Cellular characteristics of canine trophoblasts Public Deposited

http://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/j098zd38t

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  • This research investigated the development of a novel canine model to study preeclampsia. Normal canine placental development has morphologic and histologic similarities to the shallow trophoblast invasion occurring with preeclampsia in humans, which makes the dog a particularly good choice for modeling this disease and will be an improvement on existing animal models. Preeclampsia is a pregnancy-specific syndrome, occurring in mild (late onset) and severe (early onset) forms. Severe preeclampsia is a major cause of maternal, fetal, and neonatal morbidity and mortality worldwide. It affects 0.35-1.40% of human pregnancies. Despite intense investigation, the cause (and therefore the prevention or treatment) of shallow trophoblast invasion in preeclampsia remains largely unknown. In a normal human pregnancy, trophoblasts invade the endometrium and myometrium as well as the maternal blood vessels (hemochorial placentation). In preeclampsia, trophoblast invasion is shallow and vascular transformation incomplete. In contrast to the normal human placenta, trophoblasts within the canine placenta only invade to the level of the endothelial cells within the maternal blood vessels (endotheliochorial). In this way, normal canine placental development is similar to preeclampsia. The hypothesis of this research was that isolated canine trophoblasts will express similar proteins as human preeclamptic trophoblasts. The objectives of the research were to (1) isolate canine trophoblasts from fresh and cryogenically frozen placenta and (2) perform immunocytochemistry and immunohistochemistry on canine trophoblasts for proteins expressed in human preeclamptic trophoblasts. Cellular morphology was similar to that reported for trophoblasts. More than 97% of the cells cultured expressed cytokeratin-7. Although both matrix metalloproteinases (MMPs) were immunolocalized to the cytoplasm, MMP2 was found in large, coalescing granules, whereas MMP9 was more diffusely expressed throughout the cell. More cultured canine trophoblasts expressed MMP9 (54.7±3.4%) compared to MMP2 (40.3±1.8%) (p=0.02). Cryopreserving placental tissue prior to primary cell culture had no effect on cell proliferation (p=0.37). Relaxin, vascular endothelial growth factor, and tissue inhibitor of metalloproteinase 2 were positively expressed in primary canine trophoblasts. Immunohistochemical results revealed CK-7, MMP9, TIMP2 and relaxin was expressed in trophoblasts along the villous margin with MMP9, TIMP2 and relaxin extending towards the basement membrane. S100A4 was minimally expressed in the basement membrane. MMP2 was strongly expressed within the basement membrane. CK-7, MMP2, MMP9 & TIMP2 were all immunolocalized to the same cells in canine placental sections as previously described in human preeclamptic placental sections. These results have demonstrated the cellular similarities in protein expression between normal canine and human preeclamptic trophoblasts thereby confirming this model is suitable for further studies.
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