Le développement dune placentation hémochoriale requiert une invasion trophoblastique de lendomètre maternel jusquà ses couches profondes ainsi quune érosion des vaisseaux maternels jusquau niveau du tiers supérieur du myomètre. Les cellules trophoblastiques envahissent et modifient les vaisseaux utérins pour permettre une augmentation du flux sanguin vers lunité foeto-placentaire. Dautre part, lembryon qui simplante va non seulement se connecter aux vaisseaux maternels mais aussi générer des vaisseaux de novo. Le processus dangiogenèse endométriale au niveau du site dimplantation est modulé spatio-temporellement par un dialogue complexe entre des facteurs endocrines, paracrines et autocrines. Parmi ces facteurs, lhCG, sécrétée précocement par le trophoblaste, va tenir un rôle de choix. Outre son rôle lutéotrophique bien décrit, lhCG promeut linvasion trophoblastique et présente une action clé sur la tolérance maternelle de lembryon. Nos travaux ont mis en évidence un rôle direct de lhCG sur langiogène endométriale ainsi quun rôle indirect via une boucle paracrine par activation du récepteur hCG/LH à la surface des cellules endothéliales et épithéliales de lendomètre et une stimulation de lexpression du VEGF dans les cellules épithéliales sous linfluence de lhCG. Grâce à de nombreux modèles dangiogenèse in vitro, in vivo et ex-vivo, nous avons décrypté la signalisation cellulaire mise en uvre lors de la liaison de lhCG à son récepteur. De plus, nos travaux démontrent un rôle artériogène inattendu de cette hormone. Ceci revêt toute son importance dans létude de leffet angiogène de lhCG lié à des processus physiologiques comme langiogenèse au niveau du site dimplantation, pathologiques comme leffet tumorigène de lhCG sécrété en fortes quantités lors de pathologies trophoblastiques.

Galectins are proteins with 15 known members found in nearly all living organisms. They share a conserved CRD that binds beta-galactoside sugars, and functions to cross-link glycoproteins as well as glycolipid receptors on the surface of cells to initiate biological responses. Functional studies on the extracellular and intracellular roles of galectins implicate them in cell adhesion, chemoattraction and migration as well as growth, differentiation and apoptosis. Therefore, studies were conducted to identify functional roles of galectin 15 (LGALS15) during the periimplantation period of pregnancy in the sheep. The first study was designed to develop and characterize primary ovine trophectoderm cell lines for the study of the biological functions of LGALS15. Once characterized, these cell lines were used to investigate the role of LGALS15 in trophectoderm gene expression, development, growth, and survival. Two primary trophectoderm cell lines (oTr1 and oTrF) were developed, and they had characteristics similar to in vivo conceptus trophectoderm relative to gene expression, morphology, and migration and proved suitable as an in vitro model to investigate functional roles of LGALS15. The second study investigated LGALS15 function in trophectoderm cell adhesion. A dose-dependent increase in oTr cell attachment to LGALS15 was found that could be inhibited by cyclic GRGDS, but not GRADS, peptides. Mutation of the LDVRGD integrin binding sequence of LGALS15 to LADRAD decreased its ability to promote oTr cell attachment, whereas mutation of the CRD had little effect. LGALS15 induced formation of robust focal adhesions in oTr cells that were abolished by mutation of the LDVRGD sequence. The third study tested the hypothesis that LGALS15 is a secreted regulator of trophectoderm development and gene expression, as well as growth, migration, and apoptosis of trophoblast. LGALS15 moderately increased cellular proliferation, partially inhibited staurosporine elicited apoptosis, stimulated migration that was dependent on Jun N-terminal kinase (JNK), and initiated differential gene expression of oTr cells. Collectively, these results support the hypothesis that LGALS15 has a biological role in the peri-implantation stage of early pregnancy in the ovine uterus and stimulates trophectoderm cell gene expression, migration and attachment via integrin binding and activation which are critical to blastocyst elongation and implantation.

