Evaluation of human cytomegalovirus-specific CD8+ T-cells in allogeneic haematopoietic stem cell transplant recipients using pentamer and interferon-γ-enzyme-linked immunospot assays

Abstract

BackgroundReactivation of latent human cytomegalovirus (HCMV) is a frequent complication following allogeneic haematopoietic stem cell transplantation (HSCT). Evaluation of the quantity and function of HCMV-specific CD8+ T-cell responses after HSCT may play a crucial role in the prevention of HCMV reactivation. ObjectivesTo investigate the mechanism of HCMV-specific T-cell immune responses after HSCT in HCMV-specific CD8+ T cells. Study designHCMV-specific CD8+ T cells were quantified using human leucocyte antigen (HLA) pentamer staining and functionally analysed by interferon-γ-enzyme-linked immunospot (IFN-γ-ELISPOT) assay with a pp65495–503 peptide in recipients four years after HSCT. ResultsThe absolute number of pp65495–503-specific CD8+ T cells did not differ significantly (p>0.05) between samples with antigenaemia and those without antigenaemia given a mean of 54.5/μl and 40.5/μl, respectively, in 21 HLA-A* 0201 patients after HSCT. The level of pp65495–503-specific CD8+ T cells>20/μl of peripheral blood was maintained 90 days after transplantation. There was a significant difference in the spot count of IFN-γ-secreting T cells between samples with antigenaemia (mean, 507/2.5×105PBMCs) and those without antigenaemia (mean, 216/2.5×105PBMCs; p<0.05). Conclusionpp65495–503-specific CD8+ T cells may not be sufficient to control HCMV reactivation in recipients after HSCT. However, the combination of pentamer and IFN-γ-ELISPOT assays may be valuable for evaluating HCMV-specific CD8+ T cells. Further studies on HCMV-specific T-cell immune responses continue to be performed for the prevention of persistent HCMV reactivation.