CORTICOSTEROID METABOLISM IN JUVENILE HYPERTENSION

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The goals of this project are to elucidate the structure, properties, and biochemical functions of 11 beta-hydroxysteroid dehydrogenase, an enzyme system composed of 11 beta-dehydrogenase and 11-oxoreductase components. This system plays a key role in corticosteroid metabolism. We anticipate that successful accomplishment of this program will provide insight into how the 11 beta-hydroxysteroid dehydrogenase complex participates in blood pressure regulation, how disturbances in its function are related to a form of juvenile hypertension (AME), and how, through catalysis of corticosteroid metabolism, it mediates cell and tissue growth and maturation. We will use antibodies against the homogeneous 11 beta-dehydrogenase (11 beta DH) component generated in rabbits and pharmacological inhibitors of 11 beta DH, such as glycyrrhetinic acid, to test the hypothesis that the enzyme "protects" type I receptors against the mineralocorticoid activity of 11 beta- hydroxycorticosteroids by oxidizing the 11 beta-hydroxysteroids to the inactive 11-oxo forms. Studies will be performed in vitro in order to explore the molecular basis for the inhibition, and in vivo in order to develop an animal model of AME. We have determined that 11 beta DH is a glycoprotein. We will examine its glycoside structure, and the significance of the carbohydrate component for enzyme function. We will use monospecific polyclonal antibodies we have generated with homogeneous 11 beta DH to screen rat liver cDNA expression libraries incorporated into bacteriophage vectors. Our goal is to determine the nucleotide sequence of the mRNA derived from the structural gene and to deduce from this the primary amino acid sequence of 11 beta DH. We will isolate enzyme from human placenta and compare its properties with the clonally derived polypeptide and the enzyme of murine origin. The antibodies and cDNAs generated in our experiments will be used as histochemical probes to quantitate the distribution of 11 beta DH antigen and its associated mRNA within cells and tissues, and to monitor changes in enzyme level in response to hormonal intervention. We plan to purify corticosteroid 11-oxoreductase, the enzyme that catalyzes the reduction of cortisone to cortisol, and which cooperates with 11 beta DH in maintaining steroid homeostasis. Subsequent to purification, analytical studies similar to those planned for 11 beta DH will be performed. These investigations will help us understand a) how deficiency of 11 beta DH activity leads to hypertension; b) the physiological mechanism that control oxidation and reduction of corticosteroid at C-11.