Os roedores murídeos são melhor conhecidos em relação à placentação, porém descrições para outros grupos como os cricetídeos incluindo os \"camundongos do Novo Mundo\" ainda são escassos. Recentemente à placenta e as membranas fetais tem sido considerados fontes promissoras para a obtenção de células-tronco. Em particular, o saco vitelino é estruturalmente diverso e desempenha várias e importantes funções. Por essa razão, o objetivo deste trabalho foi descrever a placentação corioalantóidea e vitelina em Necromys lasiurus. Além de avaliar o potencial do saco vitelino como uma fonte de células-tronco mesenquimais. Para tanto, um total de 10 placentas variando de início ao final de gestação foram analisadas através de técnicas de histologia, imunohistoquímica e microscopia eletrônica, bem como através do cultivo e diferenciação celular, citometria de fluxo e imunocitoquímica. A placenta corioalantóidea discoidal era organizada em uma zona labiríntica, zona juncional e decidua. O labirinto era a região mais importante para as trocas materno-fetal. Próximo ao final da gestação ele apresentou uma barreira hemotricorial com espessura média de 2,41 µm. A zona juncional era composta por sincício e citotrofoblasto. Células trofoblásticas gigantes localizavamse entre a zona juncional e a decidua, assim como nas margens laterais da placenta. A morfologia do saco vitelino visceral invertido variou de acordo com a sua localização e relação com a placenta e o útero. Quando cultivadas, as células aderentes do saco vitelino formaram colônias fibroblastóide (92,13%) e expressaram marcadores de células-tronco mesenquimais e alguns para células precursoras de células-hematopoiéticas. As células apresentaram sucesso quanto às diferenciações osteogênica, adipogênica e condrogênica e não desenvolveram formação tumoral quando injetadas em camundongo nude. Com isso, essas células-tronco despontam como uma fonte terapêutica promissora para a terapia celular. / Murine rodents are well investigated in regard to placentation, but data for other groups such as cricetids including New World mice or Sigmodontinae are sparse. Recently the placenta and fetal membranes are regarded to be promising sources for obtaining stem cells. In particular, the yolk sac is structurally diverse and shows important functions throughout gestation. For this reason, this study aims to describe the chorioallantoic and yolk sac placentation in Necromys lasiurus. In addition, the potential of the yolk sac as a source for mesenchymal stem cells that are not well studied so far will be evaluated. In total, 10 individuals from early gestation to near term were investigated by means of histology, immunohistochemistry and electron microscopy as well as cell culture and differentiation, flow citrometry and immunocytochemistry. The discoidal chorioallantoic placenta was organized in a labyrinth zone, junctional zone, and decidua. The labyrinth was most import for maternal-fetal exchange processes. It possessed a hemotrichorial barrier of about 2.41 µm thickness near term. The junctional zone included syncytial areas and cytotrophoblasts. Trophoblast giant cells were located between the junctional zone and decidua as well as in the lateral margins of the placenta. The morphology of the inverted visceral yolk sac varies according to its location and relationship with the placenta and uterus. When cultured, the adherent cells of the yolk sac formed fibroblastoid colonies (92.13%) and expressed mesenchymal stem cells markers, and some hematopoietic precursor cells markers. They showed successful osteogenic, adipogenic, and chondrogenic differentiation and did not develop tumors when transferred to nude mice. Thus, these cells resulted as stem cells with promising therapeutic values for cell therapy.

