Infarct-sparing effect of myocardial postconditioning is dependent on protein kinase C signalling.
OBJECTIVE
Using non-selective and selective protein kinase C (PKC) epsilon and delta isoform inhibitors, we tested the hypothesis that the cardioprotective phenotype invoked by postconditioning (postcon) is dependent on PKC signalling. Furthermore, we determined whether postconditioning alters pPKCepsilon and/or pPKCdelta in cytosolic and mitochondrial fractions.


METHODS
Male Sprague-Dawley rats underwent 30 min left coronary artery (LCA) occlusion followed by 3 h of reperfusion. Rats were randomised to the following groups: Untreated, no intervention either before or after LCA occlusion; Postcon, 3 cycles of 10-s full reperfusion and 10-s re-occlusion, initiated immediately at the onset of reperfusion; Chelerythrine (non-selective PKC inhibitor, 5 mg/kg)+/-postcon; Rottlerin (PKCdelta inhibitor, 0.3 mg/kg)+/-postcon; KIE1-1 (PKCepsilon inhibitor, 3.8 mg/kg)+/-postcon. A subset of rats was employed to assess pPKCepsilon and/or pPKCdelta in sham, Isch/RP (30-min LCA occlusion followed by 30-min reperfusion), and postcon-treated hearts.


RESULTS
Infarct size, expressed as area of necrosis as a percentage of the area at risk, AN/AAR (%), was significantly reduced by postcon compared to control (untreated) rats (39+/-2% vs. 53+/-1% in control, P<0.001). Treatment with chelerythrine alone or the PKCepsilon antagonist KIE1-1 alone at reperfusion had no effect on infarct size compared to control. In contrast, the infarct-sparing effect of postcon was abrogated by non-selective PKC inhibition and PKCepsilon antagonism (50+/-2% and 50+/-1%, respectively, P<0.002). Inhibition of PKCdelta reduced infarct size to values comparable to that in postcon group (36+/-3% vs. 39+/-2%). However, postcon in the presence of PKCdelta inhibitor did not enhance the infarct-sparing effects (38+/-2%). In addition, pPKCepsilon in postcon hearts was significantly higher in the total cell homogenate (10338+/-1627 vs. 4165+/-608 in Isch/RP, arbitrary units), and pPKCdelta translocation to mitochondria was significantly less (>2-fold decrease) compared to Isch/RP.


CONCLUSIONS
These data suggest that postcon modulates PKC during early reperfusion by increasing PKCepsilon expression and translocation to a site other than the outer mitochondrial membrane, and limits translocation of PKCdelta to mitochondria and associated deleterious signalling.
Zatta
Amanda J
AJ
Department of Cardiothoracic Surgery, Carlyle Fraser Heart Centre/Crawford Long Hospital, Emory University School of Medicine, Atlanta, GA 30308-2225, United States. azatta@emory.edu
Kin
Hajime
H
Lee
George
G
Wang
Ningping
N
Jiang
Rong
R
Lust
Robert
R
Reeves
James G
JG
Mykytenko
James
J
Guyton
Robert A
RA
Zhao
Zhi-Qing
ZQ
Vinten-Johansen
Jakob
J
D000098:Acetophenones / Q000494:pharmacology; D000470:Alkaloids; D000818:Animals; D053119:Benzophenanthridines; D001578:Benzopyrans / Q000494:pharmacology; D015153:Blotting, Western / Q000379:methods; D003600:Cytosol / Q000201:enzymology; D008297:Male; D008929:Mitochondria, Heart / Q000201:enzymology; D023421:Models, Animal; D009203:Myocardial Infarction / Q000201:enzymology / Q000473:pathology / Q000517:prevention & control*; D015425:Myocardial Reperfusion; D009206:Myocardium / Q000201:enzymology* / Q000473:pathology; D009336:Necrosis; D010455:Peptides / Q000494:pharmacology; D010617:Phenanthridines / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051745:Protein Kinase C-delta / Q000037:antagonists & inhibitors / Q000378:metabolism; D051744:Protein Kinase C-epsilon / Q000037:antagonists & inhibitors / Q000378:metabolism; D011897:Random Allocation; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology*
D000098:Acetophenones; D000470:Alkaloids; D053119:Benzophenanthridines; D001578:Benzopyrans; C513413:KIE1-1 peptide; D010455:Peptides; D010617:Phenanthridines; C085746:rottlerin; C016299:chelerythrine; D011493:Protein Kinase C; D051745:Protein Kinase C-delta; D051744:Protein Kinase C-epsilon