Although progesterone and synthetic progestins are known to effect a number of uterine functions, as yet the molecular mechanisms of action are poorly understood. An organ culture system which employs endometrial fragments has been developed and extensively characterized such that it can now be applied to elucidating the mechanism of progestin action in the mammalian uterus. With this system, glycogen synthesis serves as a precise biologic end point of progestin action. This parameter also allows systematic examination of complex hormonal interactions on the uterus such as the priming effect of estrogen and permissive requirements for insulin. In the culture system, RNA, protein synthesis, polyamine metabolism and membrane transport are all essential components of progestin-induced glycogen synthesis. In the present study, we will identify the best progestin inducible protein for use as a probe to examine the precise biologic effects of the purified progestin receptor in primary cell culture. Specifically, we will systematically evaluate the changes in the activities of 6 enzymes and seek to correlate their activities with the level of nuclear progestin receptor. These enzymes have been identified as possibly regulated by the progestin receptor: Na-K-ATPase, hexokinase, ornithine decarboxylase, S-adenosyl-L-methionine decarboxylase, glycogen synthetase and phosphorylase. Through the use of selective inhibitors we will determine the interrelationship of these key enzymes. In addition, the progestin receptor will be purified using conventional chromatography, an antibody affinity column, and preparative isoelectric focusing. An antibody will be generated in guinea pigs against the receptor and characterized by immunoelectrophoresis. The biologic activity of the purified receptor will be determined by microinjection in primary cell culture. This basic information will facilitate development of alternate means to prevent implantation as a means of contraception and will provide a background for study of the problem of early abortion.
NIH ExPorter
NIH ExPorter