Mitochondrial transplantation normalizes transcriptional shift associated with ischemia reperfusion injury in neonatal hearts donated after circulatory death
*Ilias P. Doulamis1, *Aspasia Tzani2, *Victor Alemany3, *Rio Nomoto3, *Aybuke Celik3, *Dominic Recco3, *Mossab Saeed3, *Alvise Guariento3, *Sitaram Emani3, *Pedro J del Nido3, *James D McCully3
1. Surgery, Lahey Clinic, Brookline, MA, United States. 2. Cardiology, Brigham and Women's Hospital, Boston, MA, United States. 3. Cardiac Surgery, Boston Children's Hospital, Boston, MA, United States.
Objective: We have previously shown that mitochondrial transplantation preserves myocardial function and viability in neonatal swine hearts donated after circulatory death (DCD). Here, we investigate the transcriptomic alterations following mitochondrial transplantation in DCD neonatal hearts.
Methods: Circulatory death was induced in neonatal Yorkshire pigs by cessation of mechanical ventilation, followed by 20 min of warm ischemia time, 10 min of cold cardioplegic arrest, and then 4 h of unloaded and loaded ex situ heart perfusion (ESHP). Hearts received autologous mitochondria or vehicle after 15 min of unloaded perfusion. A sham group did not undergo warm ischemia, mimicking donation after brain death heart procurement. To study the early expressed underlying global transcriptomic changes involved in the cardioprotection conferred by mitochondrial transplantation and identify critical regulators mediating these responses, RNA-sequencing of the heart tissue was performed.
Results:
Following 4 hours of ESHP, myocardial function was significantly increased in DCD hearts receiving mitochondria compared to vehicle. Gene ontology (GO) pathway analysis showed that ATP binding and ribosomal RNA biogenesis were among the top significantly upregulated molecular pathways. GO analysis for cellular compartment driving these changes identified the mitochondrion as the predicted source. Gene set enrichment analysis revealed hallmarks of angiogenesis and cardiomyocyte proliferation upregulated in the mitochondria compared to the vehicle group, while pathways related to apoptosis, and fatty acid metabolism related to ischemia were downregulated. Upstream regulator analysis revealed increase of transcriptional factors related to angiogenesis and cardiac cell fate and decrease of p53-driven transcriptional cascades. Interestingly, comparison of the cardiac transcriptome of the mitochondrial transplantation group to the group of the sham hearts, showed no significant changes.
Conclusions: Mitochondrial transplantation in neonatal pig DCD hearts significantly enhances the preservation of myocardial function by upregulating ribosomal RNA and mitochondrial ATP-related pathways and by downregulating the immune response to ischemia. Moreover, the transcriptomic profile of the mitochondrial transplantation group was similar to that of the hearts that did not undergo any ischemic injury. Further in vivo studies are required to replicate the long-term findings of our experiments.
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