HORMONAL REGULATION OF ANGIOTENSINOGEN GENE

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The long term goal of my laboratory is to characterize the basic mechanisms of blood pressure regulation and blood flow. It is now becoming apparent that tissue specific systems exert a tremendous influence on the regulation of blood pressure and may play a major role in the genesis of hypertension. We have focused on angiotensinogen (angen), important in the circulating renin angiotensin system, but also synthesized in extra-hepatic tissues. It is likely that local generation of angen can have profound physiologic and pathologic effects in the absence of changes in circulating angen. The control of angen expression and the function of local generating systems will hopefully be elucidated by the experiments described. We aim to determine the function of local angen generation. The functional significance of CNS generation will be elucidated by detailed anatomic localization (in situ hybridization and punch biopsy studies) in combination with studies on the physiologic and pharmacologic regulation of angen mRNA levels (quantitative in situ and slot blot hybridization). The possible roles of angen in renal function will be determined by detailed anatomic localization (in situ hybridization) and studies of angen mRNA regulation by relevant physiologic manipulations. Angen may have important autocrine or paracrine influence on blood vessel function. Determination of the cellular origin (endothelium vs. smooth muscle) will be accomplished by in situ hybridization. Regulation will be studied by the quantitative in situ methods developed for brain. We aim to determine the regulation of angen gene using cell culture systems. We will characterize the hormonal regulation (slot blot and nuclear run off) by hormones and growth factors in MA10 Leydig cells and compare the results to those obtained in Reuber H35 cells. Isolation and characterization of the flanking sequences of the angen gene is required for a high level of understanding of tissue specific regulation, thus we will clone the gene encoding rat angen. We then will determine the specific genomic sequences and characterize the cellular specific nuclear factors necessary for angen gene regulation. We will use CAT assays and the gel retention method to gain information about cell specific binding factors and regulatory sequences. Finally we aim to determine the role of growth factors in angen production. Solid tumors as well as developing tissues need adequate blood flow and vascularization for continued growth. Since angiogenesis (growth) factors have now been identified, we aim to determine the role of these factors in the regulation of the angen gene. When angen gene expression is characterized as above, it will become possible to make inferences about the extent to which local generation effects normal function or causes pathologic effects.