elucida um padrão sazonal reprodutivo da espécie K. scorpioides, associando hormônios gonadais com aspectos comportamentais. Trinta e oito animais adultos tiveram seus órgãos reprodutivos caracterizados para as enzimas esteroidogênicas P450 aromatase, P450c17 e PNADPH redutase através de imunomarcação e blotting, além de índice gonadossomático, morfometria e concentração de testosterona, corticosterona e estradiol pela técnica de radioimunoensaio. As mudanças biométricas, morfometria celular e a esteroidogênese testicular entre os períodos chuvoso e seco sugerem que o estrógeno produzido pelas células de leydig podem afetar a produção e a apoptose de células germinais durante o processo de espermatogênese, e a presença das enzimas P450aromatase e P450c17 em células de leydig acompanhados com a recrudescência testicular também reforça a ideia, de que elas podem desempenhar um papel na quiescência testicular. Esse trabalho evidenciou que as enzimas citocromo P450, citocromo P450c17 e NADPH-citocromo P450- redutase estão presentes no testículo e epidídimo nos diferentes períodos climáticos e podem estar relacionados à síntese de testosterona em tartarugas concordando com os diferentes achados para biometria e espermiogênese nos períodos chuvoso e seco, o que, nos leva a sugerir um estado de quiescência durante o período seco e atividade espermatogênica no período chuvoso, semelhante ao que ocorre com as espécies que apresentam comportamento reprodutivo sazonal.

In the state of Maranhão, in the lowland region, the reptile Kinosternon scorpioides, a freshwater chelonian, popularly known as swear has social, economic and environmental value to the riparian of the city of São Bento. This study includes reproductive biological characteristics based on their natural environment in order to enable the preservation and the establishment of reproductive management plans and sustainable use of the species. To date, few studies around the world address the aspects of the reproductive cycle of the turtles in the face of the endocrine features. In South America, these studies are recent and scarce. So this is the first study that elucidates a reproductive seasonal pattern of the species K. scorpioides, by associating gonadal hormones with behavioral aspects. Thirty-eight adult animals had their reproductive organs characterized for P450 steroidogenic enzyme aromatase, P450c17 and NADPH reductase by immunostaining and immunoblotting, and gonadossomatic index, morphology and concentration of testosterone, corticosterone and estradiol by radioimmunoassay. Biometric changes, cell morphology and testicular steroidogenesis between the rainy and dry seasons suggests that the estrogen produced by the leydig cells can affect the production and apoptosis of germ cells during spermatogenesis process; and the presence of P450aromatase and P450c17 enzymes in the leydig cells accompanied with testicular recrudescence also reinforces the idea that they may play a role in testicular quiescence. This work showed that the cytochrome P450 enzymes, P450c17 and NADPH-cytochrome P450 reductase are present in the testis and epididymis at different climatic periods and may be related to testosterone synthesis in turtles agreeing to the different findings for biometrics and spermiogenesis in and dry rainy periods. These findings suggest a state of quiescence during the dry season and spermatogenic activity during the rainy season, similar to those found in species with seasonal reproductive behavior.

Variações sazonais na ultra-estrutura do ducto deferente de codorna doméstica (Coturnix coturnix) / Seasonal variability on the ultrastructure of the vas deferens of domestic quail (Coturnix coturnix)

O ducto deferente (DD) de codorna doméstica mostrou variações sazonais na ultra-estrutura do epitélio de revestimento tubular nas quatro estações do ano, sendo, contudo, mais marcantes no outono, que corresponde à fase quiescente, e na primavera, que é a fase ativa do ciclo testicular anual. Durante a quiescência outonal ocorreram redução do calibre e da luz tubular do DD e características de degeneração celular das células principais (P) do epitélio do DD. A fase ativa primaveril mostrou o calibre tubular e a dimensão luminal maiores do DD, ocorrendo grande estoque de material intra-luminal formado principalmente por espermatozoides. A ultra-estrutura do epitélio de revestimento do DD na fase ativa primaveril mostrou variabilidade das organelas citoplasmáticas relacionadas, principalmente, aos processos de síntese proteica e de endocitose. Contudo, foi observada uma maior quantidade de lisossomos no citoplasma apical de células P do DD na fase recrudescente do inverno, assim como uma escassez relativa de organelas citoplasmáticas e presença de células escuras na fase regressiva do verão.(AU)

