Long-Term Organotypic Culture of Human Atrial Myocardium to Study Atrial Fibrillation

Background: Atrial myocardium can be a valuable source for translational cardiac research. In this study, we present a method for stable organotypic culture of beating human atrial myocardium. By investigating physiological mechanisms in long-term culture, we aim to establish a novel method for discovering mechanisms of perioperative complications like postoperative atrial fibrillation.

Method: After written informed consent, atrial tissue was donated by patients undergoing open heart surgery with cardiopulmonary bypass. Trabeculae (pectinate muscles) were prepared from the right-atrial appendage and installed into cultivation chambers at 37°C. Diastolic preload was set to 1.5 mN. After an adaptation period of 2 days with 0.5 Hz pacing, stimulation frequency was set to ≥1 Hz. Contractile force was monitored continuously. Stimulation threshold, contractile refractory period, and maximum captured frequency were assessed periodically with and without β-adrenergic stimulation. After cultivation, tissues were used for MTT viability assays, qPCR, and structural investigation by confocal microscopy.

Results: We investigated the viability in culture of 24 trabeculae obtained from six patients. All cultivated trabeculae still responded to electrical stimulation after 12 days. In 12 trabeculae from three samples, we compared functional parameters directly after installation (0 days) with those after 7 and 12 days in culture. Contractile refractory period was 168 ± 11 ms at 0 days, 177 ± 15 ms at 7 days, and 196 ± 20 ms (p = 0.64) at 12 days (p = 0.24). Maximum captured frequency was 6.2 Hz at 0 days, 6.1 Hz at 7 days, and 5.7 Hz at 12 days. Stimulation threshold increased within culture, being 48.8 mA at 7 days and 60.4 mA at 12 days. MTT assays indicated comparable viability between 0 and 12 days tissues (relative absorbance 0.17 ± 0.03 vs. 0.49 ± 0.39, three samples each). Confocal microscopy suggested preserved microstructure of cardiomyocytes and tissues. After application of 10 nM isoprenaline, parameters changed as followed: mean refractory period decreased by 23 milliseconds, maximum frequency increased by 0.5 Hz, and stimulation threshold decreased by 12 mA, suggesting increased excitability in response to β-adrenergic stimulation.

Conclusion: We established a workflow for the long-term cultivation of human atrial myocardium, including its functional assessment under various conditions, such as excessive β-adrenergic stimulation. We expect that this method will reveal new insights into the pathogenesis of atrial fibrillation.

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Artikel online veröffentlicht:
03. Februar 2022