Abstract 17811: Inhibition of Mitochondrial Fission and Cardioprotection: Does Activation of Necroptosis Play a Confounding Role?
Background: Emerging evidence suggests that ischemia-reperfusion-induced translocation of dynamin-related protein 1 (DRP1) to mitochondria, and subsequent fragmentation (i.e., fission) of mitochondria, contributes to myocyte death. However, recent data from our group revealed a complex temporal relationship between fission and cell fate: pretreatment with Mdivi-1 (specific DRP1 inhibitor) attenuated both apoptotic and total cell death, while ‘delayed’ treatment (at reoxygenation) attenuated apoptosis but exacerbated necrosis. There are reports that caspase inhibition can up-regulate an alternative, kinase-mediated form of cell death termed ‘necroptosis’. Accordingly, we propose that the paradox seen with ‘delayed’ Mdivi-1 treatment may be explained by Mdivi-1-triggered caspase inhibition and resultant activation of necroptosis.
Methods: Cultured HL-1 cells underwent 2 hrs of hypoxia + reoxygenation (R) or a matched normoxic period. In Protocol 1, cells received: i) Mdivi-1 (50 μM) at 1 hr before hypoxia; ii) Mdivi-1 (50 μM) at R; or iii) vehicle. In Protocol 2, Necrostatin-1 (inhibitor of RIP1, a key kinase involved in necroptosis: 50 μM) + Mdivi-1 were co-administered at R, while, in Protocol 3, Necrostatin-1 alone was given at R. Endpoints included expression of cleaved caspase 3 (harbinger of apoptosis), and cell viability (by trypan blue staining).
Results: We confirmed that Mdivi-1attenuated caspase 3 cleavage irrespective of the timing of treatment (Fig A: left), but decreased (rather than increased) cell viability when given at R (Fig A: right). Necrostatin-1 partially rescued the exacerbated cell death seen with ‘delayed’ Mdivi-1 treatment (Fig B), but had no effect when given alone (Fig C).
Conclusion: These data: i) suggest that Mdivi-1, given at reoxygenation, exacerbates myocyte death by up-regulation of necroptosis; and ii) underscore the complex temporal interplay between inhibition of mitochondrial fission and myocyte viability.
Author Disclosures: Y. Dong: None. V.V. Undyala: None. K. Przyklenk: None.
- © 2014 by American Heart Association, Inc.