Unbiased multiparameter analysis identifies increased immunosenescence of alloreactive CD8+ T cells in association with donor-specific hyporesponsiveness

A.C.J. van der List, N.H.R. Litjens, M. Klepper, M.G.H. Betjes

Wednesday 4 march 2020

14:40 - 14:50h at Joep Nicolas zaal

Parallel session: Parallel sessie IV – Basale abstracts

Background: The risk of T-cell mediated rejection (TCMR) progressively declines in the years after kidney transplantation (KT) in most recipients. This is reflected by a decreased proliferative response of recipient T cells to donor antigen presenting cells, a phenomenon called anti-donor-specific hypo-responsiveness (DSH). The onset of DSH is more rapid and pronounced in older recipients (55+), indicative of immunosenescence/exhaustion of T lymphocytes as a contributing factor.

Methods: To test this hypothesis, expression profiles of markers associated with immunosenescence were investigated within anti-donor-specific T cells of elderly stable kidney transplant recipients before and 3-5 years after KT . Anti-donor-specific T cells were identified as CD137 positive T cells upon stimulation with donor cells and further characterized by flow cytometry using a broad panel of known markers of immunosenescence/exhaustion. Traditional analysis of flow data by visual inspection of 2D scatterplots is biased and inefficient as the number of plots increases exponentially with the amount of markers used. This problem was circumvented through an unsupervised (and therefore unbiased) clustering and dimensionality reduction technique called Flow Self-Organizing Map (FlowSOM).

Results: All recipients demonstrated DSH 3-5 years after transplantation. Within the heterogeneous pool of CD137-expressing anti-donor specific T cells, clusters of CD4+ and CD8+ T-cells with a high fold-change of cell numbers between pre- and post-transplant time points were identified and confirmed by traditional flow cytometry analysis. Clusters of CD8+CD137+ T cells highly expressing exhaustion markers CD244 and TIGIT and immune checkpoint inhibitor, CD160, make up a larger proportion of CD8+CD137+ T lymphocytes post KT. In contrast, clusters of CD4+CD137+ T cells highly expressing TIGIT with or without exhaustion marker TIM3, decrease post Tx. One CD4+CD137+ T cell cluster which decreases post KT deviates from this trend, as it has no TIGIT expression but does express the immune checkpoint inhibitor PD1.

Conclusions: The results demonstrate the richness of data obtained by unbiased analysis of immunosenescence/exhaustion expression profiles as compared to classical 2D analysis. The results indicate that immunosenescence/exhaustion may be an important underlying mechanism causing DSH in the alloreactive CD8 T cells.