SERUM PROTEIN SYNTHESIS IN THE EXTRAEMBRYONIC ENDODERM OF THE EARLY CHICK EMBRYO

摘要

The basis for this dissertation rests on a model which suggests that specific developmental events on the chick are regulated by the relative synthesis and circulating concentration of serum protein. This model is based on observations that the relative synthesis of serum protein changes with development and in response to changes in nutrition. The objective of this dissertation was to study in detail the portion of the model concerned with serum protein synthesis by the yolk sac of the early chick embryo.;In order to investigate serum protein synthesis by the yolk sac the precise germ layer or layers involved were determined. Four day yolk sacs were separated into component ectoderm, mesoderm, and endoderm layers and incubated in the presence of {('14)C}valine while for the intact yolk sac {('3)H} valine was used. When media from intact yolk sac and cell layers were co-electrophoresed within a single gel it was clear that only the endoderm synthesized all four serum proteins, transferin, (alpha) globulin-a, (alpha) globulin-(beta) and prealbumin. Immunoprecipitation of the incubation media confirmed that the endoderm layer exclusively was involved in serum protein synthesis.;When endoderm cells from the area vasculosa and area vitellina were isolated and cultured separately only endoderm cells from the area vitellina were found capable of cell attachment and migration in culture. Thus, only vitellina endoderm cells could be cultured. Eagle's minimal essential medium plus 10% fetal calf serum proved to be "optimum" medium for the culture of these cells. During culture DNA synthesis dropped to background levels.;Cultures of vitellina endoderm cells were analyzed for their ability to synthesize and secrete serum protein for a period of six days from the start of culture. For 4 of 5 serum proteins, changes in relative synthesis parallelled precisely those which occurred in vivo. One serum protein, (alpha)globulin-a, was not synthesized after 1 day of culture even though in vivo its synthesis increased dramatically. During this culture period the absolute quantity of serum protein per total protein synthesized and secreted did not change and remained approximately 50%.;Cultured vitellina endoderm cells were supplemented with conalbumin and ovalbumin to determine whether endoderm cells in vitro would respond in a manner comparable to the intact yolk sac. In such cultures the amount of serum protein per total protein synthesized and secreted did not differ from control cultures. The relative synthesis of prealbumin was shown to be affected exclusively by ovalbumin. Because this response did not parallel that which takes place in vivo it was suggested that ovalbumin acts in conjunction with other factors in vivo to regulate the synthesis of serum protein by the yolk sac.;Endoderm cells in culture were supplemented with insulin which has been shown to affect serum protein synthesis in adult liver. Such cultures synthesized relatively more prealbumin and less serum albumin compared to control cultures. Because such doses of insulin have been shown to be teratogenic in vivo it was suggested that the effect of this hormone on development may be mediated through changes in the relative synthesis and secretion of serum protein by the yolk sac.