The vas deferens (VD) of domestic quail showed seasonal variability on the fine structure of the lining tubular epithelium during the four seasons along the year, whose features were marked more relevant in the autumn and spring which respectively corresponded to the quiescent and active phases of the annual testis cycle ("cycle") in this species. During the fall quiescence had been verified tubular reduction of the VD caliber and luminal dimension followed by degenerative features of the principal epithelial cells and lack of luminal content of spermatozoa. Counter wise in the spring active phase of the "cycle" great dimension of the VD caliber and luminal dimension were observed and the luminal content was plenum of material represented prior by spermatozoa. Similar features occurred on the VD fine structure in the summer and winter according to our results. Fine structural features of the VD lining epithelium in the spring showed variability of cytoplasm organelle mainly related to active processes of endocytosis and protein synthesis, although more lysosomal activity was seen on the apical cytoplasm of principal epithelial cells in the recrudescent phase of winter. Moreover, it was verified a relative scarcity of cytoplasm organelle followed by presence of dark epithelial cells in the regressive phase of summer.(AU)

Gene expression during early follicular development and mechanisms of activation and growth of primordial follicles in mammals

Background: For decades, the mechanisms maintaining the dormancy, survival and growth of mammalian primordial follicles as well as their growth up to an early antral stage have been not well understood. In recent years, data obtained from studies on the expression and quantification of mRNA for several growth factors and from studies with genetically modified mouse models have revealed a number of molecules, whose functions are indispensable for (i) the maintenance of follicular quiescence, (ii) primordial follicle survival and (iii) activation, and/or (iv) the growth of primary follicles up to an early antral stage. Review: This review focuses on expression of mRNA and protein for growth factors, cytokines and their respective receptors in early follicles, as well as on the intrafollicular signaling cascades that lead to the in-vitro activation of primordial follicles. Furthermore, fundamental information on the levels of mRNA for growth factors at different stages of follicular development and the in-vitro effects of several locally expressed factors on ovarian follicular development will be discussed. During the transition into the primary stage or from the primary into the secondary follicle stage of goats there is a significant increase in the mRNA expression of different factors (BMP-6, BMP-15, KL, EGF, VIP, GDF-9). The detection of these different factors depends on the species. In rat ovaries, GHR is detected in oocytes, granulosa and theca cells. The presence of the mRNA for GHR, but not that for GH has been detected in rat pre-antral follicles. GHR mRNA has only been found in granulosa cells, while positive immunostaining for GHR has been observed in both oocytes and granulosa cells. Also in vitro studies with goat primordial follicles have demonstrated that different factors, estradiol and progesterone promote primordial follicle activation and/or oocyte growth. Recently, several studies have been performed with mouse primordial follicles to understand how intrafollicular cytokines and growth factors may control the fate of primordial follicles in the ovary. In rodents, anti-Mullerian hormone has been shown to inhibit mouse primordial follicle growth and to downregulate c-kit expression in rats, suggesting that, at least in rodents, the kit system plays a key role in initiation of follicular growth. In relation to control of primary and secondary follicles, several in vitro studies have demonstrated that FSH, activin-A, EGF, GH, IGF-I and IGF-II stimulate oocyte growth and follicular development in different species. Conclusion: This review updates the information on expression of mRNA and proteins for growth factors and their role in the regulation of development from primordial to early antral follicles. The growing knowledge of the molecules that control the dormancy, survival, activation and growth of primordial follicles will not only contribute for a better understanding of the physiology of the mammalian ovary, but will also enable researchers to develop more promising methods for promoting growth of oocytes from primordial follicles, the richest follicular resource in the mammalian ovary.

