Histopathological, ultrastructural, and immunohistochemical examination of changes in the placenta as a result of severe preeclampsia

Purpose: To investigate the role of hypoxia-inducible transcription factor-1 alpha (HIF-1α) and angiogenetic factor endothelin-1 (ET-1) expression in regulating hypoxia and placental development by routine histopathological methods. Methods: Twenty preeclamptic and normal placentas were used. Placenta tissue pieces were examined histopathologically after routine paraffin follow-ups. HIF-1α and ET-1 proteins were examined immunohistochemically, and placental tissues were examined ultrastructurally. Results: Increase in syncytial proliferation, endothelial damage in vessels, and increase in collagen were observed in preeclamptic placentas. As a result of preeclampsia, an increase was observed in HIF-1α and ET-1 protein levels in the placenta. Dilatation of endoplasmic reticulum and loss of cristae in mitochondria were observed in trophoblast cells in preeclamptic placental sections. Conclusion: High regulation of oxygen resulting from preeclampsia has been shown to be a critical determinant of placentagenesis and plays an important role in placental differentiation, changes in maternal and fetal blood circulation, trophoblastic invasion, and syncytial node increase. It has been thought that preeclampsia affects secretion by disrupting the endoplasmic reticulum structure and induces mitochondrial damage, and that ET-1 may potentially help in the induction of stress pathways as a result of hypoxia in preeclampsia.

Placenta specific gene targeting to study histone lysine demethylase and androgen signaling in ruminant placenta

Reproductive efficiency is critically dependent on embryo survival, establishment of a successful pregnancy and placental development. Recent advances in gene editing technology have enabled investigators to use gene knockdown and knockout approaches to better understand the role of hormone signaling in placental function and fetal growth and development. In this review, an overview of ruminant placentation will be provided, including recent data highlighting the role of histone lysine demethylase 1A and androgen signaling in ruminant placenta and pregnancy. Studies in ruminant placenta establish a role for histone lysine demethylase 1A in controlling genetic networks necessary for important cellular events such as cell proliferation and angiogenesis, as well as androgen receptor signaling during early placentation.

Placenta specific gene targeting to study histone lysine demethylase and androgen signaling in ruminant placenta

Reproductive efficiency is critically dependent on embryo survival, establishment of a successful pregnancy and placental development. Recent advances in gene editing technology have enabled investigators to use gene knockdown and knockout approaches to better understand the role of hormone signaling in placental function and fetal growth and development. In this review, an overview of ruminant placentation will be provided, including recent data highlighting the role of histone lysine demethylase 1A and androgen signaling in ruminant placenta and pregnancy. Studies in ruminant placenta establish a role for histone lysine demethylase 1A in controlling genetic networks necessary for important cellular events such as cell proliferation and angiogenesis, as well as androgen receptor signaling during early placentation.(AU)

Evolution of placentation in cattle and antelope

Bovids have enjoyed great evolutionary success as evidenced by the large number of extant species. Several important domestic animals are from this family. They derive from both subfamilies: cattle and their kin belong to Bovinae and sheep and goats to Antilopinae. The premise of this review, therefore, is that evolution of reproduction and placentation is best understood in a context that includes antelope-like bovines and antelopes. Many key features of placentation, including hormone secretion, had evolved before bovids emerged as a distinct group. Variation nevertheless occurs. Most striking is the difference in fusion of the binucleate trophoblast cell with uterine epithelium that yields a transient trinucleate cell in bovines and many antelopes, but a more persistent syncytium in wildebeest, sheep and goat. There is considerable variation in placentome number and villus branching within the placentome. Many antelopes have right-sided implantation in a bicornuate uterus whilst others have a uterus duplex. Finally, there has been continued evolution of placental hormones with tandem duplication of PAG genes in cattle, differences in glycosylation of placental lactogen and the emergence of placental growth hormone in sheep and goats. The selection pressures driving this evolution are unknown though maternal-fetal competition for nutrients is an attractive hypothesis.

Evolution of placentation in cattle and antelope

Bovids have enjoyed great evolutionary success as evidenced by the large number of extant species. Several important domestic animals are from this family. They derive from both subfamilies: cattle and their kin belong to Bovinae and sheep and goats to Antilopinae. The premise of this review, therefore, is that evolution of reproduction and placentation is best understood in a context that includes antelope-like bovines and antelopes. Many key features of placentation, including hormone secretion, had evolved before bovids emerged as a distinct group. Variation nevertheless occurs. Most striking is the difference in fusion of the binucleate trophoblast cell with uterine epithelium that yields a transient trinucleate cell in bovines and many antelopes, but a more persistent syncytium in wildebeest, sheep and goat. There is considerable variation in placentome number and villus branching within the placentome. Many antelopes have right-sided implantation in a bicornuate uterus whilst others have a uterus duplex. Finally, there has been continued evolution of placental hormones with tandem duplication of PAG genes in cattle, differences in glycosylation of placental lactogen and the emergence of placental growth hormone in sheep and goats. The selection pressures driving this evolution are unknown though maternal-fetal competition for nutrients is an attractive hypothesis.(AU)

Subplacental development in Galea spixii / Desenvolvimento subplacentário do preá (Galea spixii)

Animal models are essential to understand healthy human placentation. Guinea pig related rodents became on focus for such purposes. In particular, processes of trophoblast invasion are similar. The latter is associated with a specialized area, the subplacenta. Since previous results showed differences between the guinea pig and its close relative Galea spixii, we aimed to study subplacental development with more detail. We investigated 16 pregnant females of 14 to 55 days of gestation by means of histology, morphometrics, immunohistochemistry and electron microscopy. The overlap between the fetomaternal blood systems resulted as intimate, suggesting some exchange processes. Proliferation was revealed by three independent methods, being most active in early and mid-gestation, which was in accordance to former results. Though degeneration of tissues took place, the subplacenta was maintained towards term with access to the fetal vascularization, supporting a hypothesis about the release of substances to the fetal unit in advanced gestation. In contrast to other species, the extraplacental trophoblast showed a shift from syncytial streamers to giant cells during mid-gestation. Views on placentation in caviomorphs were influenced by the guinea pig, but our data supported recent studies that the subplacenta had a much greater placidity. In regard to subplacental grow, degeneration and likely also exchange processes, Galea and other species showed a more basal pattern of caviomorphs than the guinea pig. Such differences should be considered, when choosing most adequate animal models for special purposes in comparison to human placentation.(AU)

Modelos animais são essenciais para entender a placenta humana sadia. Neste sentido os roedores relacionados ao porquinho da índia tornaram-se foco para tal entendimento. Em particular, os processos de invasão trofoblástica são semelhantes. O último está associado a uma área especializada, a subplacenta. Uma vez que os resultados anteriores mostraram diferenças entre o porquinho da índia e seu relativo o preá, buscamos estudar o desenvolvimento subplacentário com mais detalhes. Pesquisamos 16 fêmeas gestantes de 14 a 55 dias de gestação por meio de histologia, morfometria, imuno-histoquímica e microscopia eletrônica. A sobreposição entre os sistemas sanguíneos materno e fetal apresentou-se com íntima relação, sugerindo alguns processos de troca. A proliferação foi revelada por três métodos independentes, sendo mais ativos no início e metade da gestação, o que corroborou com os resultados anteriores. Embora a degeneração dos tecidos tenha ocorrido, a subplacenta foi mantida até o termo gestacional com acesso à vascularização fetal, apoiando uma hipótese sobre a liberação de substâncias para a unidade fetal em gestação avançada. Em contraste com outras espécies, o trofoblasto extraplacentário mostrou uma mudança de flâmulas sinciciais para células gigantes durante a metade da gestação. As visualizações sobre a placentação em caviomorfos foram influenciadas pelo porquinho da índia, mas nossos dados apoiaram estudos recentes de que a subplacenta apresentava uma plasticidade muito maior. Em relação ao crescimento subplacentário, a degeneração e provavelmente também os processos de troca, o preá e outras espécies apresentaram um padrão mais basal de caviomorfos do que o porquinho da índia. Tais diferenças devem ser consideradas, ao escolher os modelos animais mais adequados para fins especiais em comparação com a placentação humana.(AU)

Subplacental development in Galea spixii / Desenvolvimento subplacentário do preá (Galea spixii)

Animal models are essential to understand healthy human placentation. Guinea pig related rodents became on focus for such purposes. In particular, processes of trophoblast invasion are similar. The latter is associated with a specialized area, the subplacenta. Since previous results showed differences between the guinea pig and its close relative Galea spixii, we aimed to study subplacental development with more detail. We investigated 16 pregnant females of 14 to 55 days of gestation by means of histology, morphometrics, immunohistochemistry and electron microscopy. The overlap between the fetomaternal blood systems resulted as intimate, suggesting some exchange processes. Proliferation was revealed by three independent methods, being most active in early and mid-gestation, which was in accordance to former results. Though degeneration of tissues took place, the subplacenta was maintained towards term with access to the fetal vascularization, supporting a hypothesis about the release of substances to the fetal unit in advanced gestation. In contrast to other species, the extraplacental trophoblast showed a shift from syncytial streamers to giant cells during mid-gestation. Views on placentation in caviomorphs were influenced by the guinea pig, but our data supported recent studies that the subplacenta had a much greater placidity. In regard to subplacental grow, degeneration and likely also exchange processes, Galea and other species showed a more basal pattern of caviomorphs than the guinea pig. Such differences should be considered, when choosing most adequate animal models for special purposes in comparison to human placentation.(AU)

Modelos animais são essenciais para entender a placenta humana sadia. Neste sentido os roedores relacionados ao porquinho da índia tornaram-se foco para tal entendimento. Em particular, os processos de invasão trofoblástica são semelhantes. O último está associado a uma área especializada, a subplacenta. Uma vez que os resultados anteriores mostraram diferenças entre o porquinho da índia e seu relativo o preá, buscamos estudar o desenvolvimento subplacentário com mais detalhes. Pesquisamos 16 fêmeas gestantes de 14 a 55 dias de gestação por meio de histologia, morfometria, imuno-histoquímica e microscopia eletrônica. A sobreposição entre os sistemas sanguíneos materno e fetal apresentou-se com íntima relação, sugerindo alguns processos de troca. A proliferação foi revelada por três métodos independentes, sendo mais ativos no início e metade da gestação, o que corroborou com os resultados anteriores. Embora a degeneração dos tecidos tenha ocorrido, a subplacenta foi mantida até o termo gestacional com acesso à vascularização fetal, apoiando uma hipótese sobre a liberação de substâncias para a unidade fetal em gestação avançada. Em contraste com outras espécies, o trofoblasto extraplacentário mostrou uma mudança de flâmulas sinciciais para células gigantes durante a metade da gestação. As visualizações sobre a placentação em caviomorfos foram influenciadas pelo porquinho da índia, mas nossos dados apoiaram estudos recentes de que a subplacenta apresentava uma plasticidade muito maior. Em relação ao crescimento subplacentário, a degeneração e provavelmente também os processos de troca, o preá e outras espécies apresentaram um padrão mais basal de caviomorfos do que o porquinho da índia. Tais diferenças devem ser consideradas, ao escolher os modelos animais mais adequados para fins especiais em comparação com a placentação humana.(AU)

