title: Reactive oxygen and reactive nitrogen as signaling molecules for caspase 3 activation in acute cardiac transplant rejection.
abstract: Apoptosis is a significant factor in cardiac dysfunction and graft failure in cardiac rejection. In this study, we examined potential signaling molecules responsible for caspase 3 activation in a model of acute cardiac allograft rejection. The roles of reactive oxygen species (ROS) and nitric oxide (NO) were determined in untreated allografts and allograft recipients treated with either cyclosporine (CsA), alpha-phenyl-t-butylnitrone (PBN, a spin-trapping agent), vitamin C (VitC), Mn(III)tetrakis (1-methyl-4-pyridyl)porphyrin); MnTmPyP, a superoxide dismutase (SOD) mimetic), or L-(1-iminoethyl)lysine) (L-NIL), an inhibitor of inducible NO synthase (iNOS) enzyme activity. Graft tissue was taken for measuring superoxide radical production, Western blotting, and direct measurement of caspase 3 activity. Activation of caspase 3 in untreated allografts was revealed by the appearance of cleaved caspase 3 from pro-caspase 3 by Western blotting and functional caspase 3 catalytic activity. CsA or PBN inhibited iNOS expression and caspase 3 activity. VitC and MnTmPyP did not alter iNOS expression or decrease NO levels but did inhibit caspase 3 activity. In contrast, L-NIL completely inhibited the increase in NO production without altering iNOS expression and inhibited caspase 3 activity. The prevention of TUNEL staining by MnTmPyP and L-NIL confirmed downstream effects of superoxide and NO on apoptosis. These studies indicate that both superoxide and NO (precursors of peroxynitrite formation) play a significant role in caspase 3 activation in cardiac allograft rejection.
authors.affiliation: Division of Transplant Surgery, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA. gmpieper@mcw.edu
authors.forename: Galen M
authors.initials: GM
authors.lastname: Pieper
authors.forename: Vani
authors.initials: V
authors.lastname: Nilakantan
authors.forename: Thanh K
authors.initials: TK
authors.lastname: Nguyen
authors.forename: Gail
authors.initials: G
authors.lastname: Hilton
authors.forename: Allan M
authors.initials: AM
authors.lastname: Roza
authors.forename: Christopher P
authors.initials: CP
authors.lastname: Johnson
mesh_terms: D000818:Animals; D017209:Apoptosis; D053148:Caspase 3 / Q000378:metabolism*; D004789:Enzyme Activation; D006084:Graft Rejection / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D016027:Heart Transplantation / Q000473:pathology; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism; D051381:Rats; D011917:Rats, Inbred Lew; D011920:Rats, Inbred WF; D026361:Reactive Nitrogen Species / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; D052247:Nitric Oxide Synthase Type II; D053148:Caspase 3