EFEITO DE AGENTES CRIOPROTETORES E DE RESFRIAMENTO EM EMBRIÕES DE MACROBRACHIUM AMAZONICUM

As técnicas de criopreservação vêm sendo amplamente utilizadas visando a preservação de tecidos e células em quiescência para o retorno ao metabolismo normal. Existem poucos relatos de criopreservação em embriões de Macrobrachium amazonicum, assim como as injúrias que este processo pode causar durante o desenvolvimento embrionário. Diante disso, o objetivo deste trabalho foi analisar os efeitos dos agentes crioprotetores e do resfriamento no estádio VIII desta espécie. Fêmeas ovígeras tiveram seus ovos e embriões retirados da câmara incubadora e pesados (0,01 g), em seguida foramcolocados em tubos Falcon® com solução crioprotetora e solução salina livre de cálcio.Os ovos e embriões foram levados para caixas térmicas e resfriados a5 °C por duas horas, ao término, foram lavados em água corrente e transportados para incubadoras, onde permaneceram por 24 horas.Uma alíquota foi fixada com formol 4% e, em seguida passou pelo processo de histologia clássica, com cortes de3 m de espessura e analisada em microscópio Leica DM 2000. Outra alíquota foi utilizada para a avalição biométrica, em que comprimento, largura e volume foram medidos e analisados em microscópio estereoscópico Leica DFC 295,utilizando o programa LAS V3.6. Os agentes crioprotetores utilizados foram: dimetilsulfóxido (1, 5 e 10 %), álcool metílico (1, 5 e 10 %), etilenoglicol (1, 5 e 10 %) e sacarose (0,5 M). Foi realizada análise de variância seguida do teste Tukey, com nível de significância de 5%. O dimetilsulfóxido (DMSO) foi o crioprotetor que menos afetou a sobrevivência dos embriões com taxas de 71,8%, 36,2%e 0% para as concentrações de 1%, 5% e 10%, respectivamente. O etilenoglicol causou 100% de mortalidade em todas as concentrações utilizadas. No tratamento com DMSO 10% foram observadas as menores medidas para comprimento e volume, comparado com as outras concentrações de DMSO utilizadas. Os ovos resfriados em álcool metílico 10% tiveram as menores medidas para comprimento, largura e volume.Não foi observada diferença estatística significativa no comprimento dos ovos resfriados nas três concentrações de etilenoglicol. Não foi possível observar nesse trabalho a interferência do resfriamento e dos crioprotetores utilizados sobre as estruturas embrionárias

The cryopreservation techniques have been widely used with the intention of preserving tissues and cells in quiescence for the return to normal metabolism. There are few reports of cryopreservation Macrobrachium amazonicum embryos, as well as injuries that this process can cause during embryonic development. Thus, the aim of this study was to analyze the effects of cryoprotectants agents and cooling in stage VIII of this species. Ovigerous females eggs andembryos were removed from the incubator chamber and weighed (0.01g), then were placed in Falcon® tubes with cryoprotectant solution and saline-free calcium solution. Eggs and embryos were in ice coolers and cooled to 5 °C then were stored for a period of two hours, at the end, were washed with tap water to remove the ACP and acclimated for 5 minutes in water at room temperature and then transferred to 50 ml incubators, where they remained for 24 hours. An aliquot was fixed with 4% formaldehyde and then passed through the histology procedure with 3 mm thick sections, stainded with hematoxylin/eosin and analyzed in a Leica DM 2000. Another aliquot was used for biometric analyse, where length, width and volume were measured and analyzed in stereoscopic microscope Leica DFC 295, using the LAS program V3.6. The cryoprotectants agents used were: dimethylsulfoxide (1, 5 and 10%), methyl alcohol (1, 5 and 10%), ethylene glycol (1, 5 and 10%) and sucrose (0.5 M). Variance analysis was used, followed by the Tukey test at 5% significance level. The dimethylsulfoxide (DMSO) cryoprotector was affected less embryo survival with rates of 71.8%, 36.2% and 0% for concentrations of 1%, 5% and 10%, respectively. The ethylene caused 100% mortality at all the concentrations used. Treatment with 10% DMSO were observed minor measures to length and volume, compared with other DMSO concentrations used. Colds eggs to methyl alcohol 10% had lower measures for length, width and volume. There was no statistically significant difference in the length of colds eggs in the three ethylene glycol concentrations. It was not possible to observe this work interference cooling and cryoprotectants used on embryonic structures

