Activation of apoptotic and inflammatory pathways in dysfunctional donor hearts

Background. Myocardial dysfunction is common after brain death, but the mec hanisms remain unclear. Apoptosis is tightly regulated by enzymes termed th e caspases. We have investigated the caspases involved in the terminal part of the apoptotic pathway in dysfunctional (nontransplanted) donor hearts a nd 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 enz yme inactivated by caspase-3. Caspase-3 activity was also measured. Histolo gic and immunocytochemical analysis for HLA Class II and Real Time polymera se chain reaction for tumor necrosis factor-alpha and interleukin 6 were pe rformed 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 wa s 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 donor s, 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 n uclease 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. 0.6 1+/-0.22 O.D. units, P=0.57) donors. Conclusions. The caspases are elevated in dysfunctional donor hearts compar ed with hearts with good ventricular function with a possible link to infla mmatory activation supporting the concept that brain death causes inflammat ory activation which can lead to apoptosis with a possible important effect on function.