Placental lesions associated with abortion and stillbirth in goats naturally infected by Neospora caninum / Lesões placentárias associadas a abortos e natimortos em cabras naturalmente infectadas pelo Neospora caninum

Neospora caninum has been described as a parasite that sporadically causes reproductive problems in goats. Several aspects of the pathogenesis of neosporosis in naturally infected goats remain to be established. The aims of the present study were to characterize the placental lesions in goats naturally infected by N. caninum and to evaluate several diagnostic techniques for effective detection of this protozoan in the goat placenta. Some placentas in this study originated from abortion and stillbirth in which there were severe lesions. The lesions were characterized mainly by necrosis involving the mesenchyme of the chorionic villi and trophoblast cells often alongside mononuclear inflammation and in some cases with neutrophilic infiltration. N. caninum DNA was detected in these placentas, but parasite structures were not visualized through immunohistochemistry (IHC). However, five of 11 placentas from N. caninum-infected goats that gave birth to healthy kids had histological lesions characterized by mononuclear inflammation. Of these 11 placentas, N. caninum DNA was detected in seven, and N. caninum tachyzoites were detected in only one of these seven placentas using IHC. The present study demonstrates that severe lesions in the placenta are associated with abortion and stillbirth in caprine neosporosis and the placental alterations are likely involved in abortion pathogenesis. Moreover, the results highlight the importance of using more than one diagnostic technique for the detection of the protozoan in placentas because N. caninum cannot be reliably detected by histological and immunohistochemical tests.(AU)

Neospora caninum é descrito como um parasito que causa problemas reprodutivos esporádicos em cabras. Muitos aspectos da patogênese da neosporose em cabras naturalmente infectadas ainda precisam ser estabelecidos. Os objetivos deste trabalho foram caracterizar as lesões placentárias em cabras naturalmente infectadas por N. caninum e avaliar as técnicas diagnósticas para a detecção efetiva do protozoário na placenta. Algumas placentas deste estudo são originárias de abortos e natimortos, nas quais havia lesões graves. As lesões foram classificadas principalmente por necrose envolvendo o mesênquima das vilosidades coriônicas e células trofoblásticas, geralmente associadas a infiltrado inflamatório mononuclear e em alguns casos infiltrado neutrofílico. O DNA do N. caninum foi detectado nestas placentas, porém estruturas parasitárias não foram visualizadas na imuno-histoquímica (IHQ). Entretanto, cinco das onze placentas de cabras infectadas, que deram à luz a conceptos saudáveis, apresentaram lesões histológicas caracterizadas por infiltrado inflamatório mononuclear. Destas 11 placentas, foi detectado DNA de N. caninum em sete e taquizoítos foram encontrados em apenas uma por meio de imuno-histoquímica. O presente estudo demonstra que abortos e natimortos na espécie caprina, causados pelo N. caninum estão associados a lesões acentuadas nas placentas, sendo que as mesmas estão envolvidas na patogênese do aborto. Os resultados também ressaltam a importância do uso de mais de uma técnica diagnóstica para a detecção do protozoário em placentas, pois o N. caninum não pode ser confiavelmente detectado somente pelos exames de histopatologia e imuno-histoquímica.(AU)

Parthenogenetic bovine embryos secrete type I interferon capable of stimulating ISG15 in luteal cell culture

Interferon tau (IFNT) is the pregnancy recognition signal in ruminants and is secreted by trophoblast cells. Paracrine action in the endometrium is well established by inhibiting luteolytic pulses of prostaglandin F2 alpha. Recently, endocrine action was documented in the corpus luteum, blood cell and liver. It was hypothesized that conditioned medium (CM) obtained from days 7, 9 and 12 parthenogenetic embryos alters luteal cell gene expression. The aim was to establish a bovine mixed luteal cell culture to evaluate cellular response associated to interferon stimulated genes, steroidogenesis and apoptosis. Conditioned medium was obtained from Days 7, 9 and 12 parthenogenetic (PA) embryos culture. Moreover, antiviral assay was performed on CM from Days 7, 9 and 12 to verify Type I interferon activity. Luteal cell culture was validated by steroidogenic and apoptotic genes (CYP11A1, HSD3B1, BAX, BCL2, AKT and XIAP mRNA expression), and concentration of progesterone as endpoint. Luteal cell culture was treated with interferon alpha (IFNA) and CM from parthenogenetic embryos. Antiviral assay revealed Type I interferon activity on CM from embryos increasing on Days 9 and 12. ISG15 mRNA was greater in the mixed luteal cells culture treated with 1, 10 and 100ng/ml of interferon alpha (IFNA) and also on Days 7, 9 and 12 CM treatments. Concentration of progesterone was not altered in luteal cell culture regardless of treatments. Steroidogenic and apoptotic genes were similar among groups in luteal cell culture treated with different doses of IFNA or CM from PA embryos. In conclusion, parthenogenetic embryo-derived CM has antiviral activity, luteal cell culture respond to Type I interferon by expressing IGS15. These data indicate this model can be used for IFNT endocrine signaling studies.

