The ultimate goal of this application is to understand the mechanisms of autoimmune-associated congenital heart block (CHB) and develop novel therapeutic approaches to reverse or prevent CHB. CHB, detected at or before birth in a structurally normal heart, is a conduction abnormality that affects fetuses and/or newborn of mothers with autoantibodies reactive with the intracellular soluble ribonucleoproteins 48kD SSB/La, 52kD SSA/Ro, and 60kD SSA/Ro. CHB is presumed to be due to the transplacental passage of autoantibodies from the mother into the fetal circulation. CHB carries substantial mortality and morbidity, with >60% of affected children requiring lifelong pacemakers. We have reproduced the clinical complete AV block in an animal model of CHB and in isolated Langendorff perfused fetal heart. These findings were correlated with maternal anti-Ro/La antibodies' inhibition of L-type (1c Ca channel. We have also reported for the first time significant sinus bradycardia unrelated to AV block. This novel observation indicates that the spectrum of conduction abnormalities extend beyond the AV node to also include the SA node. Interestingly knockout of (1D L-type Ca channel in mice resulted in sinus bradycardia and AV block, suggesting a critical role of (1D Ca channels in SA node pacemaker and AV node conduction. Our preliminary data show exclusive expression of (1D L-type Ca channel in the SA and AV node but not in the ventricles; and inhibition of (1D Ca current by maternal anti-Ro/La antibodies. This inhibition was completely reversed pharmacologically by Ca channel agonists. All together, these data provide strong evidence supporting an etiologic role of anti-Ro/La antibodies and Ca channels in the pathogenesis of CHB. However, the [process by which the antibodies cause CHB is not known and experimental approaches to reverse CHB have not yet been explored. We propose to build on our observations to delineate the molecular and cellular mechanisms of these putative maternal anti-Ro/La antibodies in the genesis of electrocardiographic abnormalities seen in the fetus and newborn, address the junctional vulnerability of the fetal heart to CHB and explore novel experimental approaches in reversing CHB. Our hypothesis is that Ca channels are the direct target for maternal anti-Ro/La antibodies, and that Ca channels play a significant role in autoantibody-mediated cardiac disorders. We will focus upon five key specific aims that are likely to provide important mechanistic and therapeutic information:
Specific Aim#1. To investigate the direct biochemical and functional interaction between maternal antibodies and L-type Ca channels, mainly (1D channels.
Specific Aim#2. To investigate the consequences of chronic, in utero, exposure of L-type Ca channels to maternal antibodies.
Specific Aim#3. To investigate the ionic basis of sinus bradycardia recently reported in animal models of CHB and clinically in affected newborn. ? Specific Aim#4. To investigate the potential mechanisms contributing to the vulnerability of fetal heart to CHB.
Specific Aim#5. To test whether upregulation of Ca channel density will reverse or prevent the development of CHB. Experiments will be conducted in transgenic animal models, isolated single cardiac myocytes and heterologous expression systems. State-of-the-art electrophysiological, biochemical and molecular techniques will be used. Obtaining information regarding the cellular and molecular mechanisms that underlies CHB is an absolute prerequisite for the development of effective means for the detection and possible prevention of maternal antibody-mediated CHB. ? ?
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