Background: During clinically effective allergen-specific immunotherapy a shift in cytokine dominance from IL-4, IL-5 predominant to IFN-γ predominant has been observed. As antigen concentration influences Th cell priming, this study aimed to determine the effect of different allergen concentrations on human house dust mite (HDM)-specific T cell production of IL-4 and IFN-γ, proliferation and apoptosis. Methods: HDM-allergic donor PBMC were cultured for 14 days with different concentrations of HDM extract (1, 10 and 100 µg/ml). T cell intracellular IL-4 and IFN-γ, division (CFSE labelling) and apoptosis (active caspase-3 staining) were analysed by flow cytometry. Proliferation was assessed by 3H-thymidine incorporation. Results: Increased CD4+IFN-γ+ and CD8+IFN-γ+ T cell numbers were observed in high allergen concentration cultures compared with low concentration cultures whereas there were no differences in CD4+IL-4+ and CD8+IL-4+ T cell numbers. CFSE cell labelling revealed that high allergen concentration favours the expansion of IFN-γ-producing CD4+ T cells. The proportion of apoptotic cells increased with allergen concentration and there was preferential apoptosis of CD4+IL-4+ T cells. HDM-induced proliferation was decreased in high allergen concentration cultures; this was reversible by IL-2 consistent with anergy. Conclusion: These results show that T cell division and apoptosis contribute to the cytokine skewing to predominant IFN-γ production by T cells observed at high allergen concentration. Thus the use of hypoallergenic T cell reactive preparations which can be given safely at higher doses than natural extracts may enhance efficacy of allergen-specific immunotherapy.

1.
Kay AB: Allergy and allergic diseases. First of two parts. N Engl J Med 2001;344:30–37.
2.
O’Hehir RE, Bal V, Quint D, Moqbel R, Kay AB, Zanders ED, Lamb JR: An in vitro model of allergen-dependent IgE synthesis by human B lymphocytes: comparison of the response of an atopic and a non-atopic individual to Dermatophagoides spp. (house dust mite). Immunology 1989;66:499–504.
3.
Wills-Karp M, Luyimbazi J, Xu X, Schofield B, Neben TY, Karp CL, Donaldson DD: Interleukin-13:central mediator of allergic asthma. Science 1998;282:2258–2261.
4.
Bousquet J, Lockey RF, Malling HJ: WHO Position Paper. Allergen immunotherapy: therapeutic vaccines for allergic disease. Allergy 1998;53(suppl):1–42.
5.
Akdis CA, Akdis M, Blesken T, Wymann D, Alkan SS, Muller U, Blaser K: Epitope-specific T cell tolerance to phospholipase A2 in bee venom immunotherapy and recovery by IL-2 and IL-15 in vitro. J Clin Invest 1996;98:1676–1683.
6.
Bellinghausen I, Metz G, Enk AH, Christmann S, Knop J, Saloga J: Insect venom immunotherapy induces interleukin-10 production and a Th2- to-Th1 shift, and changes surface marker expression in venom-allergic subjects. Eur J Immunol 1997;27:1131–1139.
7.
Ebner C, Siemann U, Bohle B, Willheim M, Wiedermann U, Schenk S, Klotz F, Ebner H, Kraft D, Scheiner O: Immunological changes during specific immunotherapy of grass pollen allergy: Reduced lymphoproliferative responses to allergen and shift from TH2 to TH1 in T-cell clones specific for Phl p 1, a major grass pollen allergen. Clin Exp Allergy 1997;27:1007–1015.
8.
Eusebius NP, Papalia L, Suphioglu C, McLellan SC, Varney M, Rolland JM, O’Hehir RE: Oligoclonal analysis of the atopic T cell response to the group 1 allergen of Cynodon dactylon (bermuda grass) pollen: pre- and post-allergen-specific immunotherapy. Int Arch Allergy Immunol 2002;127:234–244.
9.
Jutel M, Pichler WJ, Skrbic D, Urwyler A, Dahinden C, Muller UR: Bee venom immunotherapy results in decrease of IL-4 and IL-5 and increase of IFN-gamma secretion in specific allergen-stimulated T cell cultures. J Immunol 1995;154:4187–4194.
10.
Kammerer R, Chvatchko Y, Kettner A, Dufour N, Corradin G, Spertini F: Modulation of T-cell response to phospholipase A2 and phospholipase A2-derived peptides by conventional bee venom immunotherapy. J Allergy Clin Immunol 1997;100:96–103.
11.
McHugh SM, Deighton J, Stewart AG, Lachmann PJ, Ewan PW: Bee venom immunotherapy induces a shift in cytokine responses from a TH-2 to a TH-1 dominant pattern: Comparison of rush and conventional immunotherapy. Clin Exp Allergy 1995;25:828–838.