O saco vitelino é o único anexo embrionário presente em todas as espécies animais cuja função é manter o desenvolvimento primário do feto até que a placenta assuma esta função. Além disso, ele desempenha funções importantes como: nutrição do embrião, síntese proteica, atividade fagocitária, transferência de materiais e hematopoiese. A utilização de células-tronco embrionárias gera muitas discussões devido às questões éticas associadas à obtenção destas células, além de questões religiosas e também as relações de baixa plasticidade das células-tronco adultas motivam os pesquisadores ao crescente interesse em outras fontes de célulastronco. Com isso, células provenientes de tecidos como as dos anexos embrionários chamam cada vez mais atenção devido à sua facilidade de acesso, crescimento rápido e boa plasticidade. Nosso grupo tem demonstrado que as células do saco vitelino têm boa capacidade de proliferação e são multipotentes. Vários pesquisadores vêm demonstrando que o período no qual as células são obtidas pode ser de fundamental importância para a diferenciação. Sendo assim, neste projeto tivemos como objetivo estabelecer e caracterizar células-tronco de saco vitelino de cães em dois estágios gestacionais, visando analisar de forma comparativa o perfil de expressão de marcadores mesenquimais, hematopoiéticos e pluripotência de forma a entender o comportamento das mesmas. Neste trabalho observamos que as células de saco vitelino obtidas de gestações com 40 e 50 dias aderiram ao plástico, tiveram morfologia fibroblastóide, boa proliferação celular, se diferenciaram em adipócitos, condrócitos e ostéocitos e expressaram alguns marcadores mesenquimais, sugerindo assim ser uma célula-tronco mesenquimal. Além disso, não levaram à formação tumoral quando injetadas em camundongos imunossuprimidos, nudes. / The yolk sac is an embryonic attachment present in all animal species, which function is to keep the primary development of the fetus until the placenta assumes this function. Moreover, it plays important roles as: nutrition of the embryo, protein synthesis, phagocytic activity, transfer of materials and hematopoiesis. The use of embryonic stem cells generates many discussions because of the ethical and religious issues associated with the obtaintion of these cells. Also the relationship of low plasticity of adult stem cells motivates researchers to explore other sources of stem cells. Thus, cells from tissues such as the embryonic anexes call more attention, due to its ease of access, rapid growth and good plasticity. Our group has shown that cells of the yolk sac has good ability to proliferate and are multipotent. Several researchers have shown that the period in which the cells are obtained can be critical for differentiation. Therefore, in this project we aimed to establish and characterize stem cells from dog yolk sac in two stages of pregnancy, in order to analyze comparatively the expression profile of mesenchymal, hematopoietic and plurypotency markers, and study their behavior and plasticity potencial in vitro. In this study we observed that yolk sac cells obtained from pregnancies at 40 and 50 days adhered to plastic, had fibroblast like-morphology, good cell proliferation and differentiated into adipocytes and osteocytes and expressed some mesenchymal markers, therefore suggesting to be a mesenchymal stem cell. Also, they did not lead to tumor formation when injected into immunosuppressed mice, nude.

Calbindin-D28K is a cytosolic calcium binding protein found in a variety of cells that transport calcium. The chorioallantoic membrane and yolk sac of oviparous squamate reptiles (lizards and snakes) transport calcium from the eggshell and yolk to the developing embryo. I used immunohistochemistry to localize calbindin-D28K expression in the chorioallantoic membrane and yolk sac of two species of oviparous scincid lizards, Plestiodon fasciatus and Saproscincus mustelinus. Calbindin-D28K was detected in the chorioallantoic membrane and yolk sac of both lizard species by a polyclonal anti-snake calbindin antibody and a monoclonal anti-cow calbindin antibody. Calbindin-D28K was localized in the chorionic epithelium and allantoic epithelium of the chorioallantoic membrane and in endodermal cells scattered throughout the yolk mass of both species. This is the first demonstration of calbindin-D28K in allantoic epithelium and in endodermal cells of the yolk sac of lizards.

Le problème du sac-à-dos à contraintes disjonctives (DCKP) est une variante du sac-à-dos normal. C'est un problème dans lequel certains objets peuvent être incompatibles avec d'autres. Le DCKP apparait, souvent, comme sous problème d'autres problèmes d'optimisation combinatoire plus complexes.

