N-acylation of phosphatidylethanolamine (PE) with free fatty acids catalyzed by N-acyl phosphatidylethanolamine (NAPE) synthase was reported in cotyledons of 24-h-old cotton seedlings. Here I report subcellular localization of this enzyme. Differential centrifugation, sucrose density gradient fractionation,aqueous two-phase partitioning and electron microscopy techniques were utilized to elucidate subcellular site(s) of NAPE synthase. Marker enzymes were used to locate organelles in subcellular fractions. Differential centrifugation indicated that NAPE synthase is present in more than one organelle and it is a membrane bound enzyme. Sucrose density gradient fractionations indicated that NAPE synthase is present in membranes derived from endoplasmic reticulum (ER),Golgi and possibly plasma membrane (PM) but not mitochondria, glyoxysomes or plastids. Aqueous two-phase partitioning experiments with cotton and spinach tissues supported these results but Goigi appeared to be the major site of NAPE synthesis. Electron microscopy of subcellular fractions was used to examine isolated fractions to provide visual confirmation of our biochemical results. Collectively, these results indicate that NAPE is synthesized in plant ER, Golgi and possibly PM.

A presença do marsúpio fechado possibilitou aos isópodos terrestres a total independência do meio aquático para a reprodução, permitindo aos mesmos a colonização de habitats terrestres variados. Em seu interior, a prole é nutrida e oxigenada através de estruturas temporárias, chamadas cotilédones, as quais surgem durante o período ovígero das fêmeas e são exclusivas do grupo Crinocheta. O presente estudo teve como objetivo descrever padrões dos cotilédones, analisados em 37 espécies de isópodos terrestres distribuídas entre as famílias: Agnaridae, Armadillidae, Armadillidiidae, Balloniscidae, Bathytropidae, Detonidae, Dubioniscidae, Philosciidae, Platyarthridae, Porcellionidae, Pudeoniscidae, Scleropactidae e Trachelipodidae, verificando se ocorre variação de comprimento ao longo das fases de desenvolvimento embrionário, sendo que para isto foram selecionadas seis espécies: Armadillidium nasatum, A. vulgare, Atlantoscia floridana, Balloniscus sellowii, Benthana cairensis e Porcellio scaber. Armadillidium vulgare e B. sellowii também foram estimados quanto à proporção comprimento dos cotilédones/ tamanho da fêmea. Entre as 13 famílias estudadas foram identificados seis formatos (tipos) de cotilédones e sete arranjos diferentes de número e distribuição. O comprimento foi estabelecido como: curto, médio e longo. Armadillidium vulgare, A. floridana e B. cairensis não apresentaram diferença no comprimento dos cotilédones durante os estágios embrionários sendo que este foi evidenciado para as demais espécies, uma vez que B. sellowii e P. scaber apresentaram aumento da fase de ovo para embrião, diminuindo na fase de manca. Já em A. nasatum ocorre um aumento gradual até a fase de manca, onde então regridem drasticamente. O comprimento dos cotilédones de A. vulgare e B. sellowii é proporcional ao tamanho da fêmea. Durante os estudos realizados outra novidade foi registrada: a presença de uma “extensão marsupial”. Para maiores informações sobre esta estrutura, foi necessário identificar quais espécies a apresentavam, o número de indivíduos abrigados em seu interior e diferenças na estrutura entre as espécies que portavam tal extensão e as que possuem o marsúpio fechado restrito aos cinco pares de oostegitos. Seis espécies apresentaram extensão marsupial, a qual possibilita o abrigo de aproximadamente ¼ do total da prole. XI Este marsúpio não apresenta formato distendido como o encontrado nas demais espécies o que vem a influenciar diretamente a alimentação da fêmea, pois com o desenvolvimento da prole ocorrendo em direção aos órgãos desta, conforme avançam as fases, aumenta a pressão interna, comprimindo seu interior. As diferenças morfológicas registradas aqui representam estratégias reprodutivas adotadas pelas espécies, ao longo da evolução, podendo estar relacionadas (ou não) à filogenia dos grupos. / The presence of the closed brood pouch made it possible for terrestrial isopods to have full independence of the aquatic environment for reproduction, allowing them to colonize varied terrestrial habitats. Inside, the offspring is nourished and oxygenated by temporary structures, called cotyledons, which emerge during the ovigerous period of females and are unique to the group Crinocheta. This study aims to describe patterns of cotyledons, analyzed in 37 species of terrestrial isopods distributed among the families: Agnaridae, Armadillidae, Armadillidiidae, Balloniscidae, Bathytropidae, Detonidae, Dubioniscidae, "Philoscidae" Platyarthridae, Porcellionidae, Pudeoniscidae, Scleropactidae and Trachelipodidae , checking if there is variation in length along the stages of embryonic development, and for this, six species were selected as models: Armadillidium nasatum, A. vulgare, Atlantoscia floridana Balloniscus sellowii, Porcellio scaber and Benthana cairensis. Armadillidium vulgare and B. sellowii were also estimated concerning the proportion of cotyledon length / female size. Among the 13 studied families six formats (types) of cotyledons were identified as well as seven different arrangements of numbers and distribution. The length was established as: short, medium and long. Armadillidium vulgare, A. floridana and B. cairensis showed no difference in the length of the cotyledons during the embryonic stages while this was evidenced for the other species, since B. sellowii and P. scaber showed increased egg stage to the embryo, decreasing during the manca. Nevertheless in A. nasatum a gradual increase until the phase of manca occurs, which then regresses dramatically. The length of the cotyledons of A. vulgare and B. sellowii is proportional to female size. XIII During the carried out studies another new feature was recorded: the presence of an " marsupial extension." For obtaining further information about this structure it was necessary to identify the species that presented it, the number of individuals housed within, and differences in structure between species that harbored such extension and the others which had the closed pouch restricted to five pairs of oostegits. Six species showed marsupial extension, which provides shelter for approximately a quarter of the total offspring. This brood pouch does not have a distended format as the one found in other species, which directly influences the feeding of the female, because due to the fact that the offspring development occurs towards its organs as the stages advance, internal pressure increases, compressing its interior. The morphological differences reported here represent reproductive strategies adopted by the species, through evolution and could be related (or not) to the phylogeny of the groups.

En este trabajo se ha estudiado la posibilidad de desarrollar un método de regeneración y transformación para producir plantas transgénicas de albaricoquero con cotiledones y secciones de hipocotilo a partir de semillas maduras de las variedades de albaricoquero ‘Canino’, ‘Moniquí’ y ‘Dorada’ y de la subespecie de albaricoquero, Prunus armeniaca L. var. ansu Maxim, cuyas semillas se utilizan para producir patrones paraalbaricoquero y otros frutales en China. Se ha conseguido establecer un protocolo de regeneración con los cotiledones de semillas de ‘Canino’ y del patrón, Prunus armeniaca L. var. ansu Maxim obteniendo porcentajes de regeneración muy elevados (66%) que no se vieron afectados por el genotipo. Sin embargo, la falta de coincidencia de las células que regeneran con las células que son transformadas por Agrobacterium ha impedido la obtención de plantas transformadas hasta el momento. Utilizando los explantos de secciones de hipocotilos de semillas maduras de tres variedades comerciales de albaricoquero, ‘Dorada’, ‘Moniquí’ y ‘Canino’, se ha establecido un protocolo de regeneración eficiente, con un mínimo del 32% de regeneración para las tres variedades estudiadas, que ha permitido abordar experimentos de transformación. Además, se han conseguido plantas transgénicas de las variedades ‘Dorada’ y ‘Moniquí’. El principal interés de la transformación genética de especies frutales reside en la posibilidad de transformar variedades comerciales con interés agronómico y ampliamente aceptadas. Sin embargo, para que la transformación genética se convierta en una herramienta de uso común en los programas de mejora de frutales leñosos se requiere que los procedimientos sean reproducibles, de elevada eficiencia e independientes del genotipo. La utilización de material adulto garantiza la identidad genética del material vegetal y evita los problemas de juvenilidad en aquellas especies en las que estos periodos son muy largos. Con el fin de explorar la posibilidad de desarrollar un protocolo de transformación independiente del genotipo, en esta tesis se ha estudiado el desarrollo de una nueva metodología basada en la transformación de células con capacidad meristemática. Se ha conseguido transformar cuatro variedades de albaricoquero, ‘Helena’, ‘Canino’, ‘Rojo Pasión’ y ‘Lorna’, con eficiencias de transformación que se encuentran entre las más elevadas descritas en especies de Prunus. Las ventajas de esta metodología son la sencillez del protocolo utilizado, su consistencia y rapidez. Sin embargo, hemos detectado que con mucha frecuencia se producen plantas quiméricas al transformar las células meristemáticas. Otro inconveniente en los protocolos de transformación es la utilización de genes marcadores de resistencia a antibióticos o herbicidas. Estos genes producen un rechazo social e incluso existe una directiva europea (2001/18/CE) que regula las plantas transformadas con genes de resistencia a antibióticos. El uso del gen pmi como marcador de selección podría reducir el rechazo de la opinión pública y facilitar los trámites legales para la comercialización de plantas transgénicas, ya que la proteína que codifica este gen no representa un riesgo para la salud humana ni para el medio ambiente. En la presente tesis se ha estudiado el desarrollo de un protocolo de transformación genética de albaricoquero que utiliza el sistema pmi/manosa para seleccionar las plantas transformadas. Con este sistema se ha conseguido producir plantas transgénicas de albaricoquero con una eficiencia similar a la obtenida con genes marcadores de selección que confieren resistencia a antibióticos, demostrando que puede ser un sistema de selección alternativo al uso de antibióticos. / In this Ph.D. thesis a methodology to develop such a procedure has been studied, based in cells with meristematic capability. We have transformed the apricot cultivars ‘Helena’, ‘Canino’, ‘Rojo Pasión’ and ‘Lorna’, with transformation efficiencies, based on PCR analysis, among the highest reported after transformation of other Prunus species. The advantages of this methodology are that is simple, fast and consistent. However, a large frequency of chimerical plants has been detected when transforming meristematic cells. Another problem of most transformation procedures is that they use antibiotic resistance marker genes. These genes produce a social rejection and there is an European law that neither allow deliberate release of plants carrying antibiotic resistance genes, used in veterinary or medicine, after 2004 nor their commercialisation after 2008 (Directive 2001/18/EEC of the European Parliament and the Council of the European Union). Since the PMI protein has revealed no adverse effects on human health and on the environment, the use of the pmi gen as a selectable marker could reduce the social rejection, facilitating legalisation and commercialisation of transgenic plants. In this Ph.D. thesis we have evaluated a transformation procedure using the pmi/mannose system for selecting apricot transgenic plants. Transformation efficiencies were similar to those obtained using antibiotic selection, demonstrating that this system could be an alternative for producing apricot transgenic plants under safe conditions for human health and the environment.