12.
O’Brien RM, Byron KA, Varigos GA, Thomas WR: House dust mite immunotherapy results in a decrease in Der p 2-specific IFN-gamma and IL-4 expression by circulating T lymphocytes. Clin Exp Allergy 1997;27:46–51.
13.
Rolland J, O’Hehir R: Immunotherapy of allergy: Anergy, deletion, and immune deviation. Curr Opin Immunol 1998;10:640–645.
14.
Constant S, Pfeiffer C, Woodard A, Pasqualini T, Bottomly K: Extent of T cell receptor ligation can determine the functional differentiation of naive CD4+ T cells. J Exp Med 1995;182:1591–1596.
15.
Grakoui A, Donermeyer DL, Kanagawa O, Murphy KM, Allen PM: TCR-independent pathways mediate the effects of antigen dose and altered peptide ligands on Th cell polarization. J Immunol 1999;162:1923–1930.
16.
Hosken NA, Shibuya K, Heath AW, Murphy KM, O’Garra A: The effect of antigen dose on CD4+ T helper cell phenotype development in a T cell receptor-alpha beta-transgenic model. J Exp Med 1995;182:1579–1584.
17.
Rogers PR, Grey HM, Croft M: Modulation of naive CD4 T cell activation with altered peptide ligands: The nature of the peptide and presentation in the context of costimulation are critical for a sustained response. J Immunol 1998;160:3698–3704.
18.
Rogers PR, Croft M: Peptide dose, affinity, and time of differentiation can contribute to the Th1/Th2 cytokine balance. J Immunol 1999;163:1205–1213.
19.
Gett AV, Hodgkin PD: Cell division regulates the T cell cytokine repertoire, revealing a mechanism underlying immune class regulation. Proc Natl Acad Sci USA 1998;95:9488–9493.
20.
Ben-Sasson SZ, Gerstel R, Hu-Li J, Paul WE: Cell division is not a ‘clock’ measuring acquisition of competence to produce IFN-gamma or IL-4. J Immunol 2001;166:112–120.
21.
Carballido JM, Faith A, Carballido-Perrig N, Blaser K: The intensity of T cell receptor engagement determines the cytokine pattern of human allergen-specific T helper cells. Eur J Immunol 1997;27:515–521.
22.
Secrist H, DeKruyff RH, Umetsu DT: Interleukin 4 production by CD4+ T cells from allergic individuals is modulated by antigen concentration and antigen-presenting cell type. J Exp Med 1995;181:1081–1089.
23.
Sutherland MF, Drew A, Rolland JM, Slater JE, Suphioglu C, O’Hehir RE: Specific monoclonal antibodies and human immunoglobulin E show that Hev b 5 is an abundant allergen in high protein powdered latex gloves. Clin Exp Allergy 2002;32:583–589.
24.
O’Hehir RE, Yssel H, Verma S, de Vries JE, Spits H, Lamb JR: Clonal analysis of differential lymphokine production in peptide and superantigen induced T cell anergy. Int Immunol 1991;3:819–826.
25.
Tovey ER, Chapman MD, Wells CW, Platts-Mills TA: The distribution of dust mite allergen in the houses of patients with asthma. Am Rev Respir Dis 1981;124:630–635.
26.
Bousquet J, Lockey R, Malling HJ: Allergen immunotherapy: Therapeutic vaccines for allergic diseases. A WHO position paper. J Allergy Clin Immunol 1998;102:558–562.
27.
Venkataraman C, Leung S, Salvekar A, Mano H, Schindler U: Repression of IL-4-induced gene expression by IFN-gamma requires Stat1 activation. J Immunol 1999;162:4053–4061.
28.
Huang TJ, MacAry PA, Eynott P, Moussavi A, Daniel KC, Askenase PW, Kemeny DM, Chung KF: Allergen-specific Th1 cells counteract efferent Th2 cell-dependent bronchial hyperresponsiveness and eosinophilic inflammation partly via IFN-gamma. J Immunol 2001;166:207–217.
29.
Durham SR, Ying S, Varney VA, Jacobson MR, Sudderick RM, Mackay IS, Kay AB, Hamid QA: Grass pollen immunotherapy inhibits allergen-induced infiltration of CD4+ T lymphocytes and eosinophils in the nasal mucosa and increases the number of cells expressing messenger RNA for interferon-gamma. J Allergy Clin Immunol 1996;97:1356–1365.
30.
Constant SL, Bottomly K: Induction of Th1 and Th2 CD4+ T cell responses: The alternative approaches. Annu Rev Immunol 1997;15:297–322.
31.
Kadowaki N, Ho S, Antonenko S, Malefyt RW, Kastelein RA, Bazan F, Liu YJ: Subsets of human dendritic cell precursors express different toll-like receptors and respond to different microbial antigens. J Exp Med 2001;194:863–869.
