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• W2330667071 abstract "Introduction: Cross-reactive anti-viral memory T cells constitute a significant proportion of the alloresponse and potentially play a pivotal role in mediating adverse post-transplant outcomes in HLA mismatched allografts. In addition, chronic DNA viruses such as cytomegalovirus (CMV) commonly reactivate and continue to influence episodes of post-transplant morbidity and mortality. This study identified a novel cross-reactive HLA-A2-restricted CMV-specific CD8+ T cell recognising both the CMV epitope NLVPMVATV (NLV) and the HLA-B27 alloantigen. We examine the functionality and TCR clonal distribution dynamics of cross-reactive NLV-specific CD8+ T cells both preceding and following CMV reactivation and their role in mediating allograft dysfunction. Method: NLV-specific CD8+ T cell expansion was performed on HLA-A2/CMV healthy donors (n=14) and LTR (n=11) by stimulating peripheral blood mononuclear cells with NLV peptide for 13 days. Allo-reactivity of NLV-specific CD8+ T cells was assessed using single expressing HLA class I-specific transfected cell lines and multiparameter flow cytometry which included A2/NLV-tetramer staining (specificity), IFN-γ production (cytolytic ability), CFSE (proliferation) and CD107a expression (cytotoxicity). Additionally, paired TCR variable (V) alpha and beta chains from the complementarity-determining region (CDR3) of cross-reactive NLV-specific CD8+ T cells were characterised using novel single-cell multiplex nested RT-PCR for simultaneous detection of V alpha and V beta chains. The dynamics of cross-reactive NLV-specific CD8+ T cells were then analysed in relation to the patient's CMV viral loads, lung function and acute rejection events. Results: NLV-specific CD8+ T cells from 3 healthy individuals underwent initial screening for allo-reactivity using cell lines encompassing 19 HLA-A and -B alleles. Cross-reactivity was observed in one individual based on proliferation, cytokine and cytolytic activity in response to B*2705 alloantigen, representing approximately 5% of the NLV-specific CD8+ T cell population. Further investigation elucidated a hierarchical response in allelic variation of B27 (B*2709>B*2705>B*2703). Similar patterns were observed in one LTR receiving a B27 allograft and we show that the cross-reactive A2/NLV-specific CD8+ T cells gradually increased (days 13-193 post-transplant) prior to a clinically detectable CMV reactivation event (day 270) and these cross-reactive cells reduced to basal levels following viral clearance on day 909. The overall NLV-specific TCR repertoire was dominated by one TCR clone (81%), but upon stimulation with HLA-B27, the main NLV-specific TCR repertoire shifted to 100% clonality for a second cross-reactive clonotype. However, following viral clearance the TCR repertoire greatly diversified in both the NLV (70%) and B27-cross-reactive pools (53%). Conclusion: Individualised monitoring of cross-reactive antiviral T cells and their TCR repertoire will provide further insights into the potential interaction between sub-clinical CMV reactivation and long term allograft outcomes." @default.
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• W2330667071 date "2012-11-01" @default.
• W2330667071 modified "2023-09-27" @default.
• W2330667071 title "Cross-Reactive CMV-Specific T Cells Significantly Increase in Frequency and TCR Clonality in the Setting of Viral Reactivation in Human Lung Transplantation" @default.
• W2330667071 doi "https://doi.org/10.1097/00007890-201211271-02322" @default.
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