AÇÃO DE FÁRMACOS NAS CONTRAÇÕES DA MUSCULATURA DA CÉRVIX DE SUÍNOS

A manipulação farmacológica da cérvix possui potencial importante na assistência à biotécnicas da reprodução, como inseminação artificial e transferência de embriões, em espécies que o acesso transcervical ao útero é muito difícil e, portanto, necessitam de procedimento cirúrgico para aumentar as taxas de concepção. Por meio de estudos básicos das propriedades mecânicas e farmacológicas da cérvix, objetivou-se com este estudo fornecer maior conhecimento da atividade da musculatura lisa da cérvix e então apresentar e discutir possibilidades de fármacos que possam relaxá-la e, portanto facilitar a passagem transcervical. Segmentos do músculo liso circular (MC) e longitudinal (ML) foram dissecados do útero (corno e corpo) e cérvix de suínos e preparados para tensão isométrica em banhos de órgão isolados de 20mL de solução modificada de Krebs-Henseleit (pH 7,4) gaseificada com 95% O2: 5% CO2 a 37°C. Os segmentos da cérvix apresentaram maiores respostas mecânicas do que os uterinos, mas exibiram menor atividade espontânea. Todas as preparações responderam de maneira dependente à frequência de estimulação de força elétrica (EFE, 2Hz 32Hz, 300mA, por 30s). Especialmente na cérvix, após as respostas neurogênicas, houve um período de quiescência muscular importante até que as atividades espontâneas retornassem ao normal. Na maioria das preparações cervicais (MC e ML) a atropina aboliu as contrações neurogênicas, destacando um maior envolvimento para inervação colinérgica e receptoresmuscarínicos, mas uma pequena proporção das preparações se mostraram resistente à atropina. Estas respostas não foram afetadas pela fentolamina, e nem pela prazosina e fenilefrina, sugerindo a ausência de receptores alfa-adrenérgicos. L-NAME (N-nitro-L-arginina-metil-ester) não teve nenhum efeito sobre a atividade muscular cervical, excluindo o envolvimento de óxido nítrico. Diltiazem, paraverina e isoprenalina inibiram as contrações neurogênicas dos MC e ML da cérvix, consistente com importante envolvimento dos canais de cálcio sensíveis à voltagem, fosfodiesterase e receptores beta-adrenérgicos, respectivamente. Entretanto, RP73401 (3-cyclopentyloxy-N-(3,5-dichloro-4-pyridl)-4-methoxybenzamine) e UK114542 (UK114542 - 5-[2-ethoxy-5-(morpholinylacetyl)phenyl]-1,6-dihydro-1-methyl-3-propyl-7H-pyrazolo[4,3-d]pyrimidin-7-one methanesulfonate monohydrate) não tiveram efeito nenhum sobre a musculatura lisa cervical. Exposição prolongada à fenoxibenzamina reduziu as respostas neurogênicas e atividade espontânea da cérvix. Ainda, estes efeitos se mostraram irreversíveis e consistentes com a ação conhecida desta droga em ambos receptores muscarínicos e canais de cálcio, além de ser antagonista de receptores alfa-adrenérgicos. Esta ação irreversível da fenoxibenzamina pode ser particularmente útil para estudos posteriores no auxílio à penetração cervical e aumento das taxas de concepção.

