Generation of two homozygous SHOX2 knock-out human induced pluripotent stem cell lines using CRISPR/Cas9

Area of Science:

• Cardiovascular Biology
• Stem Cell Research
• Genetics

Background:

• The SHOX2 gene, a homeobox transcription factor, is implicated in cardiac development and function.
• SHOX2 dysfunction is associated with arrhythmogenic diseases, including atrial fibrillation (AF) and sinus node dysfunction.
• Understanding the precise role of SHOX2 in cardiac electrophysiology is crucial for developing targeted therapies.

Purpose of the Study:

• To generate and characterize homozygous SHOX2 knock-out human induced pluripotent stem cell (hiPSC) lines.
• To establish a cellular model for investigating the functional consequences of complete SHOX2 loss in the context of arrhythmogenic diseases.
• To provide a valuable research tool for studying SHOX2's impact on cardiac cellular function.

Main Methods:

• CRISPR/Cas9 gene editing technology was employed to create SHOX2 knock-out hiPSC lines.
• Two distinct homozygous SHOX2 knock-out lines were generated: one from a healthy control and one from a corrected AF patient line.
• Characterization included assessment of pluripotency, differentiation potential into three germ layers, and karyotype stability.

Main Results:

• Successfully generated two homozygous SHOX2 knock-out hiPSC lines.
• The generated hiPSC lines retained pluripotency and differentiation capabilities.
• Karyotype analysis confirmed the stability and normal chromosomal content of the engineered cell lines.

Conclusions:

• Homozygous SHOX2 knock-out hiPSC lines were successfully created using CRISPR/Cas9.
• These cell lines serve as a robust model for investigating the cellular impact of SHOX2 deficiency in arrhythmogenic diseases.
• This research provides a critical tool for advancing the understanding of SHOX2's role in cardiac electrophysiology and related disorders.