Activation of apoptotic and inflammatory pathways in dysfunctional donor hearts.
BACKGROUND
Myocardial dysfunction is common after brain death, but the mechanisms remain unclear. Apoptosis is tightly regulated by enzymes termed the caspases. We have investigated the caspases involved in the terminal part of the apoptotic pathway in dysfunctional (nontransplanted) donor hearts and their relation to inflammatory markers and compared them to hearts with good ventricular function (transplanted donors).


METHODS
Thirty-one donor hearts assessed for transplantation were examined. Western blotting was used to measure pro-caspase-9, caspase-3, DFF45, the activated nuclease CPAN and poly (ADP-ribose) polymerase, a DNA repair enzyme inactivated by caspase-3. Caspase-3 activity was also measured. Histologic and immunocytochemical analysis for HLA Class II and Real Time polymerase chain reaction for tumor necrosis factor-alpha and interleukin 6 were performed to detect inflammatory activation.


RESULTS
Cleaved caspase-9 was higher (5.53+/-0.6 vs. 3.64+/-0.4 O.D. units, P<0.01) in nontransplanted compared with transplanted donors and there was a trend for higher pro-caspase-9 (5.20+/-1.0 vs. 4.22+/-0.4 O.D. units, P=NS). Levels of pro-caspase-3 were higher in nontransplanted (9.66+/-0.5 vs. 5.15+/-0.5 O.D. units, P<0.00001) donors and cleavage products of caspase-3 were elevated in 14 of 14 nontransplanted and 2 of 17 transplanted donors. Intact DFF-45 (8.94+/-0.36 vs. 6.14+/-0.30 O.D. units, P<0.000005), its spliced product (2.38+/-0.35 vs. 0.4+/-0.21 O.D. units, P=0.0001) and the nuclease caspase-activated nuclease (2.01+/-0.3 vs. 0.66+/-0.16 OD units, P=0.001) were higher in nontransplanted donors. The caspase-3 substrate poly (ADP-ribose) polymerase was higher in nontransplanted (1.16+/-0.13 vs. 0.61+/-0.22 O.D. units, P=0.57) donors.


CONCLUSIONS
The caspases are elevated in dysfunctional donor hearts compared with hearts with good ventricular function with a possible link to inflammatory activation supporting the concept that brain death causes inflammatory activation which can lead to apoptosis with a possible important effect on function.
Birks
E J
EJ
Imperial College of Science, Technology and Medicine, Royal Brompton and Harefield Hospital, Middlesex, United Kingdom.
Yacoub
M H
MH
Burton
P S
PS
Owen
V
V
Pomerance
A
A
O'Halloran
A
A
Banner
N R
NR
Khaghani
A
A
Latif
N
N
D000328:Adult; D000906:Antibodies / Q000378:metabolism; D017209:Apoptosis / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D005260:Female; D006321:Heart / Q000502:physiology*; D016027:Heart Transplantation / Q000276:immunology / Q000473:pathology / Q000502:physiology; D000949:Histocompatibility Antigens Class II / Q000096:biosynthesis; D006801:Humans; D007249:Inflammation / Q000503:physiopathology*; D008297:Male; D009206:Myocardium / Q000201:enzymology; D011065:Poly(ADP-ribose) Polymerases / Q000276:immunology; D011506:Proteins / Q000276:immunology; D012333:RNA, Messenger / Q000378:metabolism; D014019:Tissue Donors; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics
D000906:Antibodies; D051017:Apoptosis Regulatory Proteins; D000949:Histocompatibility Antigens Class II; D011506:Proteins; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; C109664:caspase-activated DNase inhibitor; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases