Frequencies and TCR Repertoires of Human 2,4,6-Trinitrobenzenesulfonic Acid-specific T Cells

Abstract

Allergic contact dermatitis is a widespread T cell-mediated inflammatory skin disease, but in vitro monitoring of chemical-specific T cells remains challenging. We here introduce short-term CD154/CD137 upregulation to monitor human T cell responses to the experimental sensitizer 2,4,6-trinitrobenzenesulfonic acid (TNBS). Peripheral blood mononuclear cells (PBMC) from healthy donor buffy coats were TNBS-modified and incubated with unmodified PBMC. After 5 and 16 h, we detected TNBS-specific activated CD154+CD4+ and CD137+CD8+ T cells by multi-parameter flow cytometry, respectively. Activated cells were sorted for restimulation and bulk T cell receptor (TCR) high-throughput sequencing (HTS). Stimulation with TNBS-modified cells (3 mM) induced CD154 expression on 0.04% of CD4+ and CD137 expression on 0.60% of CD8+ memory T cells, respectively (means, n = 11–17 donors). CD69 co-expression argued for TCR-mediated activation, which was further supported by TNBS-specific restimulation of 10/13 CD154+CD4+ and 11/15 CD137+CD8+ T cell clones and lines. Major histocompatibility complex (MHC) blocking antibodies prevented activation, illustrating MHC restriction. The high frequencies of TNBS-specific T cells were associated with distinct common changes in the TCR β-chain repertoire. We observed an overrepresentation of tryptophan and lysine in the complementarity determining regions 3 (CDR3) (n = 3–5 donors), indicating a preferential interaction of these amino acids with the TNBS-induced epitopes. In summary, the detection of TNBS-specific T cells by CD154/CD137 upregulation is a fast, comprehensive and quantitative method. Combined with TCR HTS, the mechanisms of chemical allergen recognition that underlie unusually frequent T cell activation can be assessed. In the future, this approach may be adapted to detect T cells activated by additional chemical sensitizers.

CD8+ T cell clonotypes from prior SARS-CoV-2 infection predominate during the cellular immune response to mRNA vaccination

“…Amongst these, CD154 is seldom observed on activated CD8 + T cells (as tested herein, Supplemental Data Figs. 4, 5, and reported 48,49 ), such that congruency for the other markers was likely essential. The markers used for CD4 + T cells included CD200, differing from our use of CD69 and CD137.…”

Unique and common TCR repertoire features of Ni2+‐, Co2+‐, and Pd2+‐specific human CD154 + CD4+ T cells

“…The present study illustrates that Ni 2+ ‐, Co 2+ ‐, and Pd 2+ ‐specific CD4+ T cells can be detected by a CD154‐based AIM assay. AIM assays have been employed in the characterization of protein‐specific T cells but are not yet broadly established for chemical allergens 16,28,57,58,59,60 . The advantages of AIM assays include their speed, comprehensiveness, and sensitivity 28,61‐65 .…”

“…This finding argues against a preferred metal ion binding site. The activation of TCR via their CDR3 had been hypothesized and modelled for organic chemical sensitizers and was recently demonstrated for 2,4,6‐trinitrobenzenesulfonic acid (TNBS)‐specific T cells 60,87,88 . The abundance of the identified TCR repertoire features varied among individuals but combined data argued for a possible involvement in allergic immune responses.…”

“…The activation of TCR via their CDR3 had been hypothesized and modelled for organic chemical sensitizers and was recently demonstrated for 2,4,6-trinitrobenzenesulfonic acid (TNBS)-specific T cells. 60,87,88 The abundance of the identified TCR repertoire features varied among individuals but combined data argued for a possible involvement in allergic immune responses. However, the in vivo relevance of the identified metal-specific TCR requires further verification, for example, by showing a local enrichment in tissues affected by allergic reactions.…”

“…AIM assays have been employed in the characterization of proteinspecific T cells but are not yet broadly established for chemical allergens. 16,28,57,58,59,60 The advantages of AIM assays include their speed, comprehensiveness, and sensitivity. 28,[61][62][63][64][65] AIM assays are quantitative and do not require prior knowledge on the involved HLA allele, the presented antigen or self-peptide, and the chemical modifications at the TCR-self-peptide-MHC interface.…”