Abstract Body: Background: Lamin A/C (LMNA) is a nuclear intermediate filament protein that is expressed ubiquitously. LMNA mutations account for 6% of dilated cardiomyopathy, and patients affected by these mutations develop severe heart failure and lethal arrhythmias. Most LMNA variants observed in patients have unknown functional significance and an uncertain contribution to disease. A single nucleotide missense mutation (L35P) in the LMNA gene replaces leucine with proline at codon 35 of the LMNA protein. A previous study reported that congenital muscular dystrophy patients with the L35P LMNA mutation are likely to develop dilated cardiomyopathy (DCM) later in life. However, the underlying cellular pathology responsible for LMNA-related DCM is not fully understood.
Objective: To examine the functional significance of the LMNA L35P mutation in cardiomyocytes (CMs).
Methods and Results: In this study, we utilized hiPSC-derived CMs harboring the L35P mutation and its isogenic control (L35P Corrected). We developed a high-throughput in vitro assay to measure the action potential, calcium cycling, and contractility of hiPSC-CMs. Briefly, the hiPSC-CMs were cultured, labeled with fluorescent probes, and imaged with a high-speed camera at 30 frames per second. The single-cell analysis of CM functional parameters revealed that L35P mutant CMs exhibit shorter action potential duration, prolonged calcium transient duration, and reduced contractility. To identify the underlying molecular mechanism, we assessed the gene expression profile of ion channels and calcium-handling proteins. The qPCR results confirmed the differential expression of key proteins involved in the excitation-contraction coupling in L35P LMNA mutant hiPSC-CMs compared to the isogenic control. Our results indicate that the LMNA L35P mutation leads to excitation-contraction coupling defects in human iPSC-derived cardiomyocytes.
Conclusion: Our study confirms the pathogenicity of the LMNA L35P mutation in hiPSC-derived CMs and presents an in vitro model of LMNA-DCM for assessing the safety and efficacy of therapeutic drugs.