Placental lesions associated with abortion and stillbirth in goats naturally infected by Neospora caninum / Lesões placentárias associadas a abortos e natimortos em cabras naturalmente infectadas pelo Neospora caninum

Neospora caninum has been described as a parasite that sporadically causes reproductive problems in goats. Several aspects of the pathogenesis of neosporosis in naturally infected goats remain to be established. The aims of the present study were to characterize the placental lesions in goats naturally infected by N. caninum and to evaluate several diagnostic techniques for effective detection of this protozoan in the goat placenta. Some placentas in this study originated from abortion and stillbirth in which there were severe lesions. The lesions were characterized mainly by necrosis involving the mesenchyme of the chorionic villi and trophoblast cells often alongside mononuclear inflammation and in some cases with neutrophilic infiltration. N. caninum DNA was detected in these placentas, but parasite structures were not visualized through immunohistochemistry (IHC). However, five of 11 placentas from N. caninum-infected goats that gave birth to healthy kids had histological lesions characterized by mononuclear inflammation. Of these 11 placentas, N. caninum DNA was detected in seven, and N. caninum tachyzoites were detected in only one of these seven placentas using IHC. The present study demonstrates that severe lesions in the placenta are associated with abortion and stillbirth in caprine neosporosis and the placental alterations are likely involved in abortion pathogenesis. Moreover, the results highlight the importance of using more than one diagnostic technique for the detection of the protozoan in placentas because N. caninum cannot be reliably detected by histological and immunohistochemical tests.(AU)

Neospora caninum é descrito como um parasito que causa problemas reprodutivos esporádicos em cabras. Muitos aspectos da patogênese da neosporose em cabras naturalmente infectadas ainda precisam ser estabelecidos. Os objetivos deste trabalho foram caracterizar as lesões placentárias em cabras naturalmente infectadas por N. caninum e avaliar as técnicas diagnósticas para a detecção efetiva do protozoário na placenta. Algumas placentas deste estudo são originárias de abortos e natimortos, nas quais havia lesões graves. As lesões foram classificadas principalmente por necrose envolvendo o mesênquima das vilosidades coriônicas e células trofoblásticas, geralmente associadas a infiltrado inflamatório mononuclear e em alguns casos infiltrado neutrofílico. O DNA do N. caninum foi detectado nestas placentas, porém estruturas parasitárias não foram visualizadas na imuno-histoquímica (IHQ). Entretanto, cinco das onze placentas de cabras infectadas, que deram à luz a conceptos saudáveis, apresentaram lesões histológicas caracterizadas por infiltrado inflamatório mononuclear. Destas 11 placentas, foi detectado DNA de N. caninum em sete e taquizoítos foram encontrados em apenas uma por meio de imuno-histoquímica. O presente estudo demonstra que abortos e natimortos na espécie caprina, causados pelo N. caninum estão associados a lesões acentuadas nas placentas, sendo que as mesmas estão envolvidas na patogênese do aborto. Os resultados também ressaltam a importância do uso de mais de uma técnica diagnóstica para a detecção do protozoário em placentas, pois o N. caninum não pode ser confiavelmente detectado somente pelos exames de histopatologia e imuno-histoquímica.(AU)

Parthenogenetic bovine embryos secrete type I interferon capable of stimulating ISG15 in luteal cell culture

Interferon tau (IFNT) is the pregnancy recognition signal in ruminants and is secreted by trophoblast cells. Paracrine action in the endometrium is well established by inhibiting luteolytic pulses of prostaglandin F2 alpha. Recently, endocrine action was documented in the corpus luteum, blood cell and liver. It was hypothesized that conditioned medium (CM) obtained from days 7, 9 and 12 parthenogenetic embryos alters luteal cell gene expression. The aim was to establish a bovine mixed luteal cell culture to evaluate cellular response associated to interferon stimulated genes, steroidogenesis and apoptosis. Conditioned medium was obtained from Days 7, 9 and 12 parthenogenetic (PA) embryos culture. Moreover, antiviral assay was performed on CM from Days 7, 9 and 12 to verify Type I interferon activity. Luteal cell culture was validated by steroidogenic and apoptotic genes (CYP11A1, HSD3B1, BAX, BCL2, AKT and XIAP mRNA expression), and concentration of progesterone as endpoint. Luteal cell culture was treated with interferon alpha (IFNA) and CM from parthenogenetic embryos. Antiviral assay revealed Type I interferon activity on CM from embryos increasing on Days 9 and 12. ISG15 mRNA was greater in the mixed luteal cells culture treated with 1, 10 and 100ng/ml of interferon alpha (IFNA) and also on Days 7, 9 and 12 CM treatments. Concentration of progesterone was not altered in luteal cell culture regardless of treatments. Steroidogenic and apoptotic genes were similar among groups in luteal cell culture treated with different doses of IFNA or CM from PA embryos. In conclusion, parthenogenetic embryo-derived CM has antiviral activity, luteal cell culture respond to Type I interferon by expressing IGS15. These data indicate this model can be used for IFNT endocrine signaling studies.(AU)

Ultrastructure of bovine placenta during all gestational period

ABSTRACT Placentas from pregnant cows with different gestation periods were used. Placental fragments of all groups were processed and evaluated by transmission electron microscopy. After fragment analysis, bovine placenta was observed to be epitheliochorial type in early pregnancy, becoming progressively sinepiteliocorial at the beginning of the second trimester. There are no ultrastructural evidences of inflammation in the region of caruncles throughout gestation, despite the invasion of caruncle proper lamina by trophoblast cells. However, throughout pregnancy and especially at the end, there were evident signs of cell degeneration in both trophoblast and the uterine epithelium. The active trophoblast cells intensely phagocytize cellular debris. There are complex interdigitations between the surface of the trophoblast and the uterine epithelium, which is related to the increase of the exchange surface between mother and fetus. At the end of pregnancy, interdigitations disappear, favoring the detachment and expulsion of the placenta after birth.