Chez la souris, peu de choses sont connues sur les marqueurs de surface qui caractérisent les cellules souches hématopoïétiques (CSHs) embryonnaires. Durant ma thèse, j'ai étudié au niveau du placenta (Pl) et du sac vitellin (SV) les populations CD34+c-kithi, CD144+CD45+ et Sca-1+AA4.1+, déjà décrites comme enrichies en CSHs dans d'autres contextes. Le projet dans son ensemble a permis de montrer que l'enrichissement en CSHs n'implique pas les mêmes marqueurs selon le tissu considéré : dans les sites d'émergence (AGM) et d'émergence/amplification des CSHs (SV), la population la plus enrichie en CSHs a pour phénotype CD144+CD45+ ; concernant le Pl, organe d'amplification des CSHs, elle a pour phénotype CD34+c-kithiSca-1+ alors que dans le foie fœtal (FF), organe d'amplification/différenciation des CSHs, elle a pour phénotype CD34+c-kithiSca-1+AA4.1+. L'analyse moléculaire de ces populations permettra de révéler des molécules régulatrices spécifiques de l'émergence et de l'amplification des CSHs. Par ailleurs, nous avons utilisé les souris transgéniques VECR pour étudier l'origine des CSHs CD34+c-kithi du Pl et il semblerait qu'à E11.5, toutes ne proviennent pas de l'endothélium. Les résultats préliminaires réalisés sur les souris Mpl-/-, qui présentent un défaut de contenu en CSHs dans le Pl et le FF, indiquent qu'à E11.5, le potentiel hématopoïétique de la population CD34+c-kithi du Pl Mpl-/- est inférieur à celui de la population CD34+ckithi du Pl C57Bl6 ; cette différence n'est plus visible à E12.5. Le Pl constitue donc une niche transitoire d'amplification/maturation des CSHs mais il est possible qu'il puisse également produire des CSHs

Les composés organiques volatils (COV) sont une catégorie de polluants de l'air dont les techniques d'analyse et méthodes d'échantillonnage peuvent être affectées par l'humidité. Deux voies ont été explorées pour éliminer l'humidité des échantillons de COV avant leur analyse. D'une part, l'élimination de l'humidité par diffusion à travers les parois des sacs d'échantillonnage en film polymère, méthode appelée Sample Water Removal. En plaçant des sacs en Tedlar et Nalophan dans un caisson balayé d'air sec, des échantillons humides de 10 L ont été séchés en respectivement 7 h et 3,5 h. Aucune perte significative de COV jusqu'à 500 ppbv n'a été constatée avec les sacs en Tedlar et 10 µg/m3 avec le sac en Nalophan. D'autre part, l'utilisation de nouveaux adsorbants hydrophobes à base de nanostructures de carbone élaborés au laboratoire ont été testés. Durant les essais d'échantillonnage d'un mélange de COV à 500 ppbv en air humide, ces adsorbants n'ont pas donné les résultats espérés.

The extraembryonic endoderm (XEN) defines the yolk sac, a set of membranes that provide essential support for mammalian embryos. Recently, the committed XENprecursor was identified in the embryonic Inner Cell Mass (ICM) as a group of cells that intermingles with the closely related, anatomically indistinguishable epiblast (EPI)- precursor that gives rise to the fetus. In vitro, the EPI-precursor is represented by the well-known embryonic stem (ES) cell lines, but cell lines representing the XENprecursor are not known. Furthermore, since the XEN-precursor cells were discovered only very recently, the unexpected fact that they express the key pluripotency marker Oct4 has not been explored. Recently, however, our laboratory has isolated rat XEN cell lines that express Oct4, leading to the following two questions: (i) Do these new XEN cell lines represent XEN-precursor cells? (ii) Is their Oct4 expression regulated similarly as previously known from ES cells? These two questions are addressed here by lineage marker and reporter gene analyses. Whole culture analyses showed that rat XEN cell lines expressed markers of all XEN stages including XEN-precursor, primitive endoderm (PrE) and/or visceral endoderm (VE), and parietal endoderm (PE) but trophoectoderm and EPI-precursor markers were missing. In line with this, immunocytochemistry demonstrated heterogeneity and directly visualized the XEN-precursor, PrE/VE, and PE subpopulations. Low-density plating and time-dependent immunocytochemistry on resulting colonies strongly suggested that XEN-precursor cells generate the other XEN stages. Moreover, by analyzing single-cell derived clones, it was shown that culture heterogeneity results from the self-renewal and differentiation of a single cell. Reporter gene analyses using the 5’ regulatory region of the mouse Oct4 gene revealed that a DNA fragment containing the previously described distal enhancer drove reporter gene expression only in ES cells whereas inclusion of an upstream fragment led to high expression in both mouse ES and rat XEN cells. In conclusion, our rat XEN cell lines contain XEN-precursor cells that differentiate extensively, providing for the first time an in vitro model that mimics the natural process of early XEN differentiation. In addition, they regulate Oct4 gene transcription differently than ES cells suggesting heterogeneous Oct4 regulation within the mammalian ICM.

