Lack of allergy to timothy grass pollen is not a passive phenomenon but associated with the allergen-specific modulation of immune reactivity - PubMed (original) (raw)

G Seumois 2, A M Gholami 1, J A Greenbaum 3, J Lane 1, B White 1, D H Broide 4, V Schulten 1, J Sidney 1, P Bakhru 1, C Oseroff 1, E Wambre 5, E A James 5, W W Kwok 5, B Peters 1, P Vijayanand 1, A Sette 1

Affiliations

Lack of allergy to timothy grass pollen is not a passive phenomenon but associated with the allergen-specific modulation of immune reactivity

D Hinz et al. Clin Exp Allergy. 2016 May.

Abstract

Background: Timothy grass (TG) pollen is a common seasonal airborne allergen associated with symptoms ranging from mild rhinitis to severe asthma.

Objective: The aim of this study was to characterize changes in TG-specific T cell responses as a function of seasonality.

Methods: Peripheral blood mononuclear cells (PBMCs) obtained from allergic individuals and non-allergic controls, either during the pollen season or out of season, were stimulated with either TG extract or a pool of previously identified immunodominant antigenic regions.

Results: PBMCs from allergic subjects exhibit higher IL-5 and IL-10 responses in season than when collected out of season. In the case of non-allergic subjects, as expected we observed lower IL-5 responses and robust production of IFN-γ compared to allergic individuals. Strikingly, non-allergic donors exhibited an opposing pattern, with decreased immune reactivity in season. The broad down-regulation in non-allergic donors indicates that healthy individuals are not oblivious to allergen exposure, but rather react with an active modulation of responses following the antigenic stimulus provided during the pollen season. Transcriptomic analysis of allergen-specific T cells defined genes modulated in concomitance with the allergen exposure and inhibition of responses in non-allergic donors.

Conclusion and clinical relevance: Magnitude and functionality of T helper cell responses differ substantially in season vs. out of season in allergic and non-allergic subjects. The results indicate the specific and opposing modulation of immune responses following the antigenic stimulation during the pollen season. This seasonal modulation reflects the enactment of specific molecular programmes associated with health and allergic disease.

Keywords: T cell response; non-allergic; seasonality; timothy grass.

© 2015 John Wiley & Sons Ltd.

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Conflict of interest statement

CONFLICT OF INTEREST

The authors declare no conflict of interest.

Figures

Figure 1

Figure 1. Kinetics and magnitude of cytokine production in TG-allergic and non-allergic donors

A–B: Cytokine production over 21 days following restimulation with TG extract (A) or a pool of 20 predominant TG epitopes (B). Means of 10 donors are shown, respectively. Dashed line, allergics; continuous line, non-allergics. Error bars indicate SD. C–D: Seasonal cytokine responses in a cross-sectional cohort of TG allergic (C) and non-allergic (D) individuals. IFNγ, IL-5, IL-10 and IL-17 reponses elicited by P20 in a 14-day in vitro restimulation assay. Samples obtained between Jul 1st to Oct 15th are considered in-season. *P < 0.05, ** P < 0.01, Mann-Whitney U-test.

Figure 2

Figure 2. Seasonal modulation of cytokine reactivity is allergen-specific

Ratio of in- and out-of-season cytokine reactivity in TG-allergic donors tested against (A) a set of other common seasonal (trees: date palm, olive, black walnut, prickly juniper; grasses: rye grass, bermuda grass, canary grass, wheat, kentucky blue; weeds: giant ragweed, mugwort) and (B) perennial allergens (american cockroach, german cockroach, house dust mite (Der p 1), cat, dog, Alternaria sp.). All donors tested were TG-allergic and were further classified as allergic or non-allergic for other allergens on the basis of skin test reactivity (All, allergic; Ctr, not allergic to the tested seasonal or perennial allergen). PBMC, obtained in and out-of-season, were stimulated with allergen extracts for 14 days and IFNγ and IL-5 production was analyzed longitudinally (in the same donor in- and out-of-season). Donors were chosen for representative assays based on availability of longitudinal samples and type of assay. P values were determined using Mann-Whitney U-test.

Figure 3

Figure 3. Seasonal cytokine response in longitudinal samples from TG-allergic and non-allergic individuals

Cytokine responses (IFNγ, IL-5) elicited by P20 in a 14-day in vitro restimulation assay in allergic (A) and non-allergic donors (B). *P < 0.05, paired Wilcoxon one-tailed test.

Figure 4

Figure 4. A Ex vivo tetramer analysis

Numbers of TG-specific CD4+ T cells in- and out-of-season in allergic and non-allergic donors. B–C. Co-culture of longitudinal samples. PBMC from in and out-of-season donations were stimulated in vitro with P20 separately as described before. Immune reactivity in 1:1 co-cultures of in and out-of-season donations from the respective allergic (B) or non-allergic (C) donors were analyzed. The expected reactivity in absence of any inhibitory mechanisms, calculated as the average of the in and out-of-season reactivity (in terms of SFC per million PBMC) is plotted and compared to the actual observed reactivity by the end of the in vitro co-culture. *P < 0.05, as calculated by paired Wilcoxon one-tailed test.

Figure 5

Figure 5. Gene expression patterns of IFNγ- and IL-5-producing antigen-specific cells

A. MA plots comparing gene expression patterns of IFNγ- or IL-5-producing cells in allergic and non-allergic donors as a function of pollen season. Geometric mean of gene expression (x-axis) is compared with fold change in expression between in- and out-of-season samples. Differentially regulated genes were identified based on an adjusted _P_-value of < 0.05 and as well as a two-fold change in gene expression (red circles). B. Histogram of the number of genes differentially expressed between in- and out-of-season samples categorized by expression direction (black, upregulation; grey, downregulation) and by allergic status and type of molecule captured. C. Heat map gene expression of IL-5-producing cells from TCM (green) and TEM (blue) in non-allergic donors in- and out-of-season (green and red, respectively). Data from 6 non-allergic individuals (3 cross-sectional and 3 longitudinal samples) is shown. Each row indicates gene expression levels of each individual donor represented in the respective column. Shades of red are reflective of the level of downregulation; blue, upregulation; white, no change in expression with season. The heat map was generated by taking into account the distance for similarity between genes and samples using the Euclidean distance. Genes and samples were clustered using the Ward linkage method. Micro-scaled RNA-seq was performed on a total of 76 samples: 64 samples (11 donors) for each allergen-specific CD4+ subset (IFNγ-TEM, IFNγ-TCM IL-5-TEM, IL-5-TCM) and 12 samples (5 donors) for each CD4+ memory subset not producing IFNγ or IL-5 (TEM, TCM).

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