A presença do marsúpio fechado possibilitou aos isópodos terrestres a total independência do meio aquático para a reprodução, permitindo aos mesmos a colonização de habitats terrestres variados. Em seu interior, a prole é nutrida e oxigenada através de estruturas temporárias, chamadas cotilédones, as quais surgem durante o período ovígero das fêmeas e são exclusivas do grupo Crinocheta. O presente estudo teve como objetivo descrever padrões dos cotilédones, analisados em 37 espécies de isópodos terrestres distribuídas entre as famílias: Agnaridae, Armadillidae, Armadillidiidae, Balloniscidae, Bathytropidae, Detonidae, Dubioniscidae, Philosciidae, Platyarthridae, Porcellionidae, Pudeoniscidae, Scleropactidae e Trachelipodidae, verificando se ocorre variação de comprimento ao longo das fases de desenvolvimento embrionário, sendo que para isto foram selecionadas seis espécies: Armadillidium nasatum, A. vulgare, Atlantoscia floridana, Balloniscus sellowii, Benthana cairensis e Porcellio scaber. Armadillidium vulgare e B. sellowii também foram estimados quanto à proporção comprimento dos cotilédones/ tamanho da fêmea. Entre as 13 famílias estudadas foram identificados seis formatos (tipos) de cotilédones e sete arranjos diferentes de número e distribuição. O comprimento foi estabelecido como: curto, médio e longo. Armadillidium vulgare, A. floridana e B. cairensis não apresentaram diferença no comprimento dos cotilédones durante os estágios embrionários sendo que este foi evidenciado para as demais espécies, uma vez que B. sellowii e P. scaber apresentaram aumento da fase de ovo para embrião, diminuindo na fase de manca. Já em A. nasatum ocorre um aumento gradual até a fase de manca, onde então regridem drasticamente. O comprimento dos cotilédones de A. vulgare e B. sellowii é proporcional ao tamanho da fêmea. Durante os estudos realizados outra novidade foi registrada: a presença de uma “extensão marsupial”. Para maiores informações sobre esta estrutura, foi necessário identificar quais espécies a apresentavam, o número de indivíduos abrigados em seu interior e diferenças na estrutura entre as espécies que portavam tal extensão e as que possuem o marsúpio fechado restrito aos cinco pares de oostegitos. Seis espécies apresentaram extensão marsupial, a qual possibilita o abrigo de aproximadamente ¼ do total da prole. XI Este marsúpio não apresenta formato distendido como o encontrado nas demais espécies o que vem a influenciar diretamente a alimentação da fêmea, pois com o desenvolvimento da prole ocorrendo em direção aos órgãos desta, conforme avançam as fases, aumenta a pressão interna, comprimindo seu interior. As diferenças morfológicas registradas aqui representam estratégias reprodutivas adotadas pelas espécies, ao longo da evolução, podendo estar relacionadas (ou não) à filogenia dos grupos. / The presence of the closed brood pouch made it possible for terrestrial isopods to have full independence of the aquatic environment for reproduction, allowing them to colonize varied terrestrial habitats. Inside, the offspring is nourished and oxygenated by temporary structures, called cotyledons, which emerge during the ovigerous period of females and are unique to the group Crinocheta. This study aims to describe patterns of cotyledons, analyzed in 37 species of terrestrial isopods distributed among the families: Agnaridae, Armadillidae, Armadillidiidae, Balloniscidae, Bathytropidae, Detonidae, Dubioniscidae, "Philoscidae" Platyarthridae, Porcellionidae, Pudeoniscidae, Scleropactidae and Trachelipodidae , checking if there is variation in length along the stages of embryonic development, and for this, six species were selected as models: Armadillidium nasatum, A. vulgare, Atlantoscia floridana Balloniscus sellowii, Porcellio scaber and Benthana cairensis. Armadillidium vulgare and B. sellowii were also estimated concerning the proportion of cotyledon length / female size. Among the 13 studied families six formats (types) of cotyledons were identified as well as seven different arrangements of numbers and distribution. The length was established as: short, medium and long. Armadillidium vulgare, A. floridana and B. cairensis showed no difference in the length of the cotyledons during the embryonic stages while this was evidenced for the other species, since B. sellowii and P. scaber showed increased egg stage to the embryo, decreasing during the manca. Nevertheless in A. nasatum a gradual increase until the phase of manca occurs, which then regresses dramatically. The length of the cotyledons of A. vulgare and B. sellowii is proportional to female size. XIII During the carried out studies another new feature was recorded: the presence of an " marsupial extension." For obtaining further information about this structure it was necessary to identify the species that presented it, the number of individuals housed within, and differences in structure between species that harbored such extension and the others which had the closed pouch restricted to five pairs of oostegits. Six species showed marsupial extension, which provides shelter for approximately a quarter of the total offspring. This brood pouch does not have a distended format as the one found in other species, which directly influences the feeding of the female, because due to the fact that the offspring development occurs towards its organs as the stages advance, internal pressure increases, compressing its interior. The morphological differences reported here represent reproductive strategies adopted by the species, through evolution and could be related (or not) to the phylogeny of the groups.

