Abstract Body: Background: Atrial cardiomyopathy leads to devastating conditions such as atrial fibrillation and stroke, major causes of global mortality. While genetic variants in the NPPA locus are linked to these conditions, research has typically focused on coding mutations. NPPA-AS1 is a lncRNA transcribed in the antisense frame from the NPPA gene; it is expressed primarily in the atria and is upregulated in patients with cardiomyopathy and heart failure.

Hypothesis: We propose that NPPA-AS1 is a critical regulator of atrial cardiomyocyte (aCM) physiology and that its loss contributes to atrial dysfunction.

Methods: We generated NPPA-AS1-knockout (KO) human induced pluripotent stem cells (hiPSCs) and differentiated them into aCMs. Successful differentiation was confirmed by upregulation of MLC2a and several atria-enriched genes, assessed by both RT-qPCR and immunocytochemistry. Multielectrode array was used to assess electrophysiological features including beat period and field potential duration corrected for beat rate (FPDc); in KO-aCMs treated with I_KACh-activating carbamylcholine (CCh) beat period prolongation was observed, providing further support of successful iPSC-aCM differentiation.

Results: While WT- and KO-aCMs showed similar cTNT expression (indicating similar iPSC-CM differentiation efficiency), KO-aCMs showed significantly increased expression of atrial KCNA5, KCNJ3 and KCNJ5 channel subunits, and the atria-enriched transcription factor NR2F2. Compared to WT controls, KO-aCMs displayed lower contractility and aberrant electrophysiological features including prolonged baseline FPDc and paradoxical CCh-induced FPDc lengthening.

Conclusions: These preliminary findings suggest that NPPA-AS1 plays an essential regulatory role in maintaining atrial cardiomyocyte physiology. Ongoing studies are evaluating the effects of NPPA-AS1 overexpression in iPSC-aCMs and the in vivo impact of Nppa-as loss in maintaining aCM physiology using Nppa-as germline knockout mice.