Abstract 657 Establishment of a new transgenic mouse model for arrhythmias without structural heart diseases

Background

Genome-wide association studies linked polymorphisms in the Nitric oxide synthase 1 adaptor protein (NOS1AP) gene to variations in the QTc interval and sudden cardiac death. To investigate the molecular mechanism of NOS1AP we established a conditional transgenic mouse model with over-expression of Nos1ap. We postulate NOS1AP to act as an L-type calcium channel (CaV1.2) modulator via directing NOS1 to CaV1.2 causing enhanced nitrosylation of the channel.

Methods and results

Transgenic mice with conditional over-expression of Nos1ap in cardiac myocytes were used as model organism. Western blot analysis confirmed successful induction as we found a 2.5fold higher Nos1ap expression in induced compared to non-induced mice. We found no correlation (0.207) between Nos1ap and Nos1 expression levels in the myocardium and no significant differences between cGMP production as we measured 0.199 ± 0.036 pmol/mg cGMP in non-induced and 0.204 ± 0.035 pmol/mg cGMP in Nos1ap over-expressing mice (p>0.05), hence the cardiac NO amount was obviously not altered by Nos1ap over-expression.

We confirmed the interaction of Nos1ap with Nos1 and CaV1.2. Electrocardiography in Nos1ap over-expressing mice showed ventricular tachycardia and fibrillation associated with a significant decrease in QTc intervals. Survival was significantly reduced (to 60 % after 12 weeks vs. 100 % in non-induced mice).

Whole-cell patch-clamp measurements in isolated adult ventricular myocytes were performed and action potential duration at 90 % of repolarization (APD₉₀) was significantly reduced in induced transgenic Nos1ap over-expressing mice compared to control littermates. In addition, we investigated the functional effect of the human SNP rs16847548 (T/C) and found a decrease in NOS1AP expression.

Structural examination revealed constant heart dimensions and wall thickness without abnormal fibrosis content or BNP production after three months of Nos1ap over-expression compared to controls.

Conclusion

Myocardial over-expression of NOS1AP leads to short QT syndrome with increased susceptibility to ventricular arrhythmias. In accordance, APD₉₀ is significantly shortened in over-expressing animals. The human SNP rs16847548, which was correlated to LQTS in clinical studies, is located in the promoter region of NOS1AP and results in a reduced NOS1AP promoter activity in vitro, hereby providing an explanation for the prolongation of QT interval. We conclude that changes in NOS1AP expression have an impact on NOS1 guiding and binding to CaV1.2. This modulates action potential duration in cardiac myocytes and increases susceptibility to cardiac rhythm disorders.

Clin Res Cardiol (2022). https://doi.org/10.1007/s00392-022-02002-5
Establishment of a new transgenic mouse model for arrhythmias without structural heart diseases
M. Jänsch¹, M. Trum², T. Williams³, K. Schuh⁴, F. Qadri⁵, L. S. Maier², M. Bader⁵, O. Ritter⁶
¹Department of Cardiology, Nephrology and Pneumology, Brandenburg Medical School, Brandenburg; ²Klinik und Poliklinik für Innere Med. II, Kardiologie, Universitätsklinikum Regensburg, Regensburg; ³Deutsches Zentrum für Herzinsuffizienz, Universitätsklinikum Würzburg, Würzburg; ⁴Vegetative Physiologie, Institut für Physiologie, Würzburg; ⁵Molecular Biology of Peptide Hormones, Max Delbrück Center for Molecular Medicine in the Helmholtz Association, Berlin-Buch; ⁶Zentrum für Innere Medizin I, Städt. Klinikum Brandenburg, Brandenburg an der Havel;
Background Genome-wide association studies linked polymorphisms in the Nitric oxide synthase 1 adaptor protein (NOS1AP) gene to variations in the QTc interval and sudden cardiac death. To investigate the molecular mechanism of NOS1AP we established a conditional transgenic mouse model with over-expression of Nos1ap. We postulate NOS1AP to act as an L-type calcium channel (CaV1.2) modulator via directing NOS1 to CaV1.2 causing enhanced nitrosylation of the channel. Methods and results Transgenic mice with conditional over-expression of Nos1ap in cardiac myocytes were used as model organism. Western blot analysis confirmed successful induction as we found a 2.5fold higher Nos1ap expression in induced compared to non-induced mice. We found no correlation (0.207) between Nos1ap and Nos1 expression levels in the myocardium and no significant differences between cGMP production as we measured 0.199 ± 0.036 pmol/mg cGMP in non-induced and 0.204 ± 0.035 pmol/mg cGMP in Nos1ap over-expressing mice (p>0.05), hence the cardiac NO amount was obviously not altered by Nos1ap over-expression. We confirmed the interaction of Nos1ap with Nos1 and CaV1.2. Electrocardiography in Nos1ap over-expressing mice showed ventricular tachycardia and fibrillation associated with a significant decrease in QTc intervals. Survival was significantly reduced (to 60 % after 12 weeks vs. 100 % in non-induced mice). Whole-cell patch-clamp measurements in isolated adult ventricular myocytes were performed and action potential duration at 90 % of repolarization (APD₉₀) was significantly reduced in induced transgenic Nos1ap over-expressing mice compared to control littermates. In addition, we investigated the functional effect of the human SNP rs16847548 (T/C) and found a decrease in NOS1AP expression. Structural examination revealed constant heart dimensions and wall thickness without abnormal fibrosis content or BNP production after three months of Nos1ap over-expression compared to controls. Conclusion Myocardial over-expression of NOS1AP leads to short QT syndrome with increased susceptibility to ventricular arrhythmias. In accordance, APD₉₀ is significantly shortened in over-expressing animals. The human SNP rs16847548, which was correlated to LQTS in clinical studies, is located in the promoter region of NOS1AP and results in a reduced NOS1AP promoter activity in vitro, hereby providing an explanation for the prolongation of QT interval. We conclude that changes in NOS1AP expression have an impact on NOS1 guiding and binding to CaV1.2. This modulates action potential duration in cardiac myocytes and increases susceptibility to cardiac rhythm disorders.
https://dgk.org/kongress_programme/jt2022/aP1547.html