RESUMO Foram utilizadas placentas de vacas abatidas em frigorífico com diversos tempos gestacionais. Fragmentos de placentomo de todos os grupos foram processados e avaliados em microscopia eletrônica de transmissão. Após análise dos fragmentos, observou-se que a placenta bovina é do tipo epiteliocorial no início da gestação, tornando-se sinepiteliocorial progressivamente a partir do início do segundo mês de gestação. Não existem evidências ultraestruturais de inflamação na região das carúnculas durante toda a gestação, apesar da invasão da lâmina própria caruncular por células trofoblásticas. No entanto, durante toda a gestação e em especial ao seu final, foram observados sinais evidentes de degeneração celular, tanto do trofoblasto como do epitélio uterino. As células trofoblásticas ativas fagocitam intensamente os debris celulares originados dessas degenerações. Existem complexas interdigitações entre a superfície do trofoblasto e do epitélio uterino, o que estaria relacionado com o aumento da superfície de troca entre mãe e feto. Ao final da gestação, praticamente desaparecem essas interdigitações, favorecendo o descolamento e a expulsão da placenta após o parto.

Trophoblast of domestic and companion animals: basic and applied clinical perspectives

The trophoblast is the single most important functional structure of the placenta that mediates the attachment of the blastocyst to the endometrium. Trophoblast has the capability to proliferate in concert with the uterine epithelium to form the only site for nutrient and metabolic exchange between the fetus and dam throughout pregnancy. Trophoblast is made up of a remarkable versatile epithelium showing great capacity for invasion, cell fusion, hormone production, specific nutrient absorption, selective transport, active metabolism, and has the ability to resist maternal immunological attack. These functions are attributed to its inherent ability to synthesize many developmental factors or molecular regulators. While there is an abundance of publications available on the structural, functional, and clinical relevance of the placenta in various mammalian species, a comprehensive review on the comparative aspects of the trophoblast of domestic and companion animals is lacking. Besides, a timely description on the clinical perspective on the functional aspects of the trophoblast in relation to pregnancy diagnosis, placental insufficiency, pregnancy loss, and structural abnormalities of domestic and companion animals is necessary. A brief description on the basic chronology of events in each animal is followed by the applied clinical perspectives of trophoblast. Both the above aspects of trophoblasts of domestic and companion animals including the terminologies are summarized in tables to facilitate discussion.

Trophoblast of domestic and companion animals: basic and applied clinical perspectives

The trophoblast is the single most important functional structure of the placenta that mediates the attachment of the blastocyst to the endometrium. Trophoblast has the capability to proliferate in concert with the uterine epithelium to form the only site for nutrient and metabolic exchange between the fetus and dam throughout pregnancy. Trophoblast is made up of a remarkable versatile epithelium showing great capacity for invasion, cell fusion, hormone production, specific nutrient absorption, selective transport, active metabolism, and has the ability to resist maternal immunological attack. These functions are attributed to its inherent ability to synthesize many developmental factors or molecular regulators. While there is an abundance of publications available on the structural, functional, and clinical relevance of the placenta in various mammalian species, a comprehensive review on the comparative aspects of the trophoblast of domestic and companion animals is lacking. Besides, a timely description on the clinical perspective on the functional aspects of the trophoblast in relation to pregnancy diagnosis, placental insufficiency, pregnancy loss, and structural abnormalities of domestic and companion animals is necessary. A brief description on the basic chronology of events in each animal is followed by the applied clinical perspectives of trophoblast. Both the above aspects of trophoblasts of domestic and companion animals including the terminologies are summarized in tables to facilitate discussion.(AU)

Ultrastructure of bovine placenta during all gestational period / Ultraestrutura de placenta bovina durante todo o período gestacional

Placentas from pregnant cows with different gestation periods were used. Placental fragments of all groups were processed and evaluated by transmission electron microscopy. After fragment analysis, bovine placenta was observed to be epitheliochorial type in early pregnancy, becoming progressively sinepiteliocorial at the beginning of the second trimester. There are no ultrastructural evidences of inflammation in the region of caruncles throughout gestation, despite the invasion of caruncle proper lamina by trophoblast cells. However, throughout pregnancy and especially at the end, there were evident signs of cell degeneration in both trophoblast and the uterine epithelium. The active trophoblast cells intensely phagocytize cellular debris. There are complex interdigitations between the surface of the trophoblast and the uterine epithelium, which is related to the increase of the exchange surface between mother and fetus. At the end of pregnancy, interdigitations disappear, favoring the detachment and expulsion of the placenta after birth.(AU)

Foram utilizadas placentas de vacas abatidas em frigorífico com diversos tempos gestacionais. Fragmentos de placentomo de todos os grupos foram processados e avaliados em microscopia eletrônica de transmissão. Após análise dos fragmentos, observou-se que a placenta bovina é do tipo epiteliocorial no início da gestação, tornando-se sinepiteliocorial progressivamente a partir do início do segundo mês de gestação. Não existem evidências ultraestruturais de inflamação na região das carúnculas durante toda a gestação, apesar da invasão da lâmina própria caruncular por células trofoblásticas. No entanto, durante toda a gestação e em especial ao seu final, foram observados sinais evidentes de degeneração celular, tanto do trofoblasto como do epitélio uterino. As células trofoblásticas ativas fagocitam intensamente os debris celulares originados dessas degenerações. Existem complexas interdigitações entre a superfície do trofoblasto e do epitélio uterino, o que estaria relacionado com o aumento da superfície de troca entre mãe e feto. Ao final da gestação, praticamente desaparecem essas interdigitações, favorecendo o descolamento e a expulsão da placenta após o parto.(AU)

Ultrastructure of bovine placenta during all gestational period / Ultraestrutura de placenta bovina durante todo o período gestacional

