High levels of peroxynitrite are generated in the lungs of irradiated micegiven cyclophosphamide and allogeneic T cells - A potential mechanism of injury after marrow transplantation

In a murine bone-marrow transplant (BMT) model designed to determine risk f actors for lung dysfunction in irradiated mice, we reported that cyclophosp hamide (Cy)-induced injury and lethality depended on the infusion of donor spleen T cells. In the study reported here, we hypothesized that alveolar m acrophage (AM)-derived reactive oxygen/nitrogen species are associated with lung dysfunction caused by allogeneic T cells, which stimulate nitric oxid e ((NO)-N-.) production, and by Cy, which stimulates superoxide production. 'NO reacts with superoxide to form peroxynitrite, a tissue-damaging oxidan t. On Day 7 after allogeneic BMT, bronchoalveolar lavage fluid (BALF) obtai ned from mice injected with T cells contained increased levels of nitrite, which was associated with increased lactate dehydrogenase and protein level s, both of which are indices of lung injury. The injury was most severe in mice receiving both T cells and Cy. Messenger RNA (mRNA) for inducible nitr ic oxide synthase was detected only in murine lungs injected with T cells /- Cy. AMs obtained on Day 7 after BMT From mice receiving T cells +/- Cy s pontaneously generated between 20 and 40 mu M nitrite in culture, versus < 2 mu M generated by macrophages obtained from mice undergoing BMT but not r eceiving T cells. The level of 3-nitrotyrosine, the stable byproduct of the reaction of peroxynitrite with tyrosine residues, was increased in the BAL F proteins of mice injected with both T cells and Cy. We conclude that allo geneic T cells stimulate macrophage-derived 'NO, and that the addition of C y favors peroxynitrite formation. Peroxynitrite generation clarifies the de pendence of Cy-induced lung injury and lethality on the presence of allogen eic T cells.