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Interferon-gamma and interleukin-4 reciprocally regulate CD8 expression in CD8+ T cells - PubMed

  • ️Tue Jan 01 2008

Comparative Study

. 2008 Nov 11;105(45):17475-80.

doi: 10.1073/pnas.0809549105. Epub 2008 Nov 6.

Affiliations

Comparative Study

Interferon-gamma and interleukin-4 reciprocally regulate CD8 expression in CD8+ T cells

Simon H Apte et al. Proc Natl Acad Sci U S A. 2008.

Abstract

The CD8 co-receptor can modulate CD8(+) T cell function through its contributions to T cell receptor (TCR) binding and signaling. Here we show that IFN-gamma and IL-4 exert opposing effects on the expression of CD8alpha mRNA and surface CD8 protein during CD8(+) T cell activation. IL-4 caused down-regulation of surface CD8 on ovalbumin (OVA)(257-264)-specific TCR-transgenic OT-I CD8(+) T cells activated with OVA(257-264)-coated antigen presenting cells or polyclonal stimuli, and on wild type CD8(+) T cells activated with polyclonal stimuli. This effect was enhanced in each case when the cells lacked a functional IFN-gamma or IFN-gamma R gene. When WT or IFN-gamma-deficient OT-I CD8(+) T cells were analyzed 9 days after co-injection with control or IL-4-expressing OVA(+) tumor cells into RAG-2(-/-)gamma c(-/-) mice, CD8 levels were highest on WT donor cells from mice that received the control tumor and lowest on IFN-gamma-deficient donor cells from mice that received the IL-4-expressing tumor. The latter CD8(low) cells displayed markedly impaired binding of OVA(257-264)/MHC tetramers and peptide/MHC-dependent degranulation. The data reveal an unexpected role for IFN-gamma in tuning the CD8 co-receptor during primary CD8(+) T cell activation both in vitro and in vivo.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.

CD8 expression by activated CD8+ T cells is reduced in the presence of IL-4 and/or the absence of IFN-γ. CD8+ cells from OT-I (IFN-γ+/+) and OT-I x IFN-γ−/− (IFN-γ−/−) mice were stained with (A) anti-CD8α Ab and SIINFEKL tetramer or (B) Ab to CD44 and CD62L. The CD8 MFI and the percentage of double-positive cells (A), and the percentage of cells with a naïve (CD44low CD62Lhigh) phenotype (B), are shown within the frames. (C) The two CD8+ populations were cultured with anti-receptor Ab and IL-2 with or without IL-4 for 6 days then stained with anti-CD8α Ab (filled histograms) or isotype control Ab (open histograms). (D) The same CD8+ populations were cultured with SIINFEKL-coated APC or PMA and ionomycin, and IL-2 with or without IL-4. CD8 expression is shown as relative fluorescence intensity, obtained by normalizing the MFI to the highest sample in the experiment (100%). (E) CD8 expression by CD8+ cells from C57BL/6 WT (IFN-γ+/+), IFN-γR−/− or IFN-γ−/− mice was measured after culture with anti-receptor Ab, IL-2 and with or without IL-4 for 10 days. Groups were compared by unpaired t test (see Materials and Methods).

Fig. 2.
Fig. 2.

IL-4 and IFN-γ exert opposing dose-dependent effects on CD8 expression. (A and B) CD8+ cells from C57BL/6 WT (IFN-γ+/+), IFN-γR−/−, and IFN-γ−/− mice were cultured with anti-receptor Ab, IL-2 and the indicated concentrations of IFN-γ for 5 days. CD8α mRNA levels are expressed as β2M units (mean and SD, n = 3) in A, and as relative CD8α and CD8β mRNA expression (mean and SD, n = 4) in B. (C) In an independent experiment, CD8+ cells from C57BL/6 WT (IFN-γ+/+) and IFN-γ−/− mice were cultured with anti-receptor Ab, IL-2 and the indicated concentrations of IL-4 and IFN-γ for 5 days. Surface CD8 expression was analyzed by flow cytometry and expressed as relative fluorescence intensity (see legend to Fig. 1) (mean and SD, n = 5).

Fig. 3.
Fig. 3.

IL-4 and IFN-γ modulate CD8 expression in vivo. Vα2+ CD8+ cells from OT-I (IFN-γ+/+) or OT-I x IFN-γ−/− (IFN-γ−/−) mice were co-injected with EG7-IL-4+ (IL-4+) or EG7-IL-4− (Ctrl) tumor cells into RAG-2−/−γc−/− mice. After 9 days, splenic Vα2+ cells were isolated and analyzed for surface CD8 expression by flow cytometry. (A) Representative data from individual mice are shown. (B) Data from all mice in a representative experiment are shown. Each point represents one mouse; horizontal lines indicate the mean of each group. (C) CD8α mRNA expression was measured after restimulation in vitro with anti-receptor Ab for 6 days.

Fig. 4.
Fig. 4.

Modulation of CD8 levels by IL-4 and IFN-γ affects T cell function. The restimulated cell populations in Fig. 3C were co-stained with anti-CD8α Ab and (A) SIINFEKL tetramers at a dilution of 16.7 × 10−3 or (B) anti-Vα2 Ab. Values within the frames indicate the percentage of cells in that region. (C) The tetramer MFI is shown for each population following staining with the indicated tetramer dilutions in the absence of anti-CD8α Ab. (D) The restimulated cell populations were rested for 6 days then cultured with SIINFEKL or SIIGFEKL peptide-coated APC or with PMA, ionomycin and IL-2, in the presence of anti-CD107 Ab and monensin. (E) Upper frames show CD107 profiles of the indicated populations from (D); the open histogram in the left frame shows CD107 Ab binding to cells cultured with APC without peptide. Lower frames show corresponding CD8 profiles and CD8 MFI following gating on CD107− (dark) and CD107+ (light) cells.

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