UP-REGULATION AND MODULATION OF INDUCIBLE NITRIC-OXIDE SYNTHASE IN RAT CARDIAC ALLOGRAFTS WITH CHRONIC REJECTION AND TRANSPLANT ARTERIOSCLEROSIS

Background The Lewis-F344 rat cardiac transplantation model produces c ardiac allografts with chronic rejection characterized by arterioscler otic lesions composed of macrophages and smooth muscle cells. Modulati on of the inflammatory response with a diet deficient in essential fat ty acids protects against the development of intimal thickening. Littl e is known about the components of the inflammatory response mediating this process. The cytokine-inducible isoform of nitric oxide synthase (iNOS) regulates the high-output nitric oxide pathway that confers ac tivation properties to macrophages and regulates vasomotion, monocyte adherence, and smooth muscle cell proliferation in the vasculature. Th e purpose of the present study was to determine whether the iNOS pathw ay was upregulated during the course of chronic cardiac rejection. Met hods and Results We studied iNOS mRNA and protein expression patterns in a series of Lewis-F344 cardiac allografts with early and late chron ic rejection and after modulation of the inflammatory response (in an effort to attenuate arteriosclerosis). Relative gene transcript levels were measured with a P-32-dCTP reverse-transcriptase polymerase chain reaction assay designed to amplify iNOS mRNA. The distribution of the iNOS gene product was examined by immunocytochemistry with a polyclon al antibody against iNOS. NOS transcript levels increased significantl y in cardiac allografts (days 7, 14, 28, and 75) compared with paired host hearts (exposed to the same circulation) and syngrafts (P<.003). Immunostaining localized the iNOS antigen within subpopulations of mon onuclear inflammatory cells in cardiac allografts-presumably, activate d macrophages. The number of iNOS-positive mononuclear cells was 25-fo ld higher in cardiac allografts compared with paired host hearts and s yngrafts (P<.009). In cardiac allografts of 75 days or older, there al so was striking iNOS staining within some medial and intimal smooth mu scle cells in various vessels. Modulation of the inflammatory response (with a diet deficient in essential fatty acids) produced significant decreases in the intimal thickening score and in the percentage of di seased vessels in 28-day cardiac allografts compared with allografts f rom rats fed a control diet. There was a correlate decrease in iNOS tr anscript levels and in the number of iNOS-positive mononuclear cells i n the 28-day cardiac allografts from rats fed the essential fatty acid -deficient diet. Conclusions The early and persistent upregulation of iNOS in chronic cardiac rejection and the coincident reduction in arte riosclerosis and downregulation of iNOS suggest that this inducible re gulator may contribute to the inflammatory response mediating transpla nt arteriosclerosis.