Development of a Mass Spectrometry-Based Assay for Measurement of Angiotensin I and Plasma Renin Activity to Diagnose Secondary Hypertension

Supervisory Committee Dr. Christoph Borchers, Department of Biochemistry and Microbiology Supervisor Dr. Terry Pearson, Department of Biochemistry and Microbiology Departmental Member Dr. Patrick von Aderkas, Department of Biology Outside Member The renin-angiotensin-aldosterone system (RAAS) plays an essential role in maintaining plasma volume and arterial blood pressure by regulating angiotensin II levels. Dysregulation of the RAAS can result from an underlying disorder that results in a severe and untreatable form of hypertension, known as secondary hypertension. Measurement of plasma renin activity is a commonly employed method of diagnosing secondary hypertension. Plasma renin activity is quantified by determining the amount of angiotensin I generated through the enzymatic cleavage of angiotensinogen by renin. Radioimmunoassay is routinely used to measure plasma renin activity, however there are limitations to the method. With the prevalence of hypertension on the rise, there is need for a more accurate and rapid method of assessing the RAAS for diagnostic purposes and therapeutic monitoring. Multiplexed measurement of angiotensin I and angiotensin II would provide comprehensive understanding of the RAAS by determining dysregulation in the production of either molecule. In this thesis, the relationship between endogenous angiotensin I concentrations and plasma renin activity are studied in order to examine the research hypothesis that measurement of angiotensin I concentration correlates with iv plasma renin activity and whether this may provide a more accurate and rapid method of screening for hypertension when multiplexed with angiotensin II. To overcome the current limitations of radioimmunoassay for measuring plasma renin activity, a mass spectrometric-based method was developed to measure angiotensin I and plasma renin activity. Evaluation of the assay against radioimmunoassay demonstrates that the assay is reproducible and provides a linear response over a diagnostically relevant concentration range. Comparison of endogenous levels of angiotensin I with normal plasma renin activity levels show a correlation in this study (R=0.74). Comparison of plasma renin activity values by radioimmunoassay and iMALDI also show correlation (R=0.98), indicating that the iMALDI assay may provide an improved method for diagnosing secondary hypertension.