Myocardial ischemic preconditioning preserves postischemic function of the 26S proteasome through diminished oxidative damage to 19S regulatory particle subunits.
BACKGROUND
The ubiquitin proteasome system (UPS) becomes dysfunctional as a result of ischemia/reperfusion (I/R), which may lead to dysregulation of signaling pathways. Ischemic preconditioning (IPC) may prevent dysregulation by preventing UPS dysfunction through inhibition of oxidative damage.


OBJECTIVE
Examine the hypothesis that early IPC preserves postischemic UPS function thus facilitating prosurvival signaling events.


RESULTS
I/R decreased proteasome chymotryptic activity by 50% in isolated rat heart and an in vivo murine left anterior descending coronary artery occlusion model. Following IPC, proteasome activity was decreased 25% (P<0.05) in isolated heart and not different from baseline in the murine model. Enriched 26S proteasome was prepared and analyzed for protein carbonyl content. Increased (P<0.05) carbonylation in a 53-kDa band following I/R was diminished by IPC. Immunoprecipitation studies indicated that the 53-kDa carbonylation signal was of proteasomal origin. Two-dimensional gel electrophoresis resolved the 53-kDa band into spots analyzed by liquid chromatography/tandem mass spectrometry containing Rpt3/Rpt5 both of which could be immunoprecipitated conjugated to dinitrophenylhydrazine (DNPH). Higher amounts of DNPH-tagged Rpt5 were immunoprecipitated from the I/R samples and less from the IPC samples. I/R increased Bax levels by 63% (P<0.05) which was decreased by IPC. Lactacystin (lac) pretreatment of preconditioned hearts increased Bax by 140% (P<0.05) and also increased ubiquitinated proteins. Pretreatment of hearts with a proteasome inhibitor reversed the effects of IPC on postischemic Rpt5 carbonylation, cardiac function, morphology and morphometry, and ubiquitinated and signaling proteins.


CONCLUSIONS
These studies suggest that IPC protects function of the UPS by diminishing oxidative damage to 19S regulatory particle subunits allowing this complex to facilitate degradation of proapoptotic proteins.
Divald
Andras
A
FAHA, The Feinstein Institute for Medical Research, 350 Community Drive, Manhasset, NY 11030, USA.
Kivity
Shaye
S
Wang
Ping
P
Hochhauser
Edith
E
Roberts
Beth
B
Teichberg
Saul
S
Gomes
Aldrin V
AV
Powell
Saul R
SR
D000818:Animals; D054059:Coronary Occlusion / Q000503:physiopathology; D006321:Heart / Q000503:physiopathology; D019157:Ischemic Preconditioning, Myocardial; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D023421:Models, Animal; D017202:Myocardial Ischemia / Q000503:physiopathology*; D018384:Oxidative Stress / Q000502:physiology*; D046988:Proteasome Endopeptidase Complex / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D025801:Ubiquitin / Q000502:physiology; D051028:bcl-2-Associated X Protein / Q000502:physiology
D025801:Ubiquitin; D051028:bcl-2-Associated X Protein; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease; C119011:26S proteasome non-ATPase regulatory subunit 13