Counterregulation between the FcepsilonRI pathway and antiviral responses in human plasmacytoid dendritic cells - PubMed (original) (raw)

Counterregulation between the FcepsilonRI pathway and antiviral responses in human plasmacytoid dendritic cells

Michelle A Gill et al. J Immunol. 2010.

Abstract

Plasmacytoid dendritic cells (pDCs) play essential roles in directing immune responses. These cells may be particularly important in determining the nature of immune responses to viral infections in patients with allergic asthma as well those with other atopic diseases. The purposes of this study were 1) to compare the functional capacity of pDCs in patients with one type of allergic disorder, allergic asthma, and controls; 2) to determine whether IgE cross-linking affects antiviral responses of influenza-exposed pDCs; and 3) to determine whether evidence of counterregulation of FcepsilonRIalpha and IFN-alpha pathways exists in these cells. pDC function was assessed in a subset of asthma patients and in controls by measuring IFN-alpha production after exposure of purified pDCs to influenza viruses. FcepsilonRIalpha expression on pDCs was determined by flow cytometry in blood samples from patients with allergic asthma and controls. pDCs from patients with asthma secreted significantly less IFN-alpha upon exposure to influenza A (572 versus 2815; p = 0.03), and secretion was inversely correlated with serum IgE levels. Moreover, IgE cross-linking prior to viral challenge resulted in 1) abrogation of the influenza-induced pDC IFN-alpha response; 2) diminished influenza and gardiquimod-induced TLR-7 upregulation in pDCs; and 3) interruption of influenza-induced upregulation of pDC maturation/costimulatory molecules. In addition, exposure to influenza and gardiquimod resulted in upregulation of TLR-7, with concomitant downregulation of FcepsilonRIalpha expression in pDCs. These data suggest that counterregulation of FcepsilonRI and TLR-7 pathways exists in pDCs, and that IgE cross-linking impairs pDC antiviral responses.

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Figures

Figure 1

Figure 1

Purified pDCs from patients with allergic asthma secrete less IFN-α upon exposure to influenza viruses. Blood pDCs were isolated from patients with allergic asthma (n = 8) and controls (n = 9) and exposed to influenza A or B for 36 h. Mean (± SEM) IFN-α concentrations measured in culture supernatants are displayed in A (influenza A) and B (influenza B). Results of unpaired t tests are displayed.

Figure 2

Figure 2

pDCs from patients with allergic asthma express increased surface FcεRIα, and this correlates inversely with influenza-induced IFN-α secretion. A, FcεRIα expression on pDCs was determined by flow cytometry (expressed as MESF) in the allergic asthma patient group and healthy control subjects. Results of t tests are displayed. Correlation between influenza A- and B-induced IFN-α release and pDC FcεRIα expression was determined (B and C, respectively). Significant inverse correlations are demonstrated for both influenza A- and influenza B-induced IFN-a secretion and pDC FcεRIα expression. Results of Pearson correlation tests are shown. Correlation between influenza A- and B-induced IFN-α release and serum IgE concentration was determined (D and E, respectively). Significant inverse correlations are demonstrated for both influenza A- and influenza B-induced IFN-α secretion and pDC FcεRIα expression. Results of Pearson correlation tests are shown.

Figure 3

Figure 3

IgE cross-linking inhibits influenza-induced secretion of IFN-α in pDCs. Purified pDCs were cultured for 18 h with influenza A or influenza B, either alone or after preincubation with anti-IgE or rabbit IgG (as a control Ab), and IFN-α was measured in the supernatants. Data are expressed as mean ± SEM. ANOVA, p < 0.001 for A and B; results of Mann–Whitney t tests are displayed.

Figure 4

Figure 4

Exposure to influenza viruses and gardiquimod (a TLR-7 agonist) upregulates TLR-7 expression and downregulates FcεRIα expression in pDCs. A and B, Purified pDCs were cultured for 8 h with control media, influenza A, influenza B, or gardiquimod. Cells were harvested for RNA extraction and subsequent quantification of TLR-7 (A) or FcεR1α (B) mRNA by real-time RT-PCR, using hypoxanthine-guanine phosphoribosyltransferase as an endogenous control. Data are expressed as the mean ± SEM percentages of control pDC 2−ΔΔct values. Results represent five individual experiments. ANOVA, p < 0.01 for A and B. The p values displayed represent paired t test results. C, Purified pDCs were cultured with anti-IgE (5 μg/ml) or rabbit IgG (5 μg/ml) for 8 h and harvested for RNA extraction. TLR-7 mRNA was quantified as above. Data are expressed as the mean ± SEM percentages of control pDC 2−ΔΔct values of three individual experiments. ANOVA, p < 0.01; displayed p values represent results of unpaired t tests.

Figure 5

Figure 5

IgE cross-linking interferes with influenza- and gardiquimod-induced expression of TLR-7 in pDCs. A_–_C, Purified pDCs were cultured for 8 h with influenza A (A), influenza B (B), or gardiquimod (C), either alone or after preincubation with anti-IgE (5 μg/ml) or rabbit IgG (5 μg/ml). Cells were harvested for RNA extraction, generation of cDNA, and quantification of TLR-7 mRNA by RT-PCR. Hypoxanthine-guanine phosphoribosyltransferase was used as an endogenous control. Data are expressed as the mean ± SEM percentages of (A) influenza A-, (B) influenza B-, or (C) gardiquimod-stimulated pDC 2−ΔΔct values. Results represent seven individual experiments. ANOVA, p < 0.01 for A_–_C. Displayed p values represent Mann–Whitney t test results.

Figure 6

Figure 6

IgE cross-linking interferes with influenza-induced maturation of pDCs. A_–_C, Purified pDCs were cultured for 48 h with IL-3 alone (control), anti-IgE (5 μg/ml), rabbit IgG (5 μg/ml), or influenza A virus alone or after preincubation with anti-IgE or rabbit IgG. Cells were harvested, stained with CD86 PE-Cy5, CD80 FITC, and HLA-DR APC-Cy7 Abs, and the MFI of CD86 (A), CD80 (B), and HLA-DR (C) was measured by flow cytometry. Data are expressed as the mean ± SEM MFI values for displayed markers. Results of three individual experiments are displayed. Displayed p values represent results of paired t tests. Similar results for influenza B experiments are displayed in D_–_F. Data are expressed as the mean ± SEM.

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