Pharmacological manipulation of cervix has a potentially important role for aiding either artificial insemination or embryo transfer in species requiring surgical procedures to aid conception. By studying basic pharmacological and mechanical properties of the cervix, we aimed with this study to provide a better knowledge about cervical smooth muscle activity and then present and discuss insights into possible drugs for the pig that could facilitate biotechnological procedures for reproduction. Circular (CM) and longitudinal (LM) smooth muscle strips were dissected from porcine uterus (cornu and corpus) and cervix and prepared for isometric tension recording in organ-bath of 20mL of modified Krebs Henseleit solution (pH 7.4) maintained at 37oC and gassed with 95% O2 and 5% CO2. Cervical strips produced larger mechanical responses than the uterine segments, but exhibited less spontaneous activity. All preparations responded in a frequency-dependent manner to electrical field stimulation (EFS, 2Hz-32Hz, 300mA for 30s) with a contraction. Especially in cervix, after neurogenic responses, there was a period of important muscular quiescence until the resume of normal spontaneous activity. In most cervical preparations (CM and LM) atropine abolished neurogenic contractions, highlighting a major role for cholinergic nerves and muscarinic receptors, but a small proportion of preparations exhibited atropine-resistant responses. These responses were not affected by phentolamine, and neither by prazosina and phenylephrine suggesting no role for alpha-adrenoceptors. L-NAME (N-nitro-L-arginina-metil-ester) did not affect the cervical smooth muscle activity, excluding the role of nitric oxide. Diltiazem, papaverine and isoprenaline inhibited neurogenic contractions in CM and LM strips of the cervix, consistent with important roles for voltage-sensitive calcium channels, phosphodiesterase and beta-adrenoceptors, respectively. However, RP73401 (3-cyclopentyloxy-N-(3,5-dichloro-4-pyridl)-4-methoxybenzamine) and UK114542 (UK114542 - 5-[2-ethoxy-5-(morpholinylacetyl)phenyl]-1,6-dihydro-1-methyl-3-propyl-7H-pyrazolo[4,3-d]pyrimidin-7-one methanesulfonate monohydrate) did not present effect in cervical smooth muscle. Prolonged exposure to phenoxybenzamine reduced cervical neurogenic responses and spontaneous activity. Interestingly, these effects were irreversible and consistent with known action of this drug at both muscarinic receptors and calcium channels, besides being alfa-adrenoceptor antagonist. This atypical, irreversible, action of the phenoxybenzamine may be particularly useful for further studies in aiding cervical penetration and increasing conception rates.

Variação sazonal na morfologia dos ductos eferentes distais em codorna doméstica mantida em cativeiro experimental

Some expressive differences were noted on the distal efferent ducts (DED) morphology in domestic quail of the Italian variety along the year. The birds were maintained on experimental captive breeding along two consecutive years, before the morphologic studies. Morphologic differences on the DED had been more evident in spring and autumn respectively, the active and quiescent phases of the annual testis cycle. Variability more expressive was noted among the principal (P) and ciliated (C) epithelial cells and in tubular diameters of DED, during the two focused seasons. Spring features of DED were marked by relative enlargement of the passageways, with similar results in summer and winter, and characteristic organelle development on the apical cytoplasm of P cells, allowing to conclude presence of active process of endocytosis. Autumn quiescence was marked by an enfolding pattern of all the tubular passageways, lacking of spermatozoa and small amount of heterogeneous material inside their luminal compartments, followed by presence of ultrastructural degenerative features in apical cytoplasm of P cells and C cells in DED passageways. Tubular features verified in DED during winter and summer were minor and similar to those described in spring. Summer and winter were respectively considered the recrudescent and regressive phases of the testis cycle in these species, based on the spermatogenesis cycle analysed in all the seasons of the year as well as on the base the morphologic features here found.

Neste estudo diferenças expressivas foram observadas na estrutura dos ductos eferentes distais (DED) de codorna doméstica Coturnix coturnix, da variedade italiana, mantida em cativeiro experimental durante dois anos, durante as quatro estações do ano. A variabilidade morfológica dos DED foi mais marcante na primavera e no outono, eqüivalendo às fases ativa e quiescente do ciclo reprodutivo anual (ciclo testicular) nesta ave. Nessas fases do ciclo, diferenças significativas foram observadas na ultraestrutura das células principais (P) e ciliadas (C) do epitélio de revestimento tubular e nos calibres tubulares. Estes tiveram valores médios maiores na primavera, com valores similares no verão e no inverno, e valores médios menores observados no outono. Na primavera, no citoplasma apical de células P, notou-se maior complexidade organelar, inferindo-se a ocorrência de processos endocitóticos ativos. A quiescência outonal caracterizou-se por redução do calibre tubular dos DED, luz tubular vazia de espermatozóides, degenerações de organelas citoplasmáticas e "debridamentos" citoplasmáticos apicais em células P e C. No inverno e verão, correspondentes às fases recrudescente e regressiva, respectivamente, do ciclo testicular nesta espécie, os dados obtidos foram, de modo geral, similares aos observados na primavera.

