House dust mite (HDM) is the most important inducer of allergy and asthma in the world. More than thirty three group of mite allergens have been reported from HDMs and storage mites (SMs). The accurate diagnosis of mite allergy is a very important for effective treatment. For diagnosis and immunotherapy of mite allergy, the crude extract of mites have been used, which may cause side effects. Recently, specific allergens selectively developed using molecular biological technology came into the spotlight. This study intended to examine diagnostic and immunotherapeutic potentials of the recombinant allergens from house dust mite Dermatophagoides farinae.
Thirteen D. farinae-derived recombinant allergens were produced using Escherichia coli or Pichia pastoris expression system. Native Der f 4 was also purified for comparison. rDer f 1 (89.2%) and rDer f 2 (96.4%) showed high reactivity to IgE in Korean HDM-sensitized patient’s sera. rDer f 3 (61.9%), nDer f 4 (75.8%), rDer f 5 (64.9%), rDer f 7 (43.8%), rDer f 21 (66.0%), and rDer f 23 (43.3%) exhibited moderate IgE reactivity them. IgE reactivity of rDer f 10 (24.7%), rDer f 13 (3.6%), rDer f 15 (4.1%), rDer f 18 (32.0%) and rDer f 20 (38.7%) displayed low IgE reactivity. IgE reactivities against Der f 2 were shown in all patients with asthma, rhinitis or atopic dermatitis. Each serum recognized with 2 to 12 D. farinae proteins, and 190 of 194 (97.9%) recognized with three or more proteins.
A cross-reactivity of recombinant group 5 and 21 allergens from three mite species (D. farinae, Tyrophagus putrescentiae, and Blomia tropicalis) was evaluated. Cross-reactivity between rDer f 23 and rDer p 23 were also examined by using inhibition-ELISA. IgE reactivity to rDer f 5 was highly correlated with dose of rDer f 21 (r = 0.733). rTