Pulse field ablation induces nonapoptopic cardiomyocyte death in vitro accompanied by thermal effects

Pulsed field ablation (PFA) has been introduced into clinical practice as a nonthermal method for pulmonary vein isolation in atrial fibrillation. PFA is deemed to induce cell death through apoptotic pathways with minimal thermal injury. However, emerging evidence indicates discrepancies in the proposed mechanisms of cardiomyocyte death.

The objective of this study is to evaluate the thermal effects during PFA in vitro and to characterize the resulting type of cell death.

In this in vitro study, we exposed the immortalized cardiomyocyte HL-1 cell line to high-frequency irreversible electroporation using electric fields ranging from 250V/cm to 1500V/cm. Apoptosis was assessed by cleaved caspase 3 and 9 at 1, 8, and 24 hours after PFA. Total cell death was assessed 24 hours after PFA using propidium iodide. Simultaneously, we simulated temperature changes and validated using high-frequency thermosensors in vitro. Thermal contribution to cell death was further analyzed with the Arrhenius model.

The overall cardiomyocyte death increased with the increasing electric field. Caspase-3 activation was not detected under any electric field at any timepoints. Numerical simulation predicted temperature hotspots with a maximum of 83.6°C at 1500V/cm, closely correlating with experimental temperature measurements. According to Arrhenius modeling, at 1500V/cm, approximately 75.8% of cell death was attributable to thermal injury, whereas at ≤1250V/cm, it was not.

PFA of cardiomyocytes in vitro induces predominantly nonapoptotic cell death, with both electroporation and thermal mechanisms contributing in a field strength-dependent manner. In contrast, apoptotic cell death was not observed after PFA.