Recurrent early pregnancy loss in the ovine uterine gland knockout (UGKO) ewe model manifests on Day 14 of pregnancy, indicating that endometrial secretions are critical for peri-implantation conceptus development. Therefore, the following studies were conducted with fertile ewes and infertile UGKO ewes to identify candidate endometrial factors essential for normal conceptus survival, utilizing both genomics and proteomics approaches. The first study used transcriptional profiling of endometrium from Day 14 cyclic, pregnant, and bred UGKO ewes, as well as ewes treated with interferon tau (IFN??) and progesterone, to identify genes important for conceptus development. A number of novel and previously known IFN??-stimulated genes, as well as progesterone-stimulated genes were identified that are higher in fertile ewes, such as galectin-15. Interactive effects of progesterone and IFN?? regulate endometrial gene expression in a temporal and cell-type specific manner. The second study characterized the endometrial expression and hormonal regulation of galectin-15, a member of the galectin family of secreted ??-galactoside lectins. Galectin-15 was secreted into the uterine lumen by the lumenal (LE) and superficial glandular epithelium (sGE), where it may promote adhesion during implantation, as well as was phagocytosed by the trophectoderm and formed intracellular crystals. The third study determined the endometrial expression of galectin-15 throughout gestation. Galectin-15 was secreted into the uterine lumen, where it was phagocytosed by the trophectoderm/chorion, transferred through placental vasculature to the fetus, and cleared through the fetal kidney to be stored in allantoic fluid. The fourth study utilized proteomic analysis of uterine flushes and endometrial explant cultures from Day 14 cyclic, pregnant and UGKO ewes to identify differences in uterine secretions. Analyses identified several genes that were expressed by the LE and sGE and may be involved in prostaglandin production and/or pH regulation. Collectively, results of these studies suggest that transcriptional profiling and analysis of uterine secretions are effective tools to determine genes important for early pregnancy. Further, identified genes are expected to reveal novel endometrial factors and metabolic pathways for support of conceptus survival and implantation, as well as provide improvements for embryo culture methods and diagnose endometrial dysfunctions leading to infertility.

Progesterone is unequivocally required for the establishment and maintenance of pregnancy in all mammals studied. Its known functions are complex and encompass global changes in gene expression. Therefore, studies were conducted to characterize the effects of progesterone on expression of genes for endometrial factors having roles in conceptus growth, implantation and establishment of pregnancy. The first study characterized the effect of an artificially induced early increase in circulating progesterone on conceptus growth and development and regulation of expression of galectin-15 (LGALS15), a recently identified protein secreted by the ovine uterine luminal epithelium (LE). Exogenous progesterone beginning on Day 1.5 post-mating accelerated conceptus development on Days 9 and 12. On Day 12 the conceptus was functionally and morphologically advanced to produce greater quantities of interferon tau (IFNT) than blastocysts from control ewes. Further, the endometrium responded to early progesterone and IFNT with early expression of cathepsin L (CTSL), radical S-adenosyl methionine domain containing 2 (RSAD2), and LGALS15 within the endometrium. The second study identifed structural changes within the luminal epithelium which could alter the flux of factors into and out of the uterine lumen to maintain appropriate fetal/maternal communication. In this study, progesterone reduced quantities of proteins associated with both tight and adherens junctions during the elongation period. IFNT subsequently increased these proteins after conceptus elongation. The third and fourth studies identified progesterone-regulated genes which have been implicated as having importance to implantation in sheep, mouse, and human. WNT signaling was transiently downregulated by progesterone, while members of several growth factor families are upregulated including insulin-like growth factor binding proteins (IGFBPs) 1 and 3, hepatocyte growth factor (HGF) and fibroblast growth factor 7 (FGF7), which may enhance conceptus growth. Collectively, these studies assess the role of progesterone in altering gene uterine expression to establish a favorable environment for conceptus development. The long-term goals of these studies are to establish biomarkers of receptivity to conceptus development and implantation, enhance our understanding of gene and pathway regulation in early pregnancy loss, and identify genes which may be targeted in therapeutic strategies to improve reproductive success in humans and animals.

Dissertation--Göttingen--Georg-August-Universität, 2006. / Bibliogr. p. 245-269.

Thesis (Ph. D.)--University of Texas at Austin, 2001. / Vita. Includes bibliographical references. Available also from UMI/Dissertation Abstracts International.

Habilitation - Universität, Mainz, 1998.

published_or_final_version / Medicine / Master / Doctor of Medicine

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