CMV Antigen-Specific CD4<SUP>+</SUP> and CD8<SUP>+</SUP> T Cell IFNγ Expression and Proliferation Responses in Healthy CMV-Seropositive Individuals

Abstract

CMV-specific CD4 + and CD8 + T cell IFNγ expression and proliferation were measured in healthy volunteers by flow cytometry after CMV lysate or CMV pp65 or IE peptide pool stimulation. Cutoff values were set to maximize specificity (i.e., no false positive CMV-seronegatives). Sensitivity (defined as a positive response in CMV-seropositives to at least one of the 3 antigen preparations used) was 100% for CMV-specific CD4 + and CD8 + T cell IFNγ expression and CD4 + T cell proliferation and 95.4% for CMV-specific CD8 + T cell proliferation. All 22 CMV-seropositive individuals had positive responses by at least three of these four measurements. These findings support the concept that a multiplicity of antigen-specific functional immune responses and persistence of robust virus-specific CD4 + T cells are important components of protective immunity in this chronic viral infection.

Validation of T-Track® CMV to assess the functionality of cytomegalovirus-reactive cell-mediated immunity in hemodialysis patients

“…Interestingly, detection of CMV-reactive effector T cells within CMV-seronegative individuals has been described by others, both in healthy individuals and in transplant recipients, at frequencies of 2–11% among healthy donors and up to 30% in renal transplant recipients [ 13 , 63 – 65 ]. With 21% CMV-seronegative hemodialysis patients with positive T-Track® CMV results, our data are in concordance with these published studies.…”

“…Primary infection is dominated by CD8 + T cell response, preferentially targeting CMV immediate early-1 (IE-1) antigen, while long-term recovery is dominated by CD4 + T cell response and a switch of reactivity toward CMV lower matrix phosphoprotein 65 (pp65) [ 6 , 8 – 11 ]. The frequency of CMV-specific CD8 + and CD4 + T cells is highly variable, both between healthy CMV-seropositive individuals and during the course of CMV reactivation, and correlates with varying levels of protection [ 6 , 9 , 11 – 13 ]. Beside changes in T cell frequency, alterations in T cell functionality are associated with impaired response to chronic viral infection [ 14 – 17 ].…”

“…Principally, they measure the production of activation markers (e.g. cytokines such as IFN-γ) in response to antigen stimulation, using intracellular cytokine staining followed by flow cytometry [ 6 , 7 , 12 , 13 , 25 , 26 ], ELISA [ 21 , 27 – 30 ], or ELISpot [ 31 – 33 ] assays. These approaches differ not only in their read-out format but also in the antigen (peptide vs. protein) used for the ex vivo stimulation.…”

Humoral and Cellular CMV Responses in Healthy Donors; Identification of a Frequent Population of CMV-Specific, CD4+ T Cells in Seronegative Donors

“…The major advantage of using the pp65 peptide mixture compared to a lysate is that it allows CD4 and CD8 T cell responses to be examined in parallel, something that Sester et al could not do. Others have also successfully used pp65 derived peptide mixtures to examine CMV-specific CD8 T cell responses [26] , [27] .…”