Proceedings of The Physiological Society

Europhysiology 2018 (London, UK) (2018) Proc Physiol Soc 41, PCA055

Poster Communications

Mitochondrial DAMPs in ex vivo ischemia-reperfusion injury in mouse hearts

M. Torp1, T. Ranheim2, T. Flatebø1, C. Heiestad1, A. Yndestad2, K. Stensløkken1

1. University of Oslo, Oslo, Norway. 2. Oslo University Hospital, Oslo, Norway.


Background: Acute myocardial infarction results in necrosis and initiation of a local sterile inflammation activated through Damage-Associated Molecular Patterns (DAMPs). Mitochondria are highly immunogenic due to their bacterial origin and the cardiomyocyte consists of ~30% mitochondria. In this study, we attenuate Toll-like receptor 4 (TLR4) activation in isolated hearts after ischemia-reperfusion and in cardiomyocytes. We also investigate cardiomyocyte viability and inflammatory responses after exposure to mitochondrial fragments. Methods: Langendorff heart perfusion of adult C57BL/6N male mice was retrogradely perfused at constant pressure (70 mmHg). Left-ventricular pressure was monitored through a fluid-filled balloon placed in the left ventricle. The experimental set-up: 20 minutes stabilization, 35 minutes global ischemia, and 60 minutes reperfusion. TLR4 signaling was investigated by adding a TLR4 inhibitor at reperfusion. Isolated adult mouse primary cardiomyocytes were exposed to increasing concentrations of cardiac mitochondrial debris or LPS. Expression and release of cytokines were measured by qPCR and ELISA. Results: Inhibition of TLR4 at reperfusion in ex vivo Langendorff perfused mouse hearts showed reduced damage after ischemia-reperfusion. IL-6 release was significantly reduced in TLR4 inhibited hearts compared to control. Moreover, mitochondrial debris reduced viability and activated IL-6 expression in cardiomyocytes. This effect was abolished when mitochondrial debris was added in combination with TLR4 inhibitor. LPS, a known TLR4 ligand, activated the NF-κB pathway in cardiomyocytes. Conclusions: Mitochondria released after myocardial necrosis may reduce viability of neighboring cardiomyocytes. This in turn may initiate an inflammatory response in neighboring cardiomyocytes mediated in part through TLR4 and the NF-κB pathway.

Where applicable, experiments conform with Society ethical requirements