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Simultaneous Th1-Type Cytokine Expression Is a Signature of Peritoneal CD4⁺ Lymphocytes Responding to Infection with Listeria monocytogenes ¹

Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, GA 30329

The robust murine response to infection with Listeria monocytogenes makes an excellent model to study the functional development of immune cells. We investigated the cellular immune response to i.p. infection using intracellular cytokine staining to identify Ag-specific lymphocytes. CD4⁺ peritoneal exudate cells obtained 10 days postinfection predominantly coexpressed TNF-, IFN-, and IL-2 after polyclonal or Ag stimulation. A population of cells simultaneously making TNF- and IFN- was also detected but at a lower frequency. By following the kinetics of the response to Listeria, we found that CD4⁺ lymphocytes coexpressing TNF- and IFN- dominated on day 6 postinfection and then declined. From days 10–27, TNF-⁺IFN-⁺IL-2⁺ (triple-positive) was the most prevalent cytokine phenotype, and the frequency steadily declined. These characteristic cytokine expression patterns were observed in both primary and secondary responses to Listeria infection and developed even when infection was terminated with antibiotic treatment. A cytokine-assisted immunization procedure resulted in both double- and triple-positive cells, but the clear predominance of triple-positive cells required Listeria infection. Triple-positive cells were preferentially noted in the peritoneal cavity tissue site; spleen cells displayed a predominant population of double-positive T cells (TNF-⁺IFN-⁺). We speculate that the appearance of triple-positive cells represents a functionally significant subset important in host defense at nonlymphoid tissue sites.

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