BackgroundAgonistic autoantibodies directed at the α1-adrenergic receptor (α1-AAB) have been described in patients with hypertension. We implied earlier that α1-AAB might have a mechanistic role and could represent a therapeutic target.Methodology/Principal FindingsTo pursue the issue, we performed clinical and basic studies. We observed that 41 of 81 patients with refractory hypertension had α1-AAB; after immunoadsorption blood pressure was significantly reduced in these patients. Rabbits were immunized to generate α1-adrenergic receptor antibodies (α1-AB). Patient α1-AAB and rabbit α1-AB were purified using affinity chromatography and characterized both by epitope mapping and surface plasmon resonance measurements. Neonatal rat cardiomyocytes, rat vascular smooth muscle cells (VSMC), and Chinese hamster ovary cells transfected with the human α1A-adrenergic receptor were incubated with patient α1-AAB and rabbit α1-AB and the activation of signal transduction pathways was investigated by Western blot, confocal laser scanning microscopy, and gene expression. We found that phospholipase A2 group IIA (PLA2-IIA) and L-type calcium channel (Cacna1c) genes were upregulated in cardiomyocytes and VSMC after stimulation with both purified antibodies. We showed that patient α1-AAB and rabbit α1-AB result in protein kinase C alpha activation and transient extracellular-related kinase (EKR1/2) phosphorylation. Finally, we showed that the antibodies exert acute effects on intracellular Ca2+ in cardiomyocytes and induce mesentery artery segment contraction.Conclusions/SignificancePatient α1-AAB and rabbit α1-AB can induce signaling pathways important for hypertension and cardiac remodeling. Our data provide evidence for a potential clinical relevance for α1-AAB in hypertensive patients, and the notion of immunity as a possible cause of hypertension.

Background-Catecholamines hasten cardiac relaxation through ␤-adrenergic receptors, presumably by phosphorylation of several proteins, but it is unknown which receptor subtypes are involved in human ventricle. We assessed the role of ␤ 1 -and ␤ 2 -adrenergic receptors in phosphorylating proteins implicated in ventricular relaxation. Methods and Results-Right ventricular trabeculae, obtained from freshly explanted hearts of patients with dilated cardiomyopathy (nϭ5) or ischemic cardiomyopathy (nϭ5), were paced at 60 bpm. After measurement of the contractile and relaxant effects of epinephrine (10 mol/L) or zinterol (10 mol/L), mediated through ␤ 2 -adrenergic receptors, and of norepinephrine (10 mol/L), mediated through ␤ 1 -adrenergic receptors, tissues were freeze clamped. We assessed phosphorylation of phospholamban, troponin I, and C-protein, as well as specific phosphorylation of phospholamban at serine 16 and threonine 17. Data did not differ between the 2 disease groups and were therefore pooled. Epinephrine, zinterol, and norepinephrine increased contractile force to approximately the same extent, hastened the onset of relaxation by 15Ϯ3%, 5Ϯ2%, and 20Ϯ3%, respectively, and reduced the time to half-relaxation by 26Ϯ3%, 21Ϯ3%, and 37Ϯ3%. These effects of epinephrine, zinterol, and norepinephrine were associated with phosphorylation (pmol phosphate/mg protein) of phospholamban 14Ϯ3, 12Ϯ4, and 12Ϯ3; troponin I 40Ϯ7, 33Ϯ7, and 31Ϯ6; and C-protein 7.2Ϯ1.9, 9.3Ϯ1.4, and 7.5Ϯ2.0. Phosphorylation of phospholamban occurred at both Ser16 and Thr17 residues through both ␤ 1 -and ␤ 2 -adrenergic receptors. Conclusions-Norepinephrine and epinephrine hasten human ventricular relaxation and promote phosphorylation of implicated proteins through both ␤ 1 -and ␤ 2 -adrenergic receptors, thereby potentially improving diastolic function. (Circulation. 1999;99:65-72.)