Placentas from pregnant cows with different gestation periods were used. Placental fragments of all groups were processed and evaluated by transmission electron microscopy. After fragment analysis, bovine placenta was observed to be epitheliochorial type in early pregnancy, becoming progressively sinepiteliocorial at the beginning of the second trimester. There are no ultrastructural evidences of inflammation in the region of caruncles throughout gestation, despite the invasion of caruncle proper lamina by trophoblast cells. However, throughout pregnancy and especially at the end, there were evident signs of cell degeneration in both trophoblast and the uterine epithelium. The active trophoblast cells intensely phagocytize cellular debris. There are complex interdigitations between the surface of the trophoblast and the uterine epithelium, which is related to the increase of the exchange surface between mother and fetus. At the end of pregnancy, interdigitations disappear, favoring the detachment and expulsion of the placenta after birth.(AU)

Foram utilizadas placentas de vacas abatidas em frigorífico com diversos tempos gestacionais. Fragmentos de placentomo de todos os grupos foram processados e avaliados em microscopia eletrônica de transmissão. Após análise dos fragmentos, observou-se que a placenta bovina é do tipo epiteliocorial no início da gestação, tornando-se sinepiteliocorial progressivamente a partir do início do segundo mês de gestação. Não existem evidências ultraestruturais de inflamação na região das carúnculas durante toda a gestação, apesar da invasão da lâmina própria caruncular por células trofoblásticas. No entanto, durante toda a gestação e em especial ao seu final, foram observados sinais evidentes de degeneração celular, tanto do trofoblasto como do epitélio uterino. As células trofoblásticas ativas fagocitam intensamente os debris celulares originados dessas degenerações. Existem complexas interdigitações entre a superfície do trofoblasto e do epitélio uterino, o que estaria relacionado com o aumento da superfície de troca entre mãe e feto. Ao final da gestação, praticamente desaparecem essas interdigitações, favorecendo o descolamento e a expulsão da placenta após o parto.(AU)

The evolution of fetal membranes and placentation in carnivores and ungulates (Ferungulata)

Molecular phylogenetics has made a substantialcontribution to our understanding of the relationships between mammalian orders and has generated trees that can be used to examine the evolution of anatomical and physiological traits. We here summarize findings on fetal membranes and placen tation in Ferungulata, a clade comprising carnivores, pangolins, and even and odd-toed ungulates. Their Early ontogeny shows several conserve traits such as superficial attachment of the blastocyst, amnion formation by folding, a large allantoic sac and a temporary yolk sac placenta. In contrast, Several characters of the chorioallantoic placenta are derived, including the diffuse and cotyledonary placental types in ungulate sand zonary placenta in carnivores specializations of the interhaemal barrier, the presence of areolae or haemophagous regions and lack of stromal decidual cells. Ungulates Produce large amounts of placental proteins including placental lactogens and pregnancy-associated glycoproteins. Evolutionary in novations of the placental system may contribute to the high diversity of lifestyles within Ferungulata And be linked to the evolution of highly precocial offspring in ungulates.

The evolution of fetal membranes and placentation in carnivores and ungulates (Ferungulata)

Molecular phylogenetics has made a substantialcontribution to our understanding of the relationships between mammalian orders and has generated trees that can be used to examine the evolution of anatomical and physiological traits. We here summarize findings on fetal membranes and placen tation in Ferungulata, a clade comprising carnivores, pangolins, and even and odd-toed ungulates. Their Early ontogeny shows several conserve traits such as superficial attachment of the blastocyst, amnion formation by folding, a large allantoic sac and a temporary yolk sac placenta. In contrast, Several characters of the chorioallantoic placenta are derived, including the diffuse and cotyledonary placental types in ungulate sand zonary placenta in carnivores specializations of the interhaemal barrier, the presence of areolae or haemophagous regions and lack of stromal decidual cells. Ungulates Produce large amounts of placental proteins including placental lactogens and pregnancy-associated glycoproteins. Evolutionary in novations of the placental system may contribute to the high diversity of lifestyles within Ferungulata And be linked to the evolution of highly precocial offspring in ungulates.(AU)

Apoptose e maturação placentária bovina: um estudo imunohistoquímico e morfométrico / Apoptosis and maturation of the bovine placenta: an immunohistochemical and morphometric study

A liberação da placenta após o parto envolve a perda da adesão materno-fetal e ocorre somente após a maturação completa do placentoma, que está relacionada com a diminuição da celularidade dos tecidos fetal e materno. A apoptose é requerida tanto para a maturação quanto para a liberação normal da placenta após o parto. O objetivo do presente estudo foi avaliar a ocorrência de apoptose em amostras de placenta de vacas em diferentes fases de gestação. Amostras de placentomas de 15 vacas saudáveis com 4 (n=5), 6 (n=5) e 9 (n=5) meses de gestação foram colhidas e processadas rotineiramente para a histologia, imunoistoquímica e histoquímica. As lâminas obtidas foram coradas em HE, Picrosirius Red e submetidas à análise imunoistoquímica das proteínas Caspase 3, Caspase 8, Bax e Bid. O aumento no número de vasos não necessariamente se associou ao aumento do calibre destes durante a evolução da gestação. Os resultados de histomorfometria revelaram aumento da marcação para Bax e Caspases 3 e 8 em células trofoblásticas binucleadas no final da gestação, enquanto o Bid se manteve sem alteração significativa. A histomorfometria das células trofoblásticas mononucleadas revelou expressão alta para Bax no início de gestação, com diminuição aos 6 meses de gestação e aumento das imunomarcações para Caspases 3 e 8, e Bid com o avanço gestacional. Os colágenos tipo I e III não aumentaram do terço médio ao final da gestação, o que é importante para a diminuição da adesão materno-fetal. Esses resultados confirmam que as Caspases 3 e 8, e o Bax estão envolvidos nos mecanismos de ativação da apoptose pela via intrínseca mitocondrial e/ou extrínseca ao longo da gestação em células trofoblásticas binucleadas, e que nas células trofoblásticas mononucleadas o Bax deixa de ser importante, enquanto o Bid e as Caspases 3 e 8 se tornam os mais significativos.(AU)

