Cytomegalovirus-Responsive CD8+ T Cells Expand After Solid Organ Transplantation in the Absence of CMV Disease

Abstract

Cytomegalovirus (CMV) is a major cause of morbidity and mortality in solid organ transplant recipients. Approximately 60% of adults are CMV seropositive, indicating previous exposure. Following resolution of the primary infection, CMV remains in a latent state. Reactivation is controlled by memory T cells in healthy individuals; transplant recipients have reduced memory T cell function due to chronic immunosuppressive therapies. In this study, CD8⁺ T cell responses to CMV polypeptides immediate-early-1 and pp65 were analyzed in 16 CMV-seropositive kidney and heart transplant recipients longitudinally pretransplantation and posttransplantation. All patients received standard of care maintenance immunosuppression, antiviral prophylaxis, and CMV viral load monitoring, with approximately half receiving T cell-depleting induction therapy. The frequency of CMV-responsive CD8⁺ T cells, defined by the production of effector molecules in response to CMV peptides, increased during the course of 1 year posttransplantation. The increase commenced after the completion of antiviral prophylaxis, and these T cells tended to be terminally differentiated effector cells. Based on this small cohort, these data suggest that even in the absence of disease, antigenic exposure may continually shape the CMV-responsive T cell population posttransplantation.

Keywords: T cell biology; basic (laboratory) research/science; flow cytometry; immune regulation; immunobiology; infection and infectious agents; infectious disease; viral: Cytomegalovirus (CMV).

Figure 1. Pre-transplant CD8⁺ T cell responses to CMV polypeptides vary

A) Time course of blood collection and patient treatment. B) IE-1 stimulated PBMC gated for CD8⁺ T cells were stained for IFNγ, TNFα, and CD107a. Gates were defined based on an unstimulated control for that sample. Boolean gates were used to identify combinations of expression for the three functional markers. C) % polyfunctional of CD8⁺ T cells in response to IE-1 (black circles) and pp65 (white squares) in all recipients pre-transplant. Lines represent median and error bars represent interquartile range. n=16.

Figure 2. Polyfunctional CMV-responsive CD8⁺ T cells tend to be terminally differentiated effectors

A) Total and polyfunctional CD8⁺ T cells were gated for CCR7 and CD45RO expression. B) Proportion of polyfunctional CD8⁺ T cells in each memory/effector subpopulation pre-transplant. Bars stack to a total of 100% of polyfunctional cells. n=15. C) Proportion of polyfunctional CD8⁺ T cells that are effector (CD45RO⁻CCR7⁻, black squares) or effector memory (CD45RO⁺CCR7⁻, gray triangles) phenotype in response to IE-1 or pp65. Trends analyzed using mixed-effects generalized linear modeling in STATA. There were no statistically significant differences. Lines represent means and error bars represent SEM. # PT represents the number of patients at each time point. Not all recipients had samples at all time points, resulting in variable n.

Figure 3. The frequency of CMV-responsive T cells increases post-transplant

Percent of CD8⁺ T cells that are polyfunctional in response to IE-1 plus pp65 averaged in all transplant recipients (closed black circles) and normal donor controls (closed gray square). n= # PT as indicated, 7 normal donors. Values represent means and error bars represent SEM. Trends and pairwise comparisons were analyzed using mixed-effects modeling.

Figure 4. The post-transplant increase in CMV-responsive T cells is antigen-specific and independent of lymphodepletion

Percent of CD8⁺ T cells that are polyfunctional in response to IE-1 plus pp65 at each time point. A) Recipients divided between those who received lymphodepletion (left, triangles) and non-lymphodepleting induction (right, circles). B) Percent polyfunctional in response to IE-1 (open bars) or pp65 (gray bars). C) Percent of CD8⁺ T cells that are CMV-responsive (squares) or CMV non-responsive effectors (circles). For each population, the percentage is normalized to that determined at at day 0. Values represent means and error bars represent SEM. n = # PT as indicated. Trends and pairwise comparisons analyzed using mixed modeling on non-normalized data.