Newly developed radioimmunoassay for Human Angiotensin-(1-12) measurements in plasma and urine

Abstract

The dodecapeptide angiotensin-(1-12) [Ang-(1-12)] functions as an intracrine/paracrine substrate for local production of angiotensin II. We developed a reliable and specific radioimmunoassay (RIA) method for the measurement of Ang-(1-12) in human plasma and urine using an affinity purified antibody fraction directed towards the C-terminus of the human Ang-(1-12) sequence. The RIA method was applied to quantify the Ang-(1-12) in plasma and urine collected from thirty-four human subjects (29 treated with antihypertensive medicines and 5 untreated patients). Plasma Ang-(1-12) level was significantly higher (P < 0.05) in patients with systolic blood pressure ≥140 mm Hg (n = 10) compared to the group with systolic blood pressure <140 mm Hg (n = 24). No significant difference (P = 0.22) was found in spot urine between the groups. Our study also shows that the polyclonal antibody neutralizes the cleavage sites of the human Ang-(1-12) from recombinant human chymase (rhChymase) and serum angiotensin converting enzyme (ACE) mediated Ang II generating hydrolysis. Overall, this newly developed RIA method is reliable and applicable to accurately quantify the Ang-(1-12) level in clinical samples (plasma and urine). Further, our in vitro neutralization study suggests that the anti-Ang-(1-12)-antibody might be used as an in vivo therapeutic agent for preventing Ang-(1-12)/Ang II-mediated hypertension and organ damage.

Keywords: Angiotensin I; Angiotensin II; Angiotensin-(1–12); Angiotensinogen; Hypertension; Radioimmunoassay; Renin-angiotensin system.