Placental release after birth involves loss of maternal-fetal adhesion and occurs only after complete maturation of the placentoma related to the decrease in cellularity of fetal and maternal tissues. Apoptosis is required for both the normal maturation and release of the placenta after birth. The aim of this study was to evaluate the occurrence of apoptosis in samples of the placenta of cows in different stages of gestation. Samples of 15 healthy cow placentomas at 4 (n=5), 6 (n=5) and 9 (n=5) months of gestation were harvested and processed for routine histology, immunohistochemistry and histochemistry. The slides were stained with HE, PicroSirius Red and subjected to immunohistochemical analysis of proteins Caspase 3, Caspase 8, Bax and Bid. Increase in number of vessels was not associated with increase in vascular area during progression of gestation. The results of histomorphometry revealed increased labeling for Bax and Caspases 3 and 8 in trophoblastic binucleated cells in late pregnancy, where the Bid remained without significant change. Histomorphometry analyzing the mononuclear trophoblast cells showed a high expression for Bax in early pregnancy, but decreased at 6 months of gestation. Immunolabeling revealed increased Caspases 3/8 and Bid with advancing of gestation. Further evaluation of type I and III collagen showed a decrease of both types of collagens at the end of gestation, what is very important for the reduction of maternal-fetal adhesion. These results confirm that Caspases 3 and 8 and Bax are involved in the mechanisms of activation of apoptosis through mitochondrial intrinsic and/or extrinsic pathway during pregnancy in trophoblastic binucleated cells. In mononuclear trophoblast cells Bax looses importance in the apoptosis process, awhile Bid and caspases 3 and 8 become the most significant.(AU)

PAPEL DA KISSPEPTINA E SEU POTENCIAL TERAPÊUTICO NA DISFUNÇÃO FETO-PLACENTÁRIA DE RATAS HIPOTIREOIDEAS: AVALIAÇÃO DE FATORES DE CRESCIMENTO PLACENTÁRIO E MEDIADORES DE ESTRESSE CELULAR E DO INFLAMASSOMA

Alterações no nível plasmático e/ou placentário de kisspeptina (Kp) estão associadas 9 com doenças gestacionais. No entanto, embora estudos tenham sugerido que Kp 10 apresenta efeito antioxidante e imunomodulatório na interface materno-fetal e outros 11 tenham demonstrado que o hipotireoidismo causa disfunção gestacional e estresse 12 placentário, ainda não há informações sobre o papel da kisspeptina na disfunção 13 placentária causada pelo hipotireoidismo materno. Por isso, o objetivo deste trabalho 14 foi avaliar o papel da kisspeptina e seu potencial terapêutico na disfunção feto- 15 placentária de ratas hipotireoideas. Foram utilizadas Ratas Wistar adultas e o 16 hipotireoidismo foi induzido pela administração diária de propiltiouracil (PTU) 17 (4mg/Kg/dia). No Experimento 1, os animais foram divididos nos grupos controle 18 (N=18) e hipotireoideo (N=20) e foram avaliados o desenvolvimento feto-placentário e 19 a expressão gênica e proteica de Kiss1 e seu receptor (Kiss1R) na decídua e placenta 20 aos 14 e 18 dias de gestação (DG). O hipotireoidismo reduziu o peso fetal e 21 uteroplacentário, além de reduzir a expressão de mRNA de Kiss1 na decídua e 22 placenta aos 14 e 18 DG, respectivamente. Para Kiss1R, o hipotireoidismo não 23 somente reduziu a imunomarcação no triângulo Metrial e camada de células gigantes 24 aos 14 DG, como reduziu a expressão gênica na decídua e placenta no mesmo 25 período gestacional. Considerando que teve redução do sistema Kiss1/Kiss1R na 26 interface materno-fetal das ratas hipotireoideas, no Experimento 2 foi avaliado o 27 potencial terapêutico de Kisspeptina-10 (Kp10) na disfunção feto-placentária desses 28 animais. Os animais foram divididos nos grupos controle (N=8), hipotireoideo (N=8) e 29 hipotireoideo tratado com Kp10 (8µg/Kg/dia) dia sim/dia não (KpT1) (N=8) ou 30 diariamente (KpT2) (N=8). O tratamento iniciou no 8º DG e no 18º DG foram avaliados 31 o desenvolvimento feto-placentário, a histomorfometria placentária e a expressão 32 gênica e/ou proteica de fatores de crescimento (VEGF, PLGF, IGF1, IGF2 e GLUT1), 33 mediadores inflamatórios (TNF, IL10 e IL6), mediadores de estresse oxidativo (HIF, 34 SOD1, Cat, GPx1), mediadores de estresse de retículo endoplasmático (ATF4, 35 GRP78 e CHOP) e mediadores do inflamassoma (NLRP3, IL1, IL18) e piroptose 36 (Caspase1, Gasdermina D). O tratamento diário com Kp10 aumentou o peso fetal e 37 reverteu o crescimento fetal assimétrico causado pelo hipotireoidismo materno. 38 Ambos os tratamentos reestabeleceram parcialmente a morfologia da zona juncional 39 e aumentaram a vascularização no labirinto placentário. O tratamento diário com Kp10 40 aumentou a expressão gênica de fatores de crescimento (Plgf, Igf1) e de transporte 41 (Glut1) na placenta dos animais hipotireoideos, além de aumentar a expressão 42 proteica e de mRNA de enzimas antioxidantes (SOD1, Cat, GPx1). Embora o 43 tratamento diário com Kp10 não tenha alterado a maior expressão gênica e/ou 44 proteica de VEGF, HIF1, IL10, GRP78 e CHOP causada pelo hipotireoidismo, 45 especialmente na zona juncional, reduziu a imunomarcação desses mediadores na 46 zona do labirinto quando comparado ao grupo hipotireoideo. Além disso, o tratamento 47 com Kp10 bloqueou o aumento da expressão gênica placentária de mediadores do1 inflamassoma (Nlrp3, Il18) causado pelo hipotireoidismo materno. No entanto, o 2 tratamento com Kp10 aumentou a imunomarcação placentária de TNF e a expressão 3 gênica de Il1 e Caspase1 em relação aos grupos controle e hipotireoideo, enquanto 4 não foi observado diferenças na expressão placentária de Gasdermina D entre os 5 grupos experimentais. Conclui-se que a restrição de crescimento feto-placentária 6 observada em ratas com hipotireoidismo materno está associada com uma redução 7 do sistema kisspeptina/Kiss1R na interface materno-fetal. Além disso, o tratamento 8 diário com Kp10 em ratas gestantes hipotireoideas não somente melhora o 9 desenvolvimento feto-placentário e a vascularização placentária, como aumenta a 10 expressão de fatores de crescimento e enzimas antioxidantes e bloqueia a ativação 11 do inflamassoma causado pelo hipotireoidismo. 12

Changes in the plasma and / or placental level of kisspeptin (Kp) are associated with 9 pregnancy disorders. However, although studies have suggested that Kp has an 10 antioxidant and immunomodulatory effect at the maternal-fetal interface and others 11 have shown that hypothyroidism causes gestational dysfunction and placental stress, 12 there is still no information on the role of kisspeptin in placental dysfunction caused by 13 maternal hypothyroidism. Therefore, the objective of this work was to evaluate the role 14 of kisspeptin and its therapeutic potential in the fetal-placental dysfunction of 15 hypothyroid rats. Adult Wistar rats were used and hypothyroidism was induced by daily 16 administration of propylthiouracil (PTU) (4mg/Kg/day). In Experiment 1, the animals 17 were divided into the control (N=18) and hypothyroid (N=20) groups and the fetal- 18 placental development and the gene and protein expression of Kiss1 and its receptor 19 (Kiss1R) in the decidua and placenta were evaluated at 14 and 18 days of gestation 20 (DG). Hypothyroidism reduced fetal and uteroplacental weight, in addition to reducing 21 Kiss1 mRNA expression in the decidua and placenta at 14 and 18 DG, respectively. 22 For Kiss1R, hypothyroidism not only reduced immunostaining in the Metrial triangle 23 and giant cell layer at 14 DG, but also reduced gene expression in the decidua and 24 placenta in the same gestational period. Considering that there was a reduction in the 25 Kiss1/Kiss1R system in the maternal-fetal interface of the hypothyroid rats, in 26 Experiment 2 the therapeutic potential of Kisspeptin-10 (Kp10) in the fetal-placental 27 dysfunction of these animals was evaluated. The animals were divided into the control 28 (N=8), hypothyroid (N=8) and hypothyroid groups treated with Kp10 (8µg/Kg/day) 29 every other day (KpT1) (N=8) or daily (KpT2) (N=8). The treatment started in the 8th 30 DG and the fetal-placental development, the placental histomorphometry and the gene 31 and/or protein expression of growth factors (VEGF, PLGF, IGF1, IGF2 and GLUT1), 32 inflammatory mediators (TNF, IL10 and IL6), oxidative stress mediators (HIF, SOD1, 33 Cat, GPx1), endoplasmic reticulum stress mediators (ATF4, GRP78 and CHOP) and 34 inflammasome (NLRP3, IL-1, IL-18) and pyroptosis (Caspase1, Gasdermin D) 35 mediators were evaluated at 18 GD. Daily treatment with Kp10 increased fetal weight 36 and reversed asymmetric fetal growth caused by maternal hypothyroidism. Both 37 treatments partially reestablished the morphology of the junctional zone and increased 38 vascularization in the placental labyrinth. Daily treatment with Kp10 increased the gene 39 expression of growth factors (Plgf, Igf1) and transport (Glut1) in the placenta of 40 hypothyroid animals, in addition to increasing protein and mRNA expression of 41 antioxidant enzymes (SOD1, Cat, GPx1). Although daily treatment with Kp10 did not 42 alter the greater VEGF, HIF1, IL10, GRP78 and CHOP expression caused by 43 hypothyroidism, especially in the junctional zone, it reduced the immunostaining of 44 these mediators in the labyrinth zone when compared to the hypothyroid group. In 45 addition, treatment with Kp10 blocked the increase in placental gene expression of 46 inflammasome mediators (Nlrp3, Il18) caused by maternal hypothyroidism. However, 47 treatment with Kp10 increased the placental immunostaining of TNF and the gene 48 expression of Il1 and Caspase1 in relation to the control and hypothyroid groups, 49 while no differences were observed in the placental expression of Gasdermin D 50 between the experimental groups. It is concluded that the fetal-placental growth1 restriction observed in rats with maternal hypothyroidism is associated with a reduction 2 in the kisspeptin/Kiss1R system at the maternal-fetal interface. In addition, daily 3 treatment with Kp10 in hypothyroid pregnant rats not only improves fetal placental 4 development and placental vascularization but increases the expression of growth 5 factors and antioxidant enzymes and blocks the activation of the inflammasome 6 caused by hypothyroidism. 7