Background —Recent studies of β-adrenergic receptor (β-AR) subtype signaling in in vitro preparations have raised doubts as to whether the cAMP/protein kinase A (PKA) signaling is activated in the same manner in response to β 2 -AR versus β 1 -AR stimulation. Methods and Results —The present study compared, in the intact dog, the magnitude and characteristics of chronotropic, inotropic, and lusitropic effects of cAMP accumulation, PKA activation, and PKA-dependent phosphorylation of key effector proteins in response to β-AR subtype stimulation. In addition, many of these parameters and L-type Ca 2+ current ( I Ca ) were also measured in single canine ventricular myocytes. The results indicate that although the cAMP/PKA-dependent phosphorylation cascade activated by β 1 -AR stimulation could explain the resultant modulation of cardiac function, substantial β 2 -AR–mediated chronotropic, inotropic, and lusitropic responses occurred in the absence of PKA activation and phosphorylation of nonsarcolemmal proteins, including phospholamban, troponin I, C protein, and glycogen phosphorylase kinase. However, in single canine myocytes, we found that β 2 -AR–stimulated increases in both I Ca and contraction were abolished by PKA inhibition. Thus, the β 2 -AR–directed cAMP/PKA signaling modulates sarcolemmal L-type Ca 2+ channels but does not regulate PKA-dependent phosphorylation of cytoplasmic proteins. Conclusions —These results indicate that the dissociation of β 2 -AR signaling from cAMP regulatory systems is only apparent and that β 2 -AR–stimulated cAMP/PKA signaling is uncoupled from phosphorylation of nonsarcolemmal regulatory proteins involved in excitation-contraction coupling.

Abstract-Abnormal uterine perfusion detected by Doppler sonography reflects impaired trophoblast invasion, a factor involved in the pathogenesis of pregnancy complications such as preeclampsia or intrauterine growth retardation. Recent studies have demonstrated an autoantibody against the angiotensin type 1 (AT 1 ) receptor in pregnant women with preeclampsia. Our aim was to determine whether the AT 1 autoantibody precedes the clinical symptoms and is thus predictive of preeclampsia. We therefore detected this antibody in serum from second trimester pregnancies with abnormal uterine perfusion because these women show an indirect sign of inadequate trophoblast invasion. Then the AT 1 autoantibody distribution/concentration was compared with that of women at term with or without pregnancy pathology. The AT 1 autoantibody was already detectable in second trimester pregnant women with abnormal uterine perfusion before the clinical manifestation of preeclampsia (80%). However, it was also found in second trimester pregnant women with abnormal uterine perfusion who later developed intrauterine growth retardation (60%) or even had a normal course of pregnancy (62%). In the third trimester, the AT 1 autoantibody was demonstrated in 89% of patients with manifest preeclampsia, 86% of those with manifest intrauterine growth retardation, and even in healthy pregnant women at term with a history of abnormal uterine perfusion in the second trimester. We conclude that the AT 1 autoantibody is an early but nonspecific marker for preeclampsia. The generation of this antibody seems to be associated with distinct types of pregnancy disorders resulting from impaired placental development. The AT 1 autoantibody may thus be causative for pathological uteroplacental perfusion. Key Words: angiotensin II Ⅲ antibodies Ⅲ hypertension, gestational Ⅲ preeclampsia Ⅲ pregnancy Ⅲ receptors, angiotensin P reeclampsia, a serious pregnancy-specific disorder characterized by proteinuria and hypertension after the 20th week of gestation, is still a leading cause of maternal and neonatal morbidity and mortality. The lack of preventive and causal treatment is mainly attributable to the fact that the etiology and pathophysiology are not understood. 1 Impairment of placental development and trophoblast invasion is thought to be causally associated with preeclampsia. Doppler ultrasound determination of uterine perfusion is a noninvasive tool for measuring uteroplacental vascular resistance and assessing the quality of the trophoblast invasion. 2 The trophoblast failed to develop adequately in pregnancies with abnormal uterine perfusion. Trophoblast invasion into the maternal compartment is thus impaired, and the pregnant uterus is not transformed into a low-resistance bed. Precisely this pathology is thought to be a pathogenetic feature of pregnancy complications such as preeclampsia or intrauterine growth retardation (IUGR). Abnormal uterine perfusion characterizes pregnancies at risk for these complications and precedes their clinical manifestation. H...