These data were compared with sera obtained from a group of normotensive control subjects who were expected to have negligible AT1R autoantibodies. Results: Sera from each of the 13 PA patients significantly increased AT1R activation in AT1R-transfected cells compared with 20 control subjects, and this activity was inhibited by the selective AT1R blocker losartan. was inhibited by the selective AT1R blocker losartan. Sera and IgG purified from AT1R autoantibody-positive sera demonstrated significant vasoconstrictive effects in isolated rat cremaster arterioles and were blocked by losartan. Moreover, the AT1R autoantibody-positive IgG directly stimulated aldosterone production in the cultured adrenal cells and enhanced angiotensin-induced aldosterone production in these cells, and these effects were blocked by candesartan. Conclusions: These data support a probable pathophysiological role for AT1R autoantibodies in PA and thereby raise important etiological and therapeutic implications. The pathophysiological mechanisms causing excessive aldosterone production and hypertension in primary aldosteronism (PA) have challenged investigators since the first description of an aldosterone-producing adrenal adenoma (APA) (1). Extensive follow-up studies after successful resection of the tumors demonstrated residual hypertension in up to 50% of these patients (2,C5). This question was sharpened when a large number of patients with a similar phenotype and biochemical PA were demonstrated to have idiopathic adrenal hyperplasia (IAH) rather than an adenoma (6, 7); and when their adrenals were removed in three small studies, almost all retained some degree of residual hypertension (8,C10). Thus, aldosterone excess alone does not account for the complex hypertensive pathophysiology in patients with PA. Autoantibodies that activate the G protein-coupled angiotensin II type 1 receptor (AT1R) have been described in patients during acute circumstances such as preeclampsia (11), in some circumstances involving acute renal transplant rejection (12); and in a few screened hypertensive subjects (13). We hypothesized the presence of such autoantibodies would cause or contribute to the hypertension and hyperaldosteronism normally associated with IAH and also in those with an APA, irrespective of the presence or absence of an identifiable etiology such as associated mutations of the channels (14). Rossitto et al (15) recently reported the presence of autoantibodies to the AT1R, as measured by an ELISA, in subjects with an APA but not in those with IAH. They proposed use of this assay as a diagnostic tool GRI 977143 to differentiate the two different types of PA. The ELISA technique, however, has limitations with regard to sensitivity when linear peptide targets are used. Because this type of assay inherently does not measure the ability of these autoantibodies to activate the AT1R, there is need for further studies to examine the hypertensive mechanisms by which their presence might alter or participate in the pathophysiology of PA. In GRI 977143 the present study, we have examined the mechanism(s) by which these autoantibodies might interact with and affect important target tissues including arterial smooth muscle and the adrenal cortex in patients Rabbit Polyclonal to Sodium Channel-pan with biochemically confirmed PA. We retrospectively had available sera from 13 subjects with documented PA and observed all harbored some level of autoantibodies that activated AT1R-transfected Chinese hamster ovary (CHO) cells in vitro. We have demonstrated these autoantibodies increase vascular contractility in vitro similar to angiotensin II (Ang II) in the presence of a concurrently suppressed classical renin-angiotensin system. Like native Ang II, this autoantibody contractile effect appears to be largely inhibited by direct AT1R blockade. These autoantibodies also increase and/or enhance Ang II-mediated aldosterone production in vitro. These data support the possibility these activating autoantibodies, when present, may contribute to and/or modulate known pathophysiological mechanism(s) identified in patients with PA and in other subjects expressing this same AT1R-oriented autoantibody. Materials and Methods Study subjects We had accumulated sera from 13 subjects with a variable history of hypertension, hypokalemia, and a plasma aldosterone to plasma renin activity (PRA) ratio of greater than 30 (Table 1). The diagnosis of PA was confirmed by a plasma aldosterone to PRA value greater than 80 and/or by an abnormal captopril suppression test [2 h after captopril plasma aldosterone 12 ng/dL and/or plasma aldosterone to PRA ratio 12 (16)]; an 8:00 am plasma aldosterone of 6 ng/dL or greater after a high Na ( 250 mEq Na/d) diet for 3 days (two subjects) or a plasma aldosterone concentration of 6 ng/dL or greater after a saline infusion test of 2 L over 4 hours in one subject (17). All underwent a serial 2-mm section computed tomography (CT) of the adrenals to identify existing nodules. GRI 977143 Patients.