IDENTIFICATION OF DIFFERENTIALLY EXPRESSED GENES IN RAT AORTIC ALLOGRAFT VASCULOPATHY

Graft vasculopathy is an important complication of longs-surviving org an transplants, but its pathogenesis has remained elusive. We investig ated rat aortic transplants with vasculopathy, aortic transplants with out vasculopathy, and normal aortas for differentially expressed mRNA transcripts to gain further insight into the molecular mechanisms invo lved. Aortic transplants were performed in allogeneic or syngeneic rec ipients followed by removal after 1 or 5 months, RNA isolation, and di fferential display to identify mRNA transcripts the expression of whic h was modulated in conjunction with the transplant procedure and the d evelopment of vasculopathy. Using 80 random primers, 57 differentially displayed polymerase chain reaction products were identified 18 of wh ich were found in allografts but not in syngeneic grafts or normal ves sels, whereas 15 were expressed in normal vessels and syngeneic grafts but not in allografts. Of the differentially displayed amplicons, 13 were successfully reamplified nad used as probes for Northern analysis ; differential expression was confirmed in 6 instances. DNA sequence a nalysis of these PCR products revealed identity with the immunoglobuli n J chain in 2 instances, the ferritin heavy chain, a sequence related but not identical with Ras, and an established sequence tag recently isolated from a human fetal heart library; 1 sequence was not related to arty known gene. To assess whether differential mRNA expression of the J-chain gene, a gene expressed in cells of B lymphocyte lineage, w as associated with infiltration of the graft by B lymphocytes, tissue sections were stained with art antibody against the B cell marker CD45 RA. Although the number of CD45RA-positive cells was low, there was a significant increase in the number of CD45RA-positive cells in the adv entitia and intima of grafts with vasculopathy. Furthermore, immunosta ining with anti-ferritin antiserum confirmed the presence of ferritin- positive cells within the inner layer of the graft vessel wall and dis persed in the intima, media, and adventitia. The question remains as t o which of these genes are critically relevant in the pathogenesis of graft vasculopathy and whether they serve as targets for therapeutic i nterventions.