The establishment and germination in vitro can supply a high amount of explants to develop morphogenesis protocols necessary for flowering induction in vitro, haploid and double-haploid plant production, clonal propagation, somatic embryogenesis among other applications. The objective was to develop protocols for the establishment and germination in vitro of bottle gourd (Lagenaria siceraria (Mol.) Standl.) and squash (Cucurbita pepo L.) species to effort morphogenesis studies. Seed disinfection treatments were done with ethanol 70% and NaOCl. Seed germination in vitro was evaluated changing medium-osmotic pressure, light availability, tegument and auxin presence, imbibition s time and scarification methods. Cotyledonary explants were used to induce direct organogenesis. Apical and nodal segments were grown in MS medium without growth regulators. Bottle gourd seeds did not germinate in culture medium, but it occurred on four layers of germitest paper and distilled water in the proportion of 1:7.5 (w/w). Light is not necessary for bottle gourd seed germination. Osmotic pressure reduction did not increase bottle gourd and squash germination. Bottle gourd apical and nodal segments regenerate in culture medium without growth regulators. Cotyledonary explants of bottle gourd and squash induce aerial growth, but in a low proliferation rate / O estabelecimento e a germinação in vitro suprem explantes em grande quantidade para a organização de experimentos em morfogênese, a qual apresenta muitas aplicações, tais como a indução de flores in vitro, a obtenção de plantas haplóides e duplo-haplóides, a propagação clonal em massa, a embriogênese somática, entre tantas outras. O objetivo deste trabalho foi desenvolver protocolos que permitam o estabelecimento e a germinação in vitro das espécies de porongo (Lagenaria siceraria (Mol.) Standl.) e mogango (Cucurbita pepo L.) para subsidiar pesquisas em morfogênese. Foram realizados testes com álcool 70% e NaOCl (Hipoclorito de sódio) para a desinfestação de sementes e investigados vários fatores envolvidos na germinação in vitro, tais como pressão osmótica, fotoblastismo, presença do tegumento, adição de auxinas, tempo de embebição e escarificação. Explantes cotiledonares foram utilizados para a indução de organogênese direta. Ápices caulinares e segmentos nodais foram cultivados em meio MS sem a adição de reguladores de crescimento. Nenhum dos tratamentos utilizados foi eficiente para permitir a germinação de sementes inteiras de porongo em meio de cultura, porém houve germinação em papel germitest umidecido com 7,5 vezes a massa do papel. Não foi verificado fotoblastismo para o porongo. A redução da pressão osmótica não aumentou o percentual de germinação de porongo e mogango. Ápices caulinares e segmentos nodais de porongo regeneram sem a adição de reguladores de crescimento. Explantes cotiledonares podem ser utilizados para a indução de brotações adventícias, em porongo e mogango, porém a taxa de proliferação é baixa

A presença do marsúpio fechado possibilitou aos isópodos terrestres a total independência do meio aquático para a reprodução, permitindo aos mesmos a colonização de habitats terrestres variados. Em seu interior, a prole é nutrida e oxigenada através de estruturas temporárias, chamadas cotilédones, as quais surgem durante o período ovígero das fêmeas e são exclusivas do grupo Crinocheta. O presente estudo teve como objetivo descrever padrões dos cotilédones, analisados em 37 espécies de isópodos terrestres distribuídas entre as famílias: Agnaridae, Armadillidae, Armadillidiidae, Balloniscidae, Bathytropidae, Detonidae, Dubioniscidae, Philosciidae, Platyarthridae, Porcellionidae, Pudeoniscidae, Scleropactidae e Trachelipodidae, verificando se ocorre variação de comprimento ao longo das fases de desenvolvimento embrionário, sendo que para isto foram selecionadas seis espécies: Armadillidium nasatum, A. vulgare, Atlantoscia floridana, Balloniscus sellowii, Benthana cairensis e Porcellio scaber. Armadillidium vulgare e B. sellowii também foram estimados quanto à proporção comprimento dos cotilédones/ tamanho da fêmea. Entre as 13 famílias estudadas foram identificados seis formatos (tipos) de cotilédones e sete arranjos diferentes de número e distribuição. O comprimento foi estabelecido como: curto, médio e longo. Armadillidium vulgare, A. floridana e B. cairensis não apresentaram diferença no comprimento dos cotilédones durante os estágios embrionários sendo que este foi evidenciado para as demais espécies, uma vez que B. sellowii e P. scaber apresentaram aumento da fase de ovo para embrião, diminuindo na fase de manca. Já em A. nasatum ocorre um aumento gradual até a fase de manca, onde então regridem drasticamente. O comprimento dos cotilédones de A. vulgare e B. sellowii é proporcional ao tamanho da fêmea. Durante os estudos realizados outra novidade foi registrada: a presença de uma “extensão marsupial”. Para maiores informações sobre esta estrutura, foi necessário identificar quais espécies a apresentavam, o número de indivíduos abrigados em seu interior e diferenças na estrutura entre as espécies que portavam tal extensão e as que possuem o marsúpio fechado restrito aos cinco pares de oostegitos. Seis espécies apresentaram extensão marsupial, a qual possibilita o abrigo de aproximadamente ¼ do total da prole. XI Este marsúpio não apresenta formato distendido como o encontrado nas demais espécies o que vem a influenciar diretamente a alimentação da fêmea, pois com o desenvolvimento da prole ocorrendo em direção aos órgãos desta, conforme avançam as fases, aumenta a pressão interna, comprimindo seu interior. As diferenças morfológicas registradas aqui representam estratégias reprodutivas adotadas pelas espécies, ao longo da evolução, podendo estar relacionadas (ou não) à filogenia dos grupos. / The presence of the closed brood pouch made it possible for terrestrial isopods to have full independence of the aquatic environment for reproduction, allowing them to colonize varied terrestrial habitats. Inside, the offspring is nourished and oxygenated by temporary structures, called cotyledons, which emerge during the ovigerous period of females and are unique to the group Crinocheta. This