Journal Article

A minority of proliferating human CD4+ T cells in antigen-driven proliferation assays are antigen specific

Chiu, Chris Y., Foster, Abby, Lacorcia, Matthew, Pakusch, Miha, Mannering, Stuart I., Bhattacharjee, Pushpak, Cameron, Fergus J., Liu, Xin, King, Laura, Tresoldi, Eleonora

2024

Overview

Antigen-driven T-cell proliferation is often measured using fluorescent dye dilution assays, such as the CFSE-based proliferation assay. Dye dilution assays have been powerful tools to detect human CD4 + T-cell responses, particularly against autoantigens. However, it is not known how many cells within the proliferating population are specific for the stimulating antigen. Here we determined the frequency of CD4 + T cells specific for the stimulating antigen within the antigen-responsive population of CFSE-based proliferation assays. We compared CD4 + T-cell responses to a type 1 diabetes autoantigen (proinsulin C-peptide) and to a vaccine antigen (tetanus toxoid). The TCRs expressed by antigen-responsive CD4 + T cells were sequenced, and their antigen specificity was tested functionally by expressing them in a reporter T-cell line. Responses to C-peptide were weak, but detectable, in PBMC from individuals with T1D, whereas responses to tetanus toxoid were much stronger. The frequency of antigen-specific CD4 + T cells correlated with the strength of the response to antigen in the proliferation assay. However, antigen-specific CD4 + T cells were rare among antigen-responsive CD4 + T cells. For C-peptide, an average frequency of 7.5% (1%–11%, n = 4) of antigen-responsive CD4 + T cells were confirmed to be antigen specific. In the tetanus-toxoid-stimulated cultures, on average, 45% (16%–78%, n = 5) of the antigen-responsive CD4 + T cells were tetanus toxoid specific. These data show that antigen-specific CD4 + T cells are a minority of the cells that proliferate in response to antigen and have important implications for in vitro CD4 + T-cell proliferation assays.

• Publisher: Frontiers Media SA,Frontiers Media S.A
• Subject: Adult / antigen specific T cell / Antigens / Autoantigens / Autoantigens - immunology / autoimmunity / C-Peptide / CD4 antigen / CD4+ T cell / CD4-Positive T-Lymphocytes - immunology / Cell Proliferation / clonal expansion / Cloning / Diabetes mellitus (insulin dependent) / Diabetes Mellitus, Type 1 - immunology / Dyes / Epstein-Barr virus / Female / Flow cytometry / Fluorescent indicators / Genes / Humans / Hypothesis testing / Immunology / Insulin / Lymphocyte Activation - immunology / Lymphocytes / Lymphocytes T / Male / Peptides / Peripheral blood mononuclear cells / Plasmids / proliferation / Tetanus / Tetanus Toxoid - immunology / Vaccines / Young Adult