Dynamics of cell-mediated immune responses to cytomegalovirus in pediatric transplantation recipients

Abstract

CMI responses, combined with quantification of CMV DNA (DNAemia), may identify transplantation recipients at risk for invasive disease. PBMC were collected in pediatric transplantation candidates at one, three, and six months post-transplant in 10 subjects (six renal, three cardiac, one stem cell) and at single time points in eight HC and 14 children greater than one yr post-transplant (LTTx). Cells were stimulated with anti-CD3mAb or CMV pp65 peptide pools and responses assessed by IFNG enzyme-linked immunosorbent spot assay and cytokine secretion. IFNG responses to anti-CD3mAb were significantly lower pretransplant relative to HC and were further decreased at one and three months post-transplant, but recovered to levels comparable to HC by six months. Responses to pp65 among CMV-seropositive recipients followed a similar pattern but recovered by three months. CMV-seropositive LTTx and HC showed a Th1 cytokine response to pp65 stimulation. Three LTTx subjects developed CMV DNAemia; two demonstrated decreased responses to anti-CD3mAB (and pp65 in the CMV seropositive subject) at the onset of DNAemia, which recovered as DNAemia resolved. Monitoring CMI in children is feasible and may provide an adjunct biomarker to predict CMV progression and recovery.

Fig. 1

IFNG responses in SFU/10⁵ PBMC by ELISPOT to CD3mAb (a) in the longitudinal cohort (pretransplant (n = 10) and then one month (n = 10), three months (n = 8), and six months (n = 7) post-transplant) and at single time points in a cross-sectional cohort of transplant subjects greater than one yr post-transplant (LTTx) (n = 14) and HC (n = 8). IFNG responses by ELISPOT to pp65 among CMV-seropositive subjects (b) in the longitudinal cohort (pretransplant (n = 6) and then one month (n = 6), three months (n = 4), and six months (n = 3) post-transplant) and at single time points in LTTx (n = 10) subjects and HC (n = 5). Each dot (●) represents the median SFU at a particular time point for an individual subject. In the longitudinal cohort, dots are color coded to show each subject’s median IFNG response across time.

Fig. 2

TH1 (a–c) and TH2 (d–g) cytokines (pg/mL) in PBMC culture supernatants following stimulation with CD3mAB in samples obtained from subjects pretransplant (n = 3), one month post-transplant (n = 3), six months post-transplant (n = 3), subjects greater than one yr post-transplant (LTTx) (n = 10), or HC (n = 5).

Fig. 3

TH1 (a–c) and TH2 (d–g) cytokines (pg/mL) in PBMC culture supernatants following stimulation with CMV pp65 peptide pools in samples obtained from renal transplant subjects more than one yr post-transplant (LTTx) (n = 10) or from HC (n = 5).

Fig. 4

Frequency of CD4+ and CD8+ cells among total lymphocytes by flow cytometry (right Y-axis) and absolute lymphocyte count (ALC) (left Y-axis) in LTTx subjects and HC.

Fig. 5

Longitudinal IFNG responses to CD3mAb and pp65 and CMV viral load in a HSC (a), cardiac (b), and renal (c) transplant subject.