No description available.

Pre-mature hatching of fertilized eggs of cuttlefishes and squids, which are Taiwan¡¦s major fishing species, exists in the late embryonic development before yolk sacs are fully absorbed. It is so far unknown whether there is any difference in survival rate between pre-maturely developed juveniles and the fully developed ones. Hence, by laboratory incubation, this study aimed to discuss the relationship between the yolk size in the embryonic development process and embryo, as well as the difference in survival rate of juveniles developed at different developmental stages. The impact of the incubation time on the survival rate is explored in case of changing physical and chemical environmental factors (temperature, salinity, ammonia concentration, vibration etc.) The embryonic development of Sepia pharaonis can be divided into 40 stages according to the external shape and quality of the embryo. The embryo mantle length and the yolk diameter vary by time, while the increasing rate of the mantle length does not(F = 1.88, p = 0.06), increasing or decreasing in a linear relationship respectively. However, under the same environmental conditions, the consumption rate and the mantle size may vary in different batches(yolk diameter: F = 8.77, p < 0.01. mantle length: F = 92.14, p < 0.01). There is no difference in the surviving time of juveniles artificially and naturally incubated at the same developmental stage, and the surviving time will be longer if the artificially incubated juveniles are at later embryonic developmental stages(F = 34.66¡Ap < 0.01). With regard to the feeding ratio of juveniles pre-maturely hatched at different stages, the feeding ratio of the juveniles incubated after the 36th stage will increase with the developmental stages(F = 93.10¡Ap < 0.01). In this study, the temperature limit of the embryonic development of Sepia pharaonis should never be lower than 10¢J or higher than 35¢J, and the most suitable temperate range is between 17-28¢J. In case of sudden change in temperature, temperature increase can more effectively affect the pre-mature hatching than temperature decrease. In case of either the 36th or 39th stage embryos, if the temperature rises or drops by more than 10 degrees, pre-mature hatching can exist in some of the embryos. Some embryos may die if salinity is lowered suddenly below 20 psu. Meanwhile, pre-mature hatching may occur within one hour if it is suddenly lowered below 10 psu, and the unhatched embryos may die. When increasing the ammonia concentration suddenly to 1 and 5 ppm, the embryo incubation time may be lengthened compared with the group without the addition: some embryos may die if it is increased to 5 ppm. Meanwhile, most embryos are hatched pre-maturely within 17 minutes when it is increased to 1000 ppm, while those unhatched ones may die. In case of various environmental stimuli of the experiments, a higher percentage of embryos at the 39th stage got away from the hostile environment by pre-mature hatching, while a higher percentage of embryos at 36th stage continued the development until natural incubation or died. The vibration experiment is to produce vibration by a vertical vibration instrument. In case of 30-minute vibration at frequency of 350 times/minute and maximum amplitude at 2 cm, there is no effect on incubation time and mode of embryos at both stages. This factor is still open and subject to further discussion.

Je présente dans ce mémoire un bilan de mon activité scientifique effectuée au sein des groupes POC (Parallèlisation et Optimisation Combinatoire) et SID (Systèmes d'Information Distribués) de l'équipe ROI (Recherche Opérationnelle et Informatique) du laboratoire LAMIH à l'UVHC. <br /><br />Mes travaux de recherche se divisent en trois parties:<br /><br /> - l'étude du problème du sac-à-dos non borné, problème classique de l'optimisation combinatoire, dont nous mettons en évidence des propriétés fondamentales et pour lequel nous avons dérivé, implanté et mis à disposition deux algorithmes qui tirent avantage des propriétés découvertes;<br /><br /> - une approche algorithmique parallèle/distribuée pour la<br /> bio-informatique notamment le problème de repliement de protéïnes qui est un problème reconnu comme l'un des plus difficiles posés à la science informatique dans le contexte de la bio-informatique;<br /><br /> - le développement d'une plate-forme ouverte d'expérimentations pour la méthode formelle B, l'étude de la modularité du langage B et d'extensions de B pour générer des composants logiciels ainsi que l'étude de l' adjonction au langage B d'une logique temporelle.<br /> <br /> Nous montrons comment les approches utilisées dans une recherche en optimisation combinatoire d'une part et en spécification formelle d'autre part peuvent s'enrichir et se féconder mutuellement.