THE ANATOMICAL RECORD 223:72-81(1989) zy Distribution of Fibronectins and Laminin in the Early Pig Embryo VERONIQUE RICHOUX, THIERRY DARRIBERE, JEAN-CLAUDE BOUCAUT, JACQUES-EDMOND FLECHON, AND JEAN-PAUL THIERY zyxw Laboratoire de Biologie Exptrimentale, Universitt Pierre et Marie Curie, Paris (V. R., zyxw TD., J-C.B.), Unitt de Biologie du Dtveloppement; Institut National de la Recherche Agronomique, Jouy en Josas (V.R., J.-E.F.); and Ecole Normale Suptrieure, Laboratoire de Pathologie zyxw du Dtueloppement, Paris (J.2 T), France ABSTRACT Fibronectins (FN) and laminin (LN) distributions were studied in the pig embryo by indirect immunofluorescence using antiporcine FN and antimu- rine LN antibodies. Extracellular FN are first detected in the early blastocyst before endodermal cell migration. They appear between the cells and on the blastocoelic face of the inner cell mass; thus, they are located at the interface of the trophectoderm and extraem- bryonic endoderm. Mesodermal cells migrate in a tridimensional network of fibrillar FN. These glycoproteins are also in the extraembryonic membranes (chorion and yolk sac wall) contiguous to the FN-rich basement membranes of embryonic ecto- derm and endoderm. Extracellular LN appears in the blastocyst when the endoderm is already estab- lished as a continuous cellular monolayer, and is located between the trophectoderm and the extraembryonic endoderm, which produces it. Laminin also accumulates at the basal surface of the embryonic ectoderm at the onset of gastrulation. In the extraembryonic membranes, LN appears at the interface of the endoderm and mesoderm and at the interface of the trophectoderm and mesoderm. It is produced and secreted by extraembryonic mesodermal cells. Analysis of the distribution of these glycoproteins suggests that FN allow the migration of endodermal and mesodermal cells by providing them with a suitable substrate. When these cells become immobilized, they synthesize LN, probably to stabilize their interactions with the underlying extracellular material and epithelia. The early mammalian blastocyst is organized as a core of inner cell mass (ICM) surrounded by a single layer of trophectoderm. Soon after the emergence of these two cell types, primitive endodermal cells appear on the inner face of the ICM. Although it was observed in all species, the endodermal differentiation has been studied especially in the mouse blastocyst (Gardner, 1983; Ro- senstrauss et al., 1983). It is generally accepted that the primitive endoderm gives rise to either visceral or pari- etal extraembryonic endoderm. These two endodermal cell types differ in location and in morphological and biochemical properties (Enders et al., 1978; Hogan and Newman, 1984). Visceral endodermal cells, joined by intercellular apical junctions, form a continuous epithe- lium that covers the egg cylinder and produces alpha- fetoprotein (Dziadek and Timpl, 1985). Parietal endo- dermal cells resemble flattened fibroblasts, with multi- ple filopodia on the inner face of the trophectoderm. These cells have an important granular endoplasmic reticulum and synthesize most of the elements of the Reichert’s membrane, i.e., laminin, type zyxwvu IV collagen, entactin, and heparan sulfate proteoglycan (Hogan et al., 1984; Howe and Solter, 1980; Jetten et al., 1979). In ungulate embryos, however, the extraembryonic endoderm seems to form an homogenous cellular mono- zyxwv 0 1989 ALAN R. LISS, INC. layer. Primitive endodermal cells appear on the inner face of the ICM (on the 7th day of development in the pig blastocyst) and then migrate on the inner face of the trophectoderm, covering it by a continuous monolayer of flattened extraembryonic endodermal cells (Stroband et al., 1984).The biochemical properties of this extraem- bryonic endoderm are still unknown. An extracellular material has been visualized by electronic microscopy between the trophectoderm and the extraembryonic en- doderm; the cells responsible for its synthesis are unknown. From day 11 on in the pig embryo, the bilayered tro- phoblast undergoes a dramatic phase of elongation so that the initially spherical expanded blastocyst becomes tubular and finally filamentous (> 100 mm long) on days 13-14. Geisert et al. (1982) noticed some modifica- tions in the localization of endodermal cells during this period. Nevertheless, the mechanisms involved in this extensive elongation have not yet been considered at the cellular and molecular levels in these species. The epi- Received January 5, 1988; accepted May 3, 1988. Address reprint requests to V. Richoux, Laboratoire de Biologie Experimentale, U.A. CNRS 1135, Universite P. et M. Curie, Bgtiment C.30,9, quai Saint Bernard, 75005 Paris, France.