study aims to describe patterns of cotyledons, analyzed in 37 species of terrestrial isopods distributed among the families: Agnaridae, Armadillidae, Armadillidiidae, Balloniscidae, Bathytropidae, Detonidae, Dubioniscidae, "Philoscidae" Platyarthridae, Porcellionidae, Pudeoniscidae, Scleropactidae and Trachelipodidae , checking if there is variation in length along the stages of embryonic development, and for this, six species were selected as models: Armadillidium nasatum, A. vulgare, Atlantoscia floridana Balloniscus sellowii, Porcellio scaber and Benthana cairensis. Armadillidium vulgare and B. sellowii were also estimated concerning the proportion of cotyledon length / female size. Among the 13 studied families six formats (types) of cotyledons were identified as well as seven different arrangements of numbers and distribution. The length was established as: short, medium and long. Armadillidium vulgare, A. floridana and B. cairensis showed no difference in the length of the cotyledons during the embryonic stages while this was evidenced for the other species, since B. sellowii and P. scaber showed increased egg stage to the embryo, decreasing during the manca. Nevertheless in A. nasatum a gradual increase until the phase of manca occurs, which then regresses dramatically. The length of the cotyledons of A. vulgare and B. sellowii is proportional to female size. XIII During the carried out studies another new feature was recorded: the presence of an " marsupial extension." For obtaining further information about this structure it was necessary to identify the species that presented it, the number of individuals housed within, and differences in structure between species that harbored such extension and the others which had the closed pouch restricted to five pairs of oostegits. Six species showed marsupial extension, which provides shelter for approximately a quarter of the total offspring. This brood pouch does not have a distended format as the one found in other species, which directly influences the feeding of the female, because due to the fact that the offspring development occurs towards its organs as the stages advance, internal pressure increases, compressing its interior. The morphological differences reported here represent reproductive strategies adopted by the species, through evolution and could be related (or not) to the phylogeny of the groups.

Orientador: Ivany Ferraz Marques Valio / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-10T03:19:02Z (GMT). No. of bitstreams: 1 Abrao_MarluciMundin_D.pdf: 2033158 bytes, checksum: 02bd87759812f0f20bce7ff1ed06a9d8 (MD5) Previous issue date: 2007 / Doutorado / Biologia Vegetal / Doutor em Biologia Vegetal

Au cours de la grossesse, une perfusion placentaire adéquate est indispensable au bon développement du fœtus. Dans certaines maladies comme la prééclampsie, celle-ci est altérée, compromettant ainsi la vie du fœtus, mais aussi celle de sa mère. Le retrait du placenta mène à la disparition des symptômes de la prééclampsie, suggérant un rôle central de ce dernier dans la maladie. Le placenta étant dépourvu d’innervation autonome, le tonus vasculaire placentaire doit être sous le contrôle de facteurs humoraux et tissulaires. Les vaisseaux placentaires sont très réactifs aux prostanoïdes. Le rapport thromboxane A2 (TXA2)/prostacycline (PGI2) est fortement augmenté dans les placentas de grossesses avec prééclampsie. De plus, le taux d’isoprostane, marqueur du stress oxydatif, est accru dans les placentas de femmes avec prééclampsie. Finalement, la prééclampsie s’accompagne d’un stress oxydatif placentaire marqué. Les espèces réactives de l’oxygène sont connues d’une part, pour oxyder l’acide arachidonique (AA), formant ainsi des isoprostanes et d’autre part, pour augmenter la production de TXA2 dans différents tissus, suite à l’activation des cyclooxygénases (COXs). Nous proposons que : 1. les prostanoïdes sont parmi les molécules endogènes qui contrôlent le tonus vasculaire placentaire. 2. la maladie modifie la réponse aux isoprostanes dans les vaisseaux placentaires. 3. l’induction d’un stress oxydatif placentaire entraîne une réponse vasoactive par activation de la voie du métabolisme de l’AA. Nous avons tout d’abord montré, dans des placentas obtenus de grossesses normotensives, que l’U-46619, un mimétique de la TXA2, de même que l’isoprostane, 8-iso-prostaglandine E2 (8-isoPGE2), ont augmenté fortement la pression de perfusion dans les cotylédons perfusés in vitro et la tension dans les anneaux d’artères chorioniques suspendus dans des bains à organe isolé. En revanche, dans les artères chorioniques de placentas obtenus de grossesses avec prééclampsie, ces réponses étaient modifiées puisque la réponse maximale à l’U-46619 était augmentée et celle à la 8-isoPGE2 diminuée. D’autre part, nous avons montré que les réponses maximales aux deux prostanoïdes étaient augmentées dans les vaisseaux placentaires de grossesse normale ou avec prééclampsie issus d’une délivrance prématurée par rapport à ceux d’une délivrance à terme. Ceci suggère une évolution de la réactivité des artères placentaires au cours du 3e trimestre de grossesse. En outre, les vaisseaux placentaires ont répondu aux prostanoïdes de façon semblable qu’ils aient été issus d’un accouchement vaginal ou d’une césarienne élective. Ceci indique que les prostanoïdes placentaires n’interviennent pas dans le processus de délivrance. D’un autre côté, l’utilisation de bloqueurs spécifiques des récepteurs TP à la TXA2, le SQ29,548 et l’ICI192,605, et des récepteurs EP à la prostaglandine E2, l’AH6809, nous ont permis de mettre en évidence le fait que l’U-46619 et la 8-isoPGE2 pouvaient agir de façon non-sélective sur l’un ou l’autre des récepteurs. Ces résultats supportent donc nos 2 premières hypothèses : les prostanoïdes font partie des molécules endogènes qui peuvent contrôler le tonus vasculaire placentaire et la prééclampsie modifie la réponse aux isoprostanes dans les artères chorioniques d’une manière compatible avec l’augmentation de la production de ces substances qui elle, est probablement le résultat du stress oxydatif. En revanche, en ce qui concerne les substances capables de jouer la contrepartie