Variação sazonal na morfologia dos ductos eferentes distais em codorna doméstica mantida em cativeiro experimental

Some expressive differences were noted on the distal efferent ducts (DED) morphology in domestic quail of the Italian variety along the year. The birds were maintained on experimental captive breeding along two consecutive years, before the morphologic studies. Morphologic differences on the DED had been more evident in spring and autumn respectively, the active and quiescent phases of the annual testis cycle. Variability more expressive was noted among the principal (P) and ciliated (C) epithelial cells and in tubular diameters of DED, during the two focused seasons. Spring features of DED were marked by relative enlargement of the passageways, with similar results in summer and winter, and characteristic organelle development on the apical cytoplasm of P cells, allowing to conclude presence of active process of endocytosis. Autumn quiescence was marked by an enfolding pattern of all the tubular passageways, lacking of spermatozoa and small amount of heterogeneous material inside their luminal compartments, followed by presence of ultrastructural degenerative features in apical cytoplasm of P cells and C cells in DED passageways. Tubular features verified in DED during winter and summer were minor and similar to those described in spring. Summer and winter were respectively considered the recrudescent and regressive phases of the testis cycle in these species, based on the spermatogenesis cycle analysed in all the seasons of the year as well as on the base the morphologic features here found.

Neste estudo diferenças expressivas foram observadas na estrutura dos ductos eferentes distais (DED) de codorna doméstica Coturnix coturnix, da variedade italiana, mantida em cativeiro experimental durante dois anos, durante as quatro estações do ano. A variabilidade morfológica dos DED foi mais marcante na primavera e no outono, eqüivalendo às fases ativa e quiescente do ciclo reprodutivo anual (ciclo testicular) nesta ave. Nessas fases do ciclo, diferenças significativas foram observadas na ultraestrutura das células principais (P) e ciliadas (C) do epitélio de revestimento tubular e nos calibres tubulares. Estes tiveram valores médios maiores na primavera, com valores similares no verão e no inverno, e valores médios menores observados no outono. Na primavera, no citoplasma apical de células P, notou-se maior complexidade organelar, inferindo-se a ocorrência de processos endocitóticos ativos. A quiescência outonal caracterizou-se por redução do calibre tubular dos DED, luz tubular vazia de espermatozóides, degenerações de organelas citoplasmáticas e "debridamentos" citoplasmáticos apicais em células P e C. No inverno e verão, correspondentes às fases recrudescente e regressiva, respectivamente, do ciclo testicular nesta espécie, os dados obtidos foram, de modo geral, similares aos observados na primavera.

Variabilidade sazonal no ducto epididimário de codorna doméstica: observações morfológicas / Seasonal variability in epididymal duct of the domestic quail: morphologic features

O ducto epididimário (DE) de codorna doméstica mostrou, ao longo do ano, variabilidade pequena, porém muito expressiva no outono, o qual corresponde à fase quiescente do ciclo testicular anual. A morfologia do DE na primavera foi, em termos, similar à verificada no verão e inverno. Nestas fases notaram-se aumento significante do calibre tubular do DE; estocagem intraluminal de espermatozóides e ocorrência de mitocôndrias, lamelas do RE, vesículas variáveis quanto à forma, dimensões e conteúdos e presença de alguns lisossomos localizados, principalmente, no citoplasma apical das células principais (P), no epitélio epididimário. Estas características ultra-estruturais das células P parecem ser indicativas da ocorrência de processos ativos de endocitose e de secreção micromerócrina. A quiescência outonal foi caracterizada pelo aspecto anfractuoso do DE; ausência de espermatozóides e pouco material intraluminal, observados à microscopia de luz. Características ultra-estruturais degenerativas foram verificadas ao nível do citoplasma supranuclear das células P epididimárias no outono.(AU)

Small but expressive variability was noted on the epididymidis duct (ED) of domestic quail along the year, with more evidence in autumn of the quiescent phase of the annual testis cycle in this species. Spring features of ED had a general similar pattern in summer and winter. They were characterized by enlargement of epididymis tubule, storage of spermatozoa into the luminal compartment and presence of mitochondria, ER lamellae, several variable vesicles, and lysosomes localized mainly on the apical cytoplasm of principal cells (P) of the epididymal epithelium. These P cells features indicated a process of endocytosis and perhaps protein secretion. Autumn quiescence was marked by a convolute pattern of the epididymis tubule, lacking of spermatozoa and small amount of exfoliate heterogeneous material inside the luminal compartment at light microscopy. Ultrastructural degenerative features mainly apical cytoplasmic debris were seen in the supranuclear cytoplasm of lining P cells.(AU)

