Abstract 15099: Suppression of Autophagic Flux by Sequestration of p62 Promotes RIP1-mediated Necroptosis in Cardiomyocytes

Purpose: Necroptosis, a programmed form of necrosis, has been implicated in various pathological conditions including heart failure. Here we examined whether and how receptor-interacting protein kinase (RIP) 1-dependent necroptosis is functionally linked with autophagy in cardiomyocytes. Methods and results: In H9c2 cells, the necroptotic pathway was activated by TNF-α (TNF, 50 ng/ml) alone or by co-incubation of TNF with 20 mM z-VAD-fmk (zVAD), a pan-caspase inhibitor. TNF increased LDH activity in the culture medium from 26.6±4.3% of total LDH activity in the vehicle-treated control to 39.1±4.8%, suggesting induction of necrosis. Addition of zVAD to TNF (TNF/zVAD) further increased the LDH activity to 70.6±2.7%. The augmented cell death by TNF/zVAD was completely reversed by necrostatin-1 (39.4±4.0%), a RIP1 inhibitor, and was attenuated by rapamycin (10 nM), a promoter of autophagy (49.4±2.0%). Immunoblot analyses showed that TNF/zVAD significantly increased the level of LC3-II. The accumulation of LC3...