vasodilatatrice, l’utilisation d’un inhibiteur des synthases de monoxyde d’azote, le L-NAME, et celle d’inhibiteurs des COXs, l’ibuprofène, l’indométacine et le N-2PIA, ne nous a pas permis de mettre en évidence un quelconque rôle du monoxyde d’azote ou des prostanoïdes vasodilatatrices à ce niveau. Finalement, nous avons montré que l’induction d’un stress oxydatif dans les cotylédons perfusés in vitro et les artères chorioniques entraînait une vasoconstriction marquée. Celle-ci semble résulter de l’action des prostanoïdes puisqu’un blocage des récepteurs TP ou des COXs diminuait significativement la réponse maximale au peroxyde d’hydrogène. Les prostanoïdes impliquées dans la réponse au stress oxydatif proviendraient essentiellement d’une activation des COXs puisque l’étude ne nous permet pas de conclure à une quelconque implication des isoprostanes dans cette réponse. Ces observations confirment donc notre hypothèse que, dans le placenta, le stress oxydatif possède des propriétés vasoactives par activation du métabolisme de l’AA. En résumé, les résultats obtenus dans les placentas de grossesses normotensives et avec prééclampsie suggèrent que les prostanoïdes sont des molécules d’importance dans la régulation du tonus vasculaire placentaire. Le fait que la prééclampsie modifie la réponse aux prostanoïdes pourrait expliquer pourquoi la perfusion placentaire est altérée chez ces patientes. En outre, il apparaît évident qu’il existe un lien étroit entre le stress oxydatif et la voie de synthèse des prostanoïdes placentaires. Cependant d’autres études sont nécessaires pour mieux comprendre la nature de ce lien, qui pourrait, d’une certaine façon, jouer un rôle important dans le développement de la prééclampsie. / Throughout pregnancy, appropriate placental perfusion is essential for the fœtus to grow properly. In disease such as preeclampsia, placental perfusion is impaired, compromising the fœtus and mother’s lives. Placenta delivery leads to a complete disappearance of the clinical symptoms of preeclampsia. This suggests that the placenta plays a central role in the disease. Placenta being devoid of autonomous innervation, placental vascular tone needs to be under the control of humoral and tissular factors; placental arteries are very reactive to prostanoids. The thromboxane A2 (TXA2)/prostacyclin (PGI2) ratio is increased in placenta from preeclamptic women. Furthermore, in placenta from preeclamptic pregnancies, isoprostane rate is increased, which is a marker of oxidative stress. Finally, preeclampsia is characterised by an important oxidative stress. Reactive oxygen species are known to form isoprostane through the oxidation of arachidonic acid (AA) and to increase TXA2 production in various tissues following an activation of the cyclooxygenases (COXs). We postulate that: 1. prostanoids are among the endogenous molecules that control placental vascular tone. 2. preeclampsia alters responses to isoprostanes in placental vessels. 3. induced placental oxidative stress leads to vasoactive responses through the activation of the AA metabolism. We first showed in placentas from normotensive pregnancies that the TXA2 mimetic U-46619 and the isoprostane 8-isoprostaglandin E2 (8-isoPGE2) markedly increased perfusion pressure in in vitro perfused cotyledons, as well as tension in isolated chorionic arteries. However, in placentas obtained from women with preeclampsia, those responses were altered in chorionic arteries. Indeed, maximal response to U-46619 was raised by preeclampsia, while the one to 8-isoPGE2 was decreased. We then showed that preterm delivery increased maximal responses to both prostanoids compared to term delivery. This observation suggests that placental arteries reactivity evolves along the 3rd trimester of pregnancy. Nevertheless, it appeared that delivery mode had no effect on vascular responses to prostanoids, suggesting that placental prostanoids are not involved in the delivery process. The use of specific blockers of the TXA2 TP receptors, SQ29,548 and ICI192,605, and of the prostaglandin E2 EP receptors, AH6809, revealed that U-46619 and 8-isoPGE2 could mediate their effects by acting on both receptors in a non-selective manner. Therefore, these results support our two first postulates: prostanoids could be the endogenous substances controlling the placental vascular tone and preeclampsia alters responses to isoprostanes in chorionic artery rings in a way compatible with the increased production of these substances possibly through the associated oxidative stress. Moreover, we were unable to identify any vasodilator substances capable of counteracting the effects of vasoconstrictors in the placental circulation. Indeed, blocker of nitric oxide synthases, L-NAME, as well as blockers of COXs, ibuprofen, indometacin and N-2PIA, did not reveal any effect of nitric oxide and vasodilator prostanoids at this level. Finally, we showed that induction of oxidative stress in in vitro perfused cotyledons and in isolated chorionic artery rings led to marked vasoconstriction. This would result from the action of prostanoids since a blockade of TP receptors or COXs significantly decreased maximal response to hydrogen peroxide. Prostanoids involved in this response would essentially come from COX activation. Indeed, the present results did not show any concrete involvement of isoprostane substances in the response to oxidative stress. Consequently, these observations confirm our hypothesis that, in the placenta, oxidative stress presents some vasoactive properties through the activation of the AA metabolism. In summary, results obtained in placentas from normotensive and preeclamptic pregnancies suggest that prostanoids are important in the regulation of the placental vascular tone. Furthermore, responses to prostanoids in chorionic arteries are altered by preeclampsia, which could explain why the placental perfusion is impaired in the disease. Moreover, it seems clear that there is a close relationship between oxidative stress and synthesis of placental prostanoids. However, more investigations are needed to better understand the nature of this relationship, which, in some way, could play an important role in the development of preeclampsia.

