| Background and Aim: In recent years, it has been demonstrated that the renin-angiotensin system (RAS) plays a crucial role in the pathogenesis and development of cardiovascular diseases. At present it is commonly held that for angiotensin II, a dominant effective molecule of RAS, there are two receptor subtypes, i.e. AT1 and AT2. The biological activities of angiotensin II are primarily mediated by angiotensin II AT1 receptors. These receptors not only play an important role in the physiological regulation of cardiovascular activities but also their over-activity may lead to many pathophysiological consequences, and hence have a key role in the pathogenesis of various cardiovascular diseases. For the past few years, the existence of autoantibodies against cardiovascular G-protein coupled receptors including β1-and α1-adrenoceptors, M2-muscarinic receptors, etc, in the sera of patients with various cardiovascular diseases has been reported. These autoantibodies are able to specifically recognize the epitope of the second extracellular loop of the corresponding receptors and display a stimulatory "agonist-like"activities on their corresponding receptors. At variance with agonists, the antibodies stimulate the receptors without desensitization, and thus reveal permanent stimulatory effects resulting in pathological changes. This suggests the involvement of immunologic mechanism by receptor antibody in the pathogenesis of cardiovascular diseases. More recently, autoantibodies against angiotensin II AT1 receptors have been found in the sera of patients with preeclampsia and malignant hypertension. However, the implicative role of these autoantibodies in the pathogenesis of the diseases are unclear. In order to clarify the role of antibody against functionally important angiotensin II AT1 receptors in the pathogenesis of cardiovascular diseases, it is of significance to understand first the biological activities of these autoantibodies on cardiovascular system. Therefore, in this study we explored the effects of anti-AT1-receptor antibodies on cardiovascular activities in rats. The present study consists of following five experiments. I. Preparation of anti-AT1-receptor antibodies in rats Aim: To produce enough anti-AT1-receptor antibodies (AT1-R Ab) in the sera from rats immunized with antigenic peptide corresponding to the second extracellular loop of human angiotensin II AT1 receptors. Methods: Peptide synthesis, Active immunization of rat model, detection of anti-AT1-receptor antibodies by SA-ELISA assay, Affinity purification of IgG. Results: 1. Titres of AT1-R Ab began to increase from second week after active immunization (Fig 1, Table 1). At 8th week, the antibody titres in the sera of rats increased from the pre-immunization value of 1 : 25.56±9.22 to 1 :1490.00±847.58(p<0.01). No changes were observed for control group.2.Purified immunoglobulin fractions (IgG) from the positive sera were prepared using a MabTrap kit (Amersham). The concentration of IgG after purification is 2.38 mg/ml.These purified AT1-R Ab IgG were then used in the following studies on the biological activities of AT1-R Ab. II. Effects of AT1-R Ab on rat ventricular myocyte ionic currents Aim: In an attempt to clarify the role of AT1-R Ab in the pathogenesis of cardiovascular diseases, the direct effects of AT1-R Ab on the cardiac performance should be first investigated. In view of the myocardial electrical activities are the fundamental bases of various cardiac performances and that the research at cellular level is very suitable for the study of the direct action of antibody, so in this study we observed first the effects of AT1-R Ab on some ventricular ionic currents. Methds: Whole cell patch clamp techniques were used to study the effects of AT1-R Ab on rat ventricular ICa-L, INa/Ca, Ito, and Ik1 currents which were compared with those of Ang-II. The actions of Losartan, a specific antagonist of AT1-receptor, on these effects were also analyzed. Results: 1.Effects on ICa-...
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