140 LOCALIZATION OF PRIMORDIAL GERM CELLS IN DAY 21 BOVINE EMBRYOS

Abstract

In most animal species, germ cell precursors, i.e., primordial germ cells (PGCs), arise from the epiblast and then migrate to the future gonadal ridge during development. At least in the mouse, PGCs may be cultured as embryonic germ cells that remain pluripotent. PGCs are the only cells in which OCT4 expression is maintained after gastrulation. The present study aimed at identifying the localization of PGCs in Day 21 in vivo-derived bovine embryos by immunohistochemical staining against OCT4. Six embryos were obtained after slaughter of superovulated heifers 21 days after insemination. The uterine tracts were flushed and embryos fixed, paraffin-embedded, and processed for immunohistochemistry. Embryos were sagitally sectioned, and selected serial sections were immunohistochemically stained for OCT4 to identify potential PGCs. Two embryos were at the neural groove stage. At this stage of development, the primitive gut had not yet been abstricted from the yolk sac and the allantois was not visible. A weak homogeneous OCT4 staining was localized to nuclei in a well-defined region of the epiblast, which was in the process of a gradual anterior to posterior differentiation into neural and surface ectoderm. Moreover, a strong OCT4 staining was localized to a few scattered cells found in the visceral mesoderm associated with the yolk sac in the region of the endoderm-hypoblast transition at some distance from the embryo proper. Four embryos were at the neural tube/somite stage. At this stage of development, the primitive gut had been defined and only the midgut was connected to the yolk sac. Furthermore, the allantois was visible as an anchor-shaped structure at the posterior end of the embryo. A strong OCT4 staining was found in nuclei of solitary cells in the endoderm and its associated visceral mesoderm of the ventral aspect of the mid and hindgut. The described OCT4 staining corresponds well with previous findings in the pig, in which presumptive PGCs are found in the endoderm epithelium during the neural groove stage. Later, during the early somite stages, they are localized in the endoderm and visceral mesoderm of the yolk sac and allantois, and in later somite stages, they are found in the developing genital ridge. This is, however, the first study to demonstrate the localization of these cells, at least by OCT4 staining, in bovine embryos at the neural groove and neural tube/somite stages.