Efeito de hormônios e citocinas na expressão da Indoleamina 2,3-dioxigenase e na capacidade proliferativa de células de placenta bovina / Effects of hormones and citokynes in the indoleamine 2,3-dioxygenase expression and the proliferative capacity of cells from bovine placenta

A Indoleamina 2,3-dioxigenase (IDO) é uma enzima que apresenta um importante papel na prevenção da rejeição fetal. A IDO demonstra efeitos na supressão da ativação de células T por catabolizar o aminoácido essencial Triptofano. Nós estudamos a expressão da IDO em cultura celulares de placenta com a adição individual de Estradiol, Progesterona, Interferon , Triptofano e 1-Metil-Triptofano com o uso de citometria de fluxo, imunocitoquímica e quantificação celular, imunofluorescência, peroxidação lipídica, análise das fases do ciclo celular, imunoblotting e PCR em placentas bovinas nos três trimestres gestacionais. A quantificação celular revelou que em bovinos a atividade da IDO aumenta com o avanço do período gestacional e, sua expressão varia de acordo com os fatores, sendo o mesmo padrão observado pela imunofluorescência. O Imunoblotting mostrou a presença de possíveis isoformas desta proteína na espécie bovina que ainda não foram identificadas. A investigação pela lipoperoxidação revelou que os hormônios modularam diferencialmente a produção de radicais peroxidados nos três terços de gestação e, possivelmente atuam como fatores indutores de proliferação. As células placentárias apresentaram padrões diferenciados de proliferação e apoptose ao longo da gestação, principalmente nos tratamentos com Estradiol e Progesterona, sendo também variantes com outros fatores em todos os grupos. Observou-se que a Progesterona atua na maturação das células placentárias independente da concentração utilizada. Em todos os grupos uma grande proporção de células apresentava-se em estado de quiescência (G1). No terceiro trimestre foi detectado um aumento no número de células em G2/M, indicando a parada da capacidade proliferativa ou de progressão no ciclo celular (\"arrest\"). Maior taxa de apoptose foi observada nos animais de segundo trimestre gestacional. Baseados nisso, podemos concluir que a IDO é suscetível ao controle pelos mecanismos testados, o que nos leva a formular hipóteses de implantação de possíveis mecanismos terapêuticos para a reprodução com a participação da IDO

Indoleamina 2,3-dioxigenase (IDO) is an enzyme that plays an important role in preventing fetal rejection. IDO is related to the suppression of the T-cells activation due to the catabolism of essential amino acid Triptophan. We studied the expression of IDO in bovine placenta cell culture with individual supplementation of factors as Estrogen, Progesterone, Interferon , Triptophan and 1-Methyl-Triptophan. Evaluations were made by flow cytometry, immunocitochemistry and cellular quantification, lipidic peroxidation, cell cycle phases, imunoblotting, PCR and immunofluorescency in the three trimesters of pregnancy. Cellular quantification demonstrated that in bovines the activity of the IDO increases during pregnancy, and its expression is factor-dependent, which was also observed in immunofluorescency. Imunoblotting demonstrated the presence of possible unknown protein isoforms in bovine. Lipoperoxidation evaluation demonstrated that hormones distinguishingly modulated the production of peroxide radicals in the three trimesters of pregnancy and, possibly acting as inductive factors of proliferation. Placental cells demonstrated differentiated patterns of proliferation and apoptosis during the gestation, mainly in the presence of Estrogen and Progesterone, besides variant rates were also observed under other factors. Independent of the concentration, Progesterone influenced placental cells maturation. All groups presented great ratio of cells in quiescence state (G1). In third trimester, G2/M high rates indicated a pause in proliferative capacity or in cell cycle progression (arrest). High apoptosis rates were observed in animals at second pregnancy trimester. Based on the presented, we concluded that IDO is susceptible to be controlled by the applied-factors, what lead us to think about hypothetical therapeuthic mechnism for reproduction with the participation of IDO