Decreased frequency of interferon-gamma- and interleukin-2-producing cells in patients with atopic diseases measured at the single cell level.
BACKGROUND
Recently, diminished interferon-gamma (IFN-gamma) and increased interleukin (IL-4 production in peripheral blood mononuclear cells (PBMCs) from atopic patients have been described by several groups, measured as total cytokine content in culture supernatants. These studies suggested a predominance of TH2-like cells producing large amounts of IL-4 in atopic patients. It is not clear whether the reported cytokine imbalances are the result of an alteration in the distribution of specific T-cell subsets or whether intrinsic dysregulation in cytokine production is a characteristic of atopic individuals.


OBJECTIVE
This study examined the production of IFN-gamma, and IL-2 in PBMCs from atopic patients at the single cell level with the use of freshly isolated lymphocytes.


METHODS
We recently described a flow cytometric assay in which three-color analysis was used to study the production of a cytokine of interest in a T-cell subpopulation defined by two cell surface markers. PBMCs from 23 atopic patients and 14 control subjects were stimulated with phorbol ester and ionomycin for 5 hours. PBMCs from seven patients and seven control subjects were also cultured with immobilized anti-CD3 antibodies for 24 hours. Cells were fixed, made permeable, and stained for intracellular cytokines in combination with cell surface markers CD3, CD8, and CD45RO. Cytokine-producing cells were analyzed by gating on T-cell subsets.


RESULTS
IFN-gamma-producing cells were significantly decreased (p < 0.05) in CD4+ T cells but not in CD8+ T cells of atopic patients. CD45R0+ and CD45R0-T cells showed a decreased proportion of IFN-gamma-producing cells (p < 0.05 and p < 0.01, respectively). IL-2 production was diminished in all T-cell subsets (p < 0.01). The number of IL-4-producing cells was not elevated, and such cells were exclusively found in the CD45RO+ T cells. Analysis of culture supernatants of sorted CD45RO+ T cells for IL-4 and IFN-gamma production confirmed these results.


CONCLUSIONS
Our findings provide evidence that a reduced IFN-gamma production in atopic patients is due to an intrinsic defect selectively found in the CD4+ T cells. Because IL-2 production was markedly decreased but IL-4 production was unchanged, our data demonstrate a deficiency in the ability of atopic T cells to produce TH1-like cytokines on stimulation with phorbol ester, ionomycin, or anti-CD3 monoclonal antibodies.
Jung
T
T
Department of Dermatology, University of Göttingen, Germany.
Lack
G
G
Schauer
U
U
Uberück
W
W
Renz
H
H
Gelfand
E W
EW
Rieger
C H
CH
D000293:Adolescent; D000328:Adult; D002452:Cell Count; D002648:Child; D002675:Child, Preschool; D005434:Flow Cytometry; D006801:Humans; D006967:Hypersensitivity / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D007223:Infant; D007371:Interferon-gamma / Q000378:metabolism*; D007376:Interleukin-2 / Q000378:metabolism*; D015847:Interleukin-4 / Q000378:metabolism; D008214:Lymphocytes / Q000378:metabolism*; D009000:Monocytes / Q000378:metabolism* / Q000473:pathology; D012016:Reference Values; D016176:T-Lymphocyte Subsets / Q000378:metabolism
D007376:Interleukin-2; D015847:Interleukin-4; D007371:Interferon-gamma