Au cours de la grossesse, une perfusion placentaire adéquate est indispensable au bon développement du fœtus. Dans certaines maladies comme la prééclampsie, celle-ci est altérée, compromettant ainsi la vie du fœtus, mais aussi celle de sa mère. Le retrait du placenta mène à la disparition des symptômes de la prééclampsie, suggérant un rôle central de ce dernier dans la maladie. Le placenta étant dépourvu d’innervation autonome, le tonus vasculaire placentaire doit être sous le contrôle de facteurs humoraux et tissulaires. Les vaisseaux placentaires sont très réactifs aux prostanoïdes. Le rapport thromboxane A2 (TXA2)/prostacycline (PGI2) est fortement augmenté dans les placentas de grossesses avec prééclampsie. De plus, le taux d’isoprostane, marqueur du stress oxydatif, est accru dans les placentas de femmes avec prééclampsie. Finalement, la prééclampsie s’accompagne d’un stress oxydatif placentaire marqué. Les espèces réactives de l’oxygène sont connues d’une part, pour oxyder l’acide arachidonique (AA), formant ainsi des isoprostanes et d’autre part, pour augmenter la production de TXA2 dans différents tissus, suite à l’activation des cyclooxygénases (COXs). Nous proposons que : 1. les prostanoïdes sont parmi les molécules endogènes qui contrôlent le tonus vasculaire placentaire. 2. la maladie modifie la réponse aux isoprostanes dans les vaisseaux placentaires. 3. l’induction d’un stress oxydatif placentaire entraîne une réponse vasoactive par activation de la voie du métabolisme de l’AA. Nous avons tout d’abord montré, dans des placentas obtenus de grossesses normotensives, que l’U-46619, un mimétique de la TXA2, de même que l’isoprostane, 8-iso-prostaglandine E2 (8-isoPGE2), ont augmenté fortement la pression de perfusion dans les cotylédons perfusés in vitro et la tension dans les anneaux d’artères chorioniques suspendus dans des bains à organe isolé. En revanche, dans les artères chorioniques de placentas obtenus de grossesses avec prééclampsie, ces réponses étaient modifiées puisque la réponse maximale à l’U-46619 était augmentée et celle à la 8-isoPGE2 diminuée. D’autre part, nous avons montré que les réponses maximales aux deux prostanoïdes étaient augmentées dans les vaisseaux placentaires de grossesse normale ou avec prééclampsie issus d’une délivrance prématurée par rapport à ceux d’une délivrance à terme. Ceci suggère une évolution de la réactivité des artères placentaires au cours du 3e trimestre de grossesse. En outre, les vaisseaux placentaires ont répondu aux prostanoïdes de façon semblable qu’ils aient été issus d’un accouchement vaginal ou d’une césarienne élective. Ceci indique que les prostanoïdes placentaires n’interviennent pas dans le processus de délivrance. D’un autre côté, l’utilisation de bloqueurs spécifiques des récepteurs TP à la TXA2, le SQ29,548 et l’ICI192,605, et des récepteurs EP à la prostaglandine E2, l’AH6809, nous ont permis de mettre en évidence le fait que l’U-46619 et la 8-isoPGE2 pouvaient agir de façon non-sélective sur l’un ou l’autre des récepteurs. Ces résultats supportent donc nos 2 premières hypothèses : les prostanoïdes font partie des molécules endogènes qui peuvent contrôler le tonus vasculaire placentaire et la prééclampsie modifie la réponse aux isoprostanes dans les artères chorioniques d’une manière compatible avec l’augmentation de la production de ces substances qui elle, est probablement le résultat du stress oxydatif. En revanche, en ce qui concerne les substances capables de jouer la contrepartie vasodilatatrice, l’utilisation d’un inhibiteur des synthases de monoxyde d’azote, le L-NAME, et celle d’inhibiteurs des COXs, l’ibuprofène, l’indométacine et le N-2PIA, ne nous a pas permis de mettre en évidence un quelconque rôle du monoxyde d’azote ou des prostanoïdes vasodilatatrices à ce niveau. Finalement, nous avons montré que l’induction d’un stress oxydatif dans les cotylédons perfusés in vitro et les artères chorioniques entraînait une vasoconstriction marquée. Celle-ci semble résulter de l’action des prostanoïdes puisqu’un blocage des récepteurs TP ou des COXs diminuait significativement la réponse maximale au peroxyde d’hydrogène. Les prostanoïdes impliquées dans la réponse au stress oxydatif proviendraient essentiellement d’une activation des COXs puisque l’étude ne nous permet pas de conclure à une quelconque implication des isoprostanes dans cette réponse. Ces observations confirment donc notre hypothèse que, dans le placenta, le stress oxydatif possède des propriétés vasoactives par activation du métabolisme de l’AA. En résumé, les résultats obtenus dans les placentas de grossesses normotensives et avec prééclampsie suggèrent que les prostanoïdes sont des molécules d’importance dans la régulation du tonus vasculaire placentaire. Le fait que la prééclampsie modifie la réponse aux prostanoïdes pourrait expliquer pourquoi la perfusion