P002 - CHARACTERIZING AN ARRHYTHMIA-RELATED TITIN MUTATION USING PATIENT STEM CELL-DERIVED ATRIAL CARDIOMYOCYTES

Friday, October 28, 2022

4:35 PM – 4:45 PM EDT

Location: Room 215

Presenter(s)

Kate Huang, n/a

PhD Candidate
University of British Columbia
University of British Columbia
Vancouver, British Columbia, Canada

First Author(s)

Kate Huang, n/a

PhD Candidate
University of British Columbia
University of British Columbia
Vancouver, British Columbia, Canada

Author(s)

HH

Haojun Huang, n/a

PhD Candidate
Vancouver, British Columbia, Canada
MA

Mishal Ashraf, n/a

Vancouver, British Columbia, Canada
AS

Ardin Sacayanan, n/a

Vancouver, British Columbia, Canada
HL

Hattie Luo, n/a

Vancouver, British Columbia, Canada
LR

Leili Rohani, n/a

Post-doctoral Fellow
Vancouver, British Columbia, Canada
Jason Roberts, MD

Associate Professor
McMaster University and Population Health Research Institute
Population Health Research Institute and McMaster University
Hamilton, Ontario, Canada
GT

Glen F. Tibbits, PhD

Professor
Port moody, British Columbia, Canada

Senior Author(s)

Liam R. Brunham, MD, PhD, FRCPC, FACP

Associate Professor
University Of British Columbia
University of British Columbia
Vancouver, British Columbia, Canada
Zachary Laksman, n/a

Assistant Professor
University Of British Columbia
University of British Columbia
Vancouver, British Columbia, Canada

Supervisor(s)

Zachary Laksman, n/a

Assistant Professor
University Of British Columbia
University of British Columbia
Vancouver, British Columbia, Canada

Background: Atrial fibrillation (AF) is a common arrhythmia that is linked to a greater risk of ischemic stroke and heart failure. Multiple genetic studies have established an association between protein truncating variants in the titin gene and increased risk of AF in the presence or absence of cardiomyopathy. Titin truncating variants are a known cause of dilated cardiomyopathy, thus existing studies have focused on the effects of these variants in ventricular cardiomyocytes. The structural and functional consequences of titin truncating variants in atrial cardiomyocytes and how such variants lead to arrhythmias such as AF are unclear. Our objective was to investigate the cellular effects of titin truncating variants identified in patients with unexplained AF.

METHODS AND RESULTS: We identified a heterozygous titin truncating variant in a patient with early-onset atrial fibrillation and generated induced pluripotent stem cell lines (iPSCs) of the patient. We used CRISPR/Cas9 homology-directed repair to perform genome-editing of the patient iPSCs and corrected the titin truncating variant to wildtype. We differentiated patient iPSCs with the titin truncating variant and wildtype iPSCs into ventricular-like and atrial-like cardiomyocytes. We characterized cellular electrophysiology by optically mapping voltage and calcium transients and assessed the organization of sarcomere structures within cardiomyocytes through immunofluorescent staining of sarcomere proteins and confocal microscopy. iPSC-derived atrial cardiomyocytes displayed cell type-specific characteristics including faster beat rates (mean ± sem, beats/minute: 150.9 ± 21.8 vs. 21.9 ± 3.6; p = 0.01) and shorter rate-corrected action potentials (cAPD80: 195.9 ± 23.8 vs. 394.7 ± 42.8 ms; p = 0.048) compared to ventricular cardiomyocytes. Analysis of sarcomere organization showed poorer structural alignment in iPSC-derived atrial cardiomyocytes with the titin truncating variant compared to wildtype (% organization: 66.3 ± 6.8 vs. 88.0 ± 2.9; p = 0.03; Figure 1A and 1B). Similarly, iPSC-derived ventricular cardiomyocytes with the titin truncating variant showed poorer sarcomere organization compared to wildtype (62.0 ± 3.9 vs. 82.9 ± 2.9; p = 0.008; Figure 1C).

Conclusion: Titin truncating variants lead to abnormal sarcomere organization in both atrial and ventricular iPSC-derived cardiomyocytes, and this phenotype can be reverted through CRISPR/Cas9 correction of the titin truncating variant to wildtype. These findings further our understanding of the role of titin in the atria and provide insight to the mechanisms by which titin truncating variants may promote arrhythmogenesis.