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Flagellin-Specific CD4 Cytokine Production in Crohn Disease and Controls Is Limited to a Small Subset of Antigen-Induced CD40L+ T Cells

Nadine N. Morgan and Peter J. Mannon
J Immunol January 15, 2021, 206 (2) 345-354; DOI: https://doi.org/10.4049/jimmunol.2000918
Nadine N. Morgan
*Department of Medicine and Program in Immunology, University of Alabama at Birmingham, Birmingham, AL 35294; and
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Peter J. Mannon
†Birmingham VA Medical Center, Medical Service and Department of Medicine, University of Alabama at Birmingham, Birmingham, AL 35233
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Key Points

  • Only a small subset of flagellin-induced CD40LhiCD4 cells express cytokines.

  • This CD40LhiCD4 cytokine production is restricted to T effector memory cells.

Abstract

Flagellin is an immunodominant Ag in Crohn disease, with many patients showing anti-flagellin Abs. To study the clonality of flagellin-reactive CD4 cells in Crohn patients, we used a common CD154-based enrichment method following short-term Ag exposure to identify Ag-reactive CD4 cells. CD154 expression and cytokine production following Ag exposure compared with negative control responses (no Ag exposure) revealed that only a small fraction of CD154-enriched cells could be defined by Ag-reactive cytokine responses. This was especially true for low-frequency flagellin-reactive CD4 cells compared with polyclonal stimulation or Candida albicans Ag exposure. Moreover, we found that culture conditions used for the assay contributed to background CD40L (CD154) expression in the CD154-enriched CD4 cells. Using a cut-off rule based on flow cytometry results of the negative control CD154-enriched CD4 cells, we could reliably find the fraction of Ag-reactive cells in the CD154-enriched population. Ag-reactive CD4 cytokine production was restricted to CD4 cells with an effector memory phenotype and the highest levels of induced CD154 expression. This has important implications for identifying Ag-specific T cells of interest for single cell cloning, phenotyping, and transcriptomics.

Footnotes

  • This work was supported by U.S. Department of Veterans Affairs Grant CX0001530 and Impact funds from University of Alabama at Birmingham.

  • The online version of this article contains supplemental material.

  • Abbreviations used in this article:

    CD
    Crohn disease
    IBD
    inflammatory bowel disease
    TEM
    effector memory T cell.

  • Received August 6, 2020.
  • Accepted November 12, 2020.
  • Copyright © 2021 by The American Association of Immunologists, Inc.
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The Journal of Immunology: 206 (2)
The Journal of Immunology
Vol. 206, Issue 2
15 Jan 2021
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Flagellin-Specific CD4 Cytokine Production in Crohn Disease and Controls Is Limited to a Small Subset of Antigen-Induced CD40L+ T Cells
Nadine N. Morgan, Peter J. Mannon
The Journal of Immunology January 15, 2021, 206 (2) 345-354; DOI: 10.4049/jimmunol.2000918

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Flagellin-Specific CD4 Cytokine Production in Crohn Disease and Controls Is Limited to a Small Subset of Antigen-Induced CD40L+ T Cells
Nadine N. Morgan, Peter J. Mannon
The Journal of Immunology January 15, 2021, 206 (2) 345-354; DOI: 10.4049/jimmunol.2000918
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