32.
Zarember KA, Godowski PJ: Tissue expression of human Toll-like receptors and differential regulation of Toll-like receptor mRNAs in leukocytes in response to microbes, their products, and cytokines. J Immunol 2002;168:554–561.
33.
Larche M: Inhibition of human T-cell responses by allergen peptides. Immunology 2001;104:377–382.
34.
Schwartz RH: T Cell Anergy. Annu Rev Immunol 2003;21:305–334.
35.
Groux H, Bigler M, de Vries JE, Roncarolo MG: Interleukin-10 induces a long-term antigen-specific anergic state in human CD4+ T cells. J Exp Med 1996;184:19–29.
36.
Akdis CA, Blesken T, Akdis M, Wuthrich B, Blaser K: Role of interleukin 10 in specific immunotherapy. J Clin Invest 1998;102:98–106.
37.
Vandenbark AA, Barnes D, Finn T, Bourdette DN, Whitham R, Robey I, Kaleeba J, Bebo BF, Jr., Miller SD, Offner H, Chou YK: Differential susceptibility of human T(h)1 versus T(h) 2 cells to induction of anergy and apoptosis by ECDI/antigen-coupled antigen-presenting cells. Int Immunol 2000;12:57–66.
38.
Janssen O, Stocker A, Sanzenbacher R, Oberg HH, Siddiqi MA, Kabelitz D: Differential regulation of activation-induced cell death in individual human T cell clones. Int Arch Allergy Immunol 2000;121:183–193.
39.
Guerra F, Carracedo J, Solana-Lara R, Sanchez-Guijo P, Ramirez R: TH2 lymphocytes from atopic patients treated with immunotherapy undergo rapid apoptosis after culture with specific allergens. J Allergy Clin Immunol 2001;107:647–653.
40.
Seneviratne SL, Jones L, King AS, Black A, Powell S, McMichael AJ, Ogg GS: Allergen-specific CD8(+) T cells and atopic disease. J Clin Invest 2002;110:1283–1291.
41.
Renz H, Lack G, Saloga J, Schwinzer R, Bradley K, Loader J, Kupfer A, Larsen GL, Gelfand EW: Inhibition of IgE production and normalization of airways responsiveness by sensitized CD8 T cells in a mouse model of allergen-induced sensitization. J Immunol 1994;152:351–360.
42.
Suzuki M, Maghni K, Molet S, Shimbara A, Hamid QA, Martin JG: IFN-gamma secretion by CD8T cells inhibits allergen-induced airway eosinophilia but not late airway responses. J Allergy Clin Immunol 2002;109:803–809.
43.
Auphan-Anezin N, Verdeil G, Schmitt-Verhulst AM: Distinct thresholds for CD8 T cell activation lead to functional heterogeneity: CD8 T cell priming can occur independently of cell division. J Immunol 2003;170:2442–2448.
44.
O’Brien RM, Xu H, Rolland JM, Byron KA, Thomas WR: Allergen-specific production of interferon-gamma by peripheral blood mononuclear cells and CD8 T cells in allergic disease and following immunotherapy. Clin Exp Allergy 2000;30:333–340.
45.
Majori M, Caminati A, Corradi M, Brianti E, Scarpa S, Pesci A: T-cell cytokine pattern at three time points during specific immunotherapy for mite-sensitive asthma. Clin Exp Allergy 2000;30:341–347.
46.
Levings MK, Bacchetta R, Schulz U, Roncarolo MG: The role of IL-10 and TGF-beta in the differentiation and effector function of T regulatory cells. Int Arch Allergy Immunol 2002;129:263–276.
47.
Muller U, Akdis CA, Fricker M, Akdis M, Blesken T, Bettens F, Blaser K: Successful immunotherapy with T-cell epitope peptides of bee venom phospholipase A2 induces specific T-cell anergy in patients allergic to bee venom. J Allergy Clin Immunol 1998;101:747–754.
48.
Ferreira F, Hirtenlehner K, Jilek A, Godnik-Cvar J, Breiteneder H, Grimm R, Hoffmann-Sommergruber K, Scheiner O, Kraft D, Breitenbach M, Rheinberger HJ, Ebner C: Dissection of immunoglobulin E and T lymphocyte reactivity of isoforms of the major birch pollen allergen Bet v 1:potential use of hypoallergenic isoforms for immunotherapy. J Exp Med 1996;183:599–609.
49.
Smith AM, Chapman MD: Reduction in IgE binding to allergen variants generated by site-directed mutagenesis: contribution of disulfide bonds to the antigenic structure of the major house dust mite allergen Der p 2. Mol Immunol 1996;33:399–405.
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