Heart transplant rejection is characterized pathologically by myocyte necrosis and apoptosis associated with interstitial mononuclear cell infiltration. Any one of these components can be targeted for noninvasive detection of transplant rejection. During apoptotic cell death, phosphatidylserine, a phospholipid that is normally confined to the inner leaflet of cell membrane bilayer, gets exteriorized. Technetium-99m-labeled annexin-V, an endogenous protein that has high affinity for binding to phosphatidylserine, has been administered intravenously for noninvasive identification of apoptotic cell death. In the present study of 18 cardiac allograft recipients, 13 patients had negative and five had positive myocardial uptake of annexin. These latter five demonstrated at least moderate transplant rejection and caspase-3 staining, suggesting apoptosis in their biopsy specimens. This study reveals the clinical feasibility and safety of annexin-V imaging for noninvasive detection of transplant rejection by targeting cell membrane phospholipid alterations that are commonly associated with the process of apoptosis.

LOH analysis suggests that multiple tumor suppressor genes play a role in the development of human TCC. The human homolog of the Drosophila PTCH was recently cloned and mapped to the BCNS locus on 9q22.3, a chromosomal region commonly deleted in TCCs. We ®rst examined the steady state mRNA transcription of the PTCH, SMOH and GLI3 genes of the HH signal transduction pathway in TCC cell lines and normal urothelium. Normal urothelium and TCC cell lines express these three genes within the PTCH signal transduction pathway. We then screened for PTCH mutations in`hot spot' exons 6, 8, 13 and 16 by PCR/ SSCP analysis of genomic DNAs from 54 TCC tumor samples and control autologous peripheral blood lymphocytes. DNA sequence analysis con®rmed TCCspeci®c mutations in two of 54 patients (3.7%). These mutations resulted a single amino acid substitution and two frame shifts. One tumor had PTCH mutations in exon 16 as well as exon 13 and one tumor had a mutation in exon 13 alone. Both TCC tumors that contained PTCH mutations had a loss of heterozygosity at 9q. Although the PTCH protein has an unknown function in urothelial cells, the detection of the PTCH, SMOH and GLI3 transcripts in normal urothelium and TCC cell lines and rare PTCH mutations in tumor samples suggest that the HH pathway may have a role in controlling the proliferation of urothelial cells and that PTCH mutations may contribute to the development of a subset of TCCs.

A panel of 18 superficial or invasive transitional cell carcinomas (TCCs) was analyzed for chromosome 9 deletions by performing a high-density loss of heterozygosity (LOH) analysis. Twenty-five microsatellite loci were assayed by the polymerase chain reaction (PCR) and 7 restriction fragment length polymorphism (RFLP) loci were analyzed by Southern blotting. Concordant results were obtained with these methods, including direct comparisons at 2 loci. Chromosome 9 LOH was observed in 13 (72%) of 18 informative cases, including 10 superficial lesions. In contrast, LOH on chromosomes 10, 15, 20 and 21 was < or = 8%. Evidence for missing copies of chromosome 9 was observed in 7 of 13 LOH cases. Comparison of cases with subchromosomal LOH implicated the region between the D9S55 locus on 9p12 and the argininosuccinate synthetase (ASS) locus on 9q34.1 as the location of a tumor suppressor gene relevant to TCC.