Monday, September 27, 2010 2:00 PM - American Society for ...

69-PSENSITIZATION BY HLA-C CAN RENDER HLA-B ANTIGENS AS UNACCEPTABLEMISMATCHES.Marilyn Marrari, Rene J. Duquesnoy. Transplant Pathology, University of Pittsburgh Medical Center,Pittsburgh, PA, USA.Aim: Recent reports have shown that anti-HLA-C antibodies (Abs) are associated with rejection and graftfailure. DNA-based methods can accurately determine HLA-C types, and sensitive screening assays such asLuminex can identify HLA-C Abs. HLA-C antigens (Ags) display considerable amino acid polymorphismthat can be translated into a structurally defined epitope repertoire. HLAMatchmaker can be used toidentify donor-specific Ab reactivity patterns against mismatched critical components of epitopes (eplets)found on HLA-C Ags.Methods: We analyzed Ab reactivity patterns in sera from patients mismatched for HLA-C with rejectedrenal transplants that were screened in Luminex assays using class I single allele beads. Specificabsorptions were performed using microbeads bearing single class I alleles and eluates were tested byLuminex.Results: We recently reported a case of sensitization of a B*4403 patient to the 156DA epitope on anHLA-Cw*0704 mismatch which led to Abs reacting with the 156DA epitope-carrying B*4402 allele andother HLA-B Ags, including B*0801, B*3701, B*4101, B*4201, B*4501 and B*8201 (Hum Immun 71:176-8, 2010). We report here four more cases whereby absorption/elution assays clearly demonstrate thatAbs induced by a HLA-C mismatch reacted with epitopes shared with HLA-B Ags:[table1]Conclusions: Our findings demonstrate the importance of HLA-C mismatching in humoralallosensitization. Some HLA-C Abs are specific for epitopes shared with HLA-B Ags which then becomeunacceptable mismatches although the patient may have never been exposed to them. HLAMatchmaker isuseful in understanding donor-specifc Ab reactivity patterns and the determination of HLA mismatchacceptability for transplantation.70-PAUTOANTIBODIES AGAINST NUCLEOLIN IN KIDNEY TRANSPLANT RECIPIENTS.Zhiqiang Qin 1 , Bhavna Lavingia 2 , Yizhou Zou 1 , Peter Stastny 1,2 . 1 Internal Medicine - TransplantImmunology, UT Southwestern Medical Center, Dallas, TX, USA; 2 Pathology, UT Southwestern MedicalCenter, Dallas, TX, USA.Aim: Nucleolin is expressed on the surface of endothelial cells in the course of angiogenesis.Autoantibodies against nucleolin have been previously described in patients with chronic graft-versus-hostdisease.We investigated antibodies from kidney transplant recipients which react with endothelial cells.Methods: Sera from 50 kidney transplant recipients were investigated for antibodies against humanumbilical vein endothelial cells (HUVEC). One serum was used for immunoprecipitation and proteinidentification. Presence of nucleolin was confirmed with a nucleolin-specific monoclonal antibody.Nucleolin isolated from endothelial cell lysates was used for analysis of patient sera by Western blotting.The effect of antibodies against nucleolin on HUVEC was studied by assessing cell proliferation, apoptosisand in vitro tube formation.Results: Antibodies against nucleolin identified a band of approximately 100 kDa by SDS-PAGE. Resultsof mass spectrometry showed that sequences of nucleolin were identified. We tested 50 sera obtained fromkidney transplant recipients after rejection and found that 7 (14%) had antibodies against nucleolinenrichedpreparations obtained from endothelial cell lysates by Western blot analysis. The frequency ofsuch antibodies should be higher if a more sensitive assay is used. After incubation with antibodies againstnucleolin the number of HUVEC was decreased, possibly by apoptosis. Tube formation by endothelial cellsin vitro was inhibited.Conclusions: Autoantibodies against nucleolin were identified in kidney transplant recipients who hadrecently undergone immunological rejection of their allografts. Such antibodies have previously beenreported in bone marrow transplant recipients with chronic graft-versus-host-disease. The potential role ofthese autoantibodies in organ allograft rejection is suggested by their effect on angiogenesis and inhibitionof endothelial cell proliferation.71-P