Marietta, Eric

Abstract Airway eosinophilic inflammation is a characteristic feature of allergic asthma. Exposure to allergens produced by the German cockroach (Blattella germanica) is a risk factor for allergic disease in genetically predisposed individuals, and has been linked to an increase in asthma morbidity among cockroach-sensitive inner city children. To determine the role and contribution of specific HLA class II in the pathogenesis of allergic airway inflammation in cockroach-induced asthma, we generated double-transgenic, double-knockout mice expressing human HLA-DQ8, HLA-DQ6, and CD4 molecules in the absence of mouse class II and mouse CD4. Mice were actively immunized and later challenged intranasally with cockroach allergen extract. These mice developed bronchoalveolar lavage fluid (BALF) eosinophilia and pulmonary eosinophilia. This was accompanied by an increase in total protein levels, IL-5, and IL-13 in BALF. There were also elevated levels of cockroach-specific serum IgG1 and total serum IgE. Histological analysis revealed peribronchial and perivascular eosinophilic inflammation in cockroach-treated mice. Other pathologic changes in the airways were epithelial cell hypertrophy and mucus production. Treatment with anti-DQ mAb significantly reduced pulmonary and BALF eosinophilia in cockroach allergen-sensitized mice. Aβ0 mice and transgenic mice expressing human CD4 molecule alone (without class II) or human HLA-DQ8 molecule (without CD4) treated in the same fashion showed no eosinophilia in bronchoalveolar fluid and no pulmonary parenchymal inflammation. Our results provide direct evidence that HLA-DQ molecules and CD4 T cells mediate cockroach-induced eosinophilic inflammation in the airways.

Abstract To investigate the role of HLA-DQ molecules and/or CD4+ T cells in the pathogenesis of allergic asthma, we generated HLA-DQ6 and HLA-DQ8 transgenic mice lacking endogenous class II (Aβnull) and CD4 genes and challenged them intranasally with short ragweed allergenic extract (SRW). We found that DQ6/CD4null mice developed a strong eosinophilic infiltration into the bronchoalveolar lavage and lung tissue, while DQ8/CD4null mice were normal. However, neither cytokines nor eosinophil peroxidase in the bronchoalveolar lavage of DQ6/CD4null mice was found. In addition, the airway reactivity to methacholine was elevated moderately in DQ6/CD4null mice compared with the high response in DQ/CD4+ counterparts and was only partially augmented by CD4+ T cell transfer. The DQ6/CD4null mice showed Th1/Th2-type cytokines and SRW-specific Abs in the immune sera in contrast to a direct Th2 response observed in DQ6/CD4+ mice. The proliferative response of spleen mononuclear cells and peribronchial lymph node cells demonstrated that the response to SRW in DQ6/CD4null mice was mediated by HLA-DQ-restricted CD4−CD8−NK1.1− T cells. FACS analysis of PBMC and spleen mononuclear cells demonstrated an expansion of double-negative (DN) CD4−CD8−TCRαβ+ T cells in SRW-treated DQ6/CD4null mice. These cells produced IL-4, IL-5, IL-13, and IFN-γ when stimulated with immobilized anti-CD3. IL-5 ELISPOT assay revealed that DN T cells were the cellular origin of IL-5 in allergen-challenged DQ6/CD4null mice. Our study shows a role for HLA-DQ-restricted CD4+ and DN T cells in the allergic response.

Abstract Active delivery of recombinant autoantigens or allergens at the intestinal mucosa by genetically modified Lactococcus lactis (LL) provides a novel therapeutic approach for the induction of tolerance. Celiac disease is associated with either HLA-DQ2- or HLA-DQ8-restricted responses to specific antigenic epitopes of gliadin, and may be treated by induction of Ag-specific tolerance. We investigated whether oral administration of LL-delivered DQ8-specific gliadin epitope induces Ag-specific tolerance. LL was engineered to secrete a deamidated DQ8 gliadin epitope (LL-eDQ8d) and the induction of Ag-specific tolerance was studied in NOD AB° DQ8 transgenic mice. Tolerance was assessed by delayed-type hypersensitivity reaction, cytokine measurements, eDQ8d-specific proliferation, and regulatory T cell analysis. Oral administration of LL-eDQ8d induced suppression of local and systemic DQ8-restricted T cell responses in NOD AB° DQ8 transgenic mice. Treatment resulted in an Ag-specific decrease of the proliferative capacity of inguinal lymph node (ILN) cells and lamina propria cells. Production of IL-10 and TGF-β and a significant induction of Foxp3+ regulatory T cells were associated with the eDQ8d-specific suppression induced by LL-eDQ8d. These data provide support for the development of effective therapeutic approaches for gluten-sensitive disorders using orally administered Ag-secreting LL. Such treatments may be effective even in the setting of established hypersensitivity.