Autoimmune Associated Congenital Heart Block: Pathogenic Cross Reactivity With Cardiac L-Type Calcium Channels.

Abstract

Congenital heart block (CHB) is an autoimmune disease associated with autoantibodies against ribonucleoproteins SSB/La, and SSA/Ro. Atrioventricular block is the hallmarks of CHB, however sinus bradycardia has been reported in animal models of CHB. Interestingly, knockout of the neuroendocrine Cav1.3 Ca channel results in significant sinus bradycardia and AV block. The hypothesis being tested here is that Cav1.3 Ca channel is a target for maternal autoantibodies and thus plays a role in the development of CHB. RT-PCR and immunostaining showed expression of Cav1.3 Ca channel in the human fetal heart. IgG containing ant-SSA/Ro –SSB/La autoantibodies from mothers with CHB children but not from mothers with healthy children inhibited Cav1.3 Ca current and cross-reacted with Cav1.3 Ca channel protein. Because Cav1.3 and Cav1.2 Ca channels co-exist in native cardiomyocytes, silencing of the Cav1.2 Ca channel was performed to allow characterization of Cav1.3 Ca channel in cardiac myocytes. The use of a lentiviral vector-based system to deliver siRNA to rat neonatal myocytes (RNM) resulted in ~100% transfection efficiency and 91% silencing of the Cav1.2 gene. Electrophysiological recording using the patch-clamp technique of the silenced myocytes also showed the respective inhibition of the Cav1.2 current which correlated with the biochemical results. GST Fusion proteins of the extracellular regions (S5-S6) of each of the four domains I-IV of Cav1.3 Ca channel protein were prepared and tested for reactivity with sera from mothers with CHB children using enzyme-linked immunosorbent assay (ELISA). Serum samples from 17 of 118 (14.4%) patients who had a child with CHB reacted with the Extra-cellular loop E1 corresponding to Domain I S5-S6 region. Rescue and worsening of electrocardiographic abnormalities was achieved in two immunized transgenic mouse models of CHB (Cav1.2 overexpression and Cav1.3 knockout mice). The data demonstrate 1) the expression of Cav1.3 Ca channel in human fetal heart; 2) the inhibition of Cav1.3 Ca Channel by maternal IgG, and direct cross-reactivity of with the Cav1.3 protein; 3) the extracellular loop Domain I S5-S6 of Cav1.3 Ca channel is a possible antigenic target of pathological autoantibodies in neonatal lupus; 4) overexpression of cardiac Cav1.2 reduces the electrocardiographic abnormalities seen in the pups of immunized mothers, whereas Cav1.3 knockout exhibited severe conduction abnormalities and increased mortality supporting a central role of L-type Ca channel in pathogenesis of CHB.