placentaire est altérée chez ces patientes. En outre, il apparaît évident qu’il existe un lien étroit entre le stress oxydatif et la voie de synthèse des prostanoïdes placentaires. Cependant d’autres études sont nécessaires pour mieux comprendre la nature de ce lien, qui pourrait, d’une certaine façon, jouer un rôle important dans le développement de la prééclampsie. / Throughout pregnancy, appropriate placental perfusion is essential for the fœtus to grow properly. In disease such as preeclampsia, placental perfusion is impaired, compromising the fœtus and mother’s lives. Placenta delivery leads to a complete disappearance of the clinical symptoms of preeclampsia. This suggests that the placenta plays a central role in the disease. Placenta being devoid of autonomous innervation, placental vascular tone needs to be under the control of humoral and tissular factors; placental arteries are very reactive to prostanoids. The thromboxane A2 (TXA2)/prostacyclin (PGI2) ratio is increased in placenta from preeclamptic women. Furthermore, in placenta from preeclamptic pregnancies, isoprostane rate is increased, which is a marker of oxidative stress. Finally, preeclampsia is characterised by an important oxidative stress. Reactive oxygen species are known to form isoprostane through the oxidation of arachidonic acid (AA) and to increase TXA2 production in various tissues following an activation of the cyclooxygenases (COXs). We postulate that: 1. prostanoids are among the endogenous molecules that control placental vascular tone. 2. preeclampsia alters responses to isoprostanes in placental vessels. 3. induced placental oxidative stress leads to vasoactive responses through the activation of the AA metabolism. We first showed in placentas from normotensive pregnancies that the TXA2 mimetic U-46619 and the isoprostane 8-isoprostaglandin E2 (8-isoPGE2) markedly increased perfusion pressure in in vitro perfused cotyledons, as well as tension in isolated chorionic arteries. However, in placentas obtained from women with preeclampsia, those responses were altered in chorionic arteries. Indeed, maximal response to U-46619 was raised by preeclampsia, while the one to 8-isoPGE2 was decreased. We then showed that preterm delivery increased maximal responses to both prostanoids compared to term delivery. This observation suggests that placental arteries reactivity evolves along the 3rd trimester of pregnancy. Nevertheless, it appeared that delivery mode had no effect on vascular responses to prostanoids, suggesting that placental prostanoids are not involved in the delivery process. The use of specific blockers of the TXA2 TP receptors, SQ29,548 and ICI192,605, and of the prostaglandin E2 EP receptors, AH6809, revealed that U-46619 and 8-isoPGE2 could mediate their effects by acting on both receptors in a non-selective manner. Therefore, these results support our two first postulates: prostanoids could be the endogenous substances controlling the placental vascular tone and preeclampsia alters responses to isoprostanes in chorionic artery rings in a way compatible with the increased production of these substances possibly through the associated oxidative stress. Moreover, we were unable to identify any vasodilator substances capable of counteracting the effects of vasoconstrictors in the placental circulation. Indeed, blocker of nitric oxide synthases, L-NAME, as well as blockers of COXs, ibuprofen, indometacin and N-2PIA, did not reveal any effect of nitric oxide and vasodilator prostanoids at this level. Finally, we showed that induction of oxidative stress in in vitro perfused cotyledons and in isolated chorionic artery rings led to marked vasoconstriction. This would result from the action of prostanoids since a blockade of TP receptors or COXs significantly decreased maximal response to hydrogen peroxide. Prostanoids involved in this response would essentially come from COX activation. Indeed, the present results did not show any concrete involvement of isoprostane substances in the response to oxidative stress. Consequently, these observations confirm our hypothesis that, in the placenta, oxidative stress presents some vasoactive properties through the activation of the AA metabolism. In summary, results obtained in placentas from normotensive and preeclamptic pregnancies suggest that prostanoids are important in the regulation of the placental vascular tone. Furthermore, responses to prostanoids in chorionic arteries are altered by preeclampsia, which could explain why the placental perfusion is impaired in the disease. Moreover, it seems clear that there is a close relationship between oxidative stress and synthesis of placental prostanoids. However, more investigations are needed to better understand the nature of this relationship, which, in some way, could play an important role in the development of preeclampsia.