Differentiation of class I-and class II-directed donor-specific alloreactivity in bronchoalveolar lavage lymphocytes from lung transplant recipients

Previous studies have demonstrated that donor antigen-specific primed-lymphocyte-test (PLT) reactivity of bronchoalveolar lavage lymphocytes is strongly associated with acute pulmonary rejection and with oblitera-tive bronchiolitis (OB); however, a systematic analysis of PLT reactivity as being class I-or II-directed has not been performed. To assess reactivity directed against individual class I or II antigens, we tested a total of 67 BAL-derived lymphocyte samples from 26 recipiente for alloreactivity in the PLT, using a pool of allogeneic cells and selected homozygous typing cells (HTCs) representing the HLA class I and II antigens expressed by the recipient and donor cells. The results obtained by PLT were correlated with the clinical statue of the recipient with regard to rejection, infection, and OB. In 9 of 10 cases where transbronchial biopsy results were consistent with rejection, donor antigen-specific allogeneic PLT reactivity was observed and, more specifically, could be determined to be directed toward donor class II antigen in 8 of these cases. For 3 of 4 recipients tested chronologically, positive donor antigen-specific PLT reactivity was observed at the time of and 2-3 V4 months prior to the diagnosis of rejection by transbronchial biopsy. During periods of acute infection, donor antigen-specific PLT reactivity was not observed; instead, nonspecific PLT reactivity of BAL-derived cells (i.e., reactivity that did not correlate with any defined HLA antigens) was observed as well as reactivity associated with the self-antigens expressed by the recipients' cells. The PLT reactivity of BAL-derived cells from a recipient diagnosed with OB correlated specifically with one of the disparate donor class I antigens (HLA-B44). In 23 cases, BAL cells were propagated in the presence of autologous cells and rIL-2, thereby allowing for sufficient numbers of cells to test with a panel of 29 HTCs and to analyze for cell surface phenotype. The cultured BAL cells from 4 recipients undergoing a rejection episode demonstrated a predominant CD4* phenotype consistent with the class II-directed reactivity observed in PLT. However, these results did not demonstrate a phenotype distinctive from the 7 BAL results obtained from 4 quiescent recipients. In marked contrast, the cultured BAL cells obtained from 4 recipients diagnosed with OB demonstrated a predominant CD8* phenotype, with 6092% of the cultured cells being CD8*. These results are consistent with the class I-directed reactivity observed in PLT. The results of these studies demonstrate the advantage of using HTCs to analyze the PLT reactivity of BAL-derived lymphocytes and to monitor the reactivity of chronological BAL lymphocyte populations as being class I-and/or class II-directed. Although previous studies have demonstrated donor antigen-directed reactivity associated with OB, our studies indicate more specifically that donor-specific class I-directed reactivity may be relevant to the development of OB. These studies are of import not only in the monitoring of recipients but also in the investigation of the immunologic basis of pulmonary disease.