Work will continue on the progesterone-induced proteins of the porcine uterus 1) using electron microscopy and immunocytochemical tecniques involving Protein A-gold particcles, we shall define the location of uteroferrin (Uf) and plasmin inhibitor (PI) in the secretory cells of the uterus and determine whether the two proteins follow a similar pathway of secretion. Similarly, we shall demonstrate the localization of these proteins in the absorptive cells of the conceptus at Day 14 and Day 60 of pregnancy in order to understand their movement to the yolk sac and placental venous drainage respectively. Their distribution will be compared with that of immunoglobulins derived from maternal plasma. 2) We shall raise monoclonal antibodies against uterine secretory proteins and use these reagents to purify and immunocytochemically localize minor components of the secretions. 3) The PI will be sequenced and its inhibitory action on plasmin and plasminogen activator (PA) investigated in detail. Because PI may function to protect Uf and other proteins from lysosomal proteolysis, we shall test its ability to inhibit lysosomal proteases. 4) PA from secretions during proestrus will be purified ans immunocytochemically localized in the uterus. We shall determine whether PA (by local formatiom in plasmin) plays a role in effecting the movement of proteins (e.g. Uf) from uterine glands to stroma during luteal regression. 5) A cDNA of UF mRNA will be cloned from a library of cDNAs generated from endometrial mRNA's expressed during mid pregnancy. This cDNA copy of Uf mRNA will be employed to measure the amounts of Uf mRNA during pregnancy, the estrous cycle and after hormone replacement therapy to determine whether progesterone control of Uf synthesis is correlated with levels of its mRNA. Ultimately these experiments will be repeated with other proteins to see if their is a coordinated appearance of the mRNAs for the different progesterone-induced components. 6) We shall investigate the movement of 59Fe supplied as Uf from the reticuloendothelial cells of the fetal liver to erythroblasts in adjacent blood islands using autoradiography. We shall also determine whether there is a change in fetal erythropoiesis around midpregnancy. 7) Finally, we shall define the mechanism whereby estrogen triggers the rapid release of proteins from secretory granules of uterine glandular epithelium, and determine whether the mechanism involves catecholamines and increases in cAMP. These studies will provide an understanding of the function of uterine proteins in pregnancy and how the uterine environment might be manipulated artificially.
