Abstract. The purpose of the study was to determine and compare the secretion of RBP by bovine spherical, elongating and filamentous conceptuses, and to identify the cellular location of RBP in developing conceptuses by immmunocytochemistry. Bovine conceptuses were removed from the uterus between days 13 and 22 of pregnancy. Events of early bovine embryonic development were observed. The conceptuses underwent a transformation from a spherical to a filamentous morphology during the periattachment period of placentation. Isolated conceptuses were cultured in a modified minimum essential medium in the presence of radiolabeled amino acids. Presence of retinol-binding protein (RBP) in culture medium was determined by immunoprecipitation using bovine placental anti-RBP serum. Presence of immunoreactive RBP in detectable quantities in spherical blastocyst (day 13) culture medium was evident. Increased amounts of RBP were clearly detected in cultures on days 14 and 15, the time of elongating conceptuses. RBP was abundant in cultures on day 22, when the conceptuses were filamentous. Cellular sources of RBP in day 15 and 22 conceptuses were determined by immunocytochemistry with anti-RBP serum. Strong immunoreactive RBP was localized in trophectoderm of day 15 conceptuses, and in epithelial cells lining the chorion and allantois of day 22 conceptuses. RBP originating from the conceptus may serve to transport retinol locally from the uterus to embryonic tissues. Secretion of RBP by spherical blastocysts (day 13) suggests that retinol is needed for initiating the early events of embryonic development. Increased secretion of RBP on days 14 and 15, the time of elongating conceptuses, was coincident with the timing of rapid trophoblast hyperplasia and remodeling. Greater secretion of RBP by the filamentous conceptus (day 22) coincided with the timing of differentiation of the chorion and allantois as well as growth of the embryo proper. Production of RBP by the developing conceptuses may be indicative of an important role for conceptus RBP in the transport, availability and metabolism of retinol during early pregnancy.

Bovine yolk sac at day 24 of pregnancy, and placental membranes (chorion and allantois) from days 70 and 100 of pregnancy were isolated and cultured in a modified minimum essential medium in the presence of [ 35 S]methionine. Proteins synthesized and secreted by isolated bovine yolk sac, chorion and allantois were analyzed by fluorography of two-dimensional polyacrylamide gel electrophoresis. Serum-like proteins,transferrin, α-fetoprotein, α1-antitrypsin and α1-acid glycoprotein ,were the major protein products of yolk sac. A 21 kDa protein produced by yolk sac was identified immunochemically as retinol-binding protein (RBP). Chorion and allantios from days 70 and 100 of pregnancy were active in protein synthesis and secretion. Both chorion and allantois did not secret serum-like proteins but secreted a number of neutral-to-acidic proteins including RBP. Secretory proteins produced by the yolk sac, chorion and allantois may play important roles in the embryonic development and the successful outcome of pregnancy. Antiserum against bovine placental RBP was employed to the immunocytochemistry by immunoperoxidase method. Immunoreactive RBP was localized in epithelial cells and island-like cell clones of yolk sac. Immunostaining for RBP was detected in simple columnar epithelium of chorion and in simple squamous epithelium of allantois. In the present study, proteins synthesized and secreted by yolk sac at day 24 of pregnancy, chorion and allantois from days 70 and 100 of pregnancy were characterized In addition, RBP was localized in yolk sac, chorion and allantois by immunoperoxidase method. The immunoperoxidase method has been proven to be a very effective technique to identify the cellular source of protein synthesis in extraembryonic membranes.