Post-transplant lymphoproliferative disorder (PTLD) continues to be a serious problem in clinical transplantation. PTLD represents a spectrum of abnormal lymphoproliferations but most originate in B cells and are associated with Epstein Barr virus (EBV). A major contributing factor for PTLD is the impaired anti-viral immunity that results from post-transplant immunosuppression. Still, we lack a mechanistic understanding of why some patients, but not others, develop EBV+ PTLD. One possibility is that patients at risk of EBV-associated PTLD have distinct defects in their immune response to EBV.

In order to more extensively probe the immune response to EBV in transplant patients we have utilized a novel cytometry platform, Cytometry Time of Flight (CyTOF). CyTOF utilizes heavy metal ion tagged antibodies for mass cytometry that permits simultaneous analysis of 35+ phenotypic and functional parameters on a single, EBV-specific cell. PBMC from 18 pediatric liver transplant recipients and 3 healthy donors were isolated from peripheral blood and stimulated with peptide pools containing 15mer peptides with 11 aa overlapping sequences covering the entire length of lytic cycle (BZLF1, BMLF1) and latent cycle (EBNA1, EBNA3a, LMP1 and LMP2) EBV proteins. CyTOF analysis revealed robust responses of CD4 and CD8 T cells in healthy, seropositive donors to EBV latent and lytic peptides as evidenced by IL-2, IFN-Γ, TNF-Α, MIP-1Β production and CD107a mobilization. In contrast, transplant recipients showed diminished CD4 T cell responses compared to CD8 T cell responses to both lytic and latent peptides. In addition, CD4 and CD8 T cells were unable to respond to lytic peptides in a subset of patients and mobilization of CD107a was impaired. Overall, patients who were seropositive before transplant mounted stronger EBV-specific T cell responses than patients who seroconverted post-transplant. Thus, specific defects in the T cell response to EBV can be identified in transplant recipients and may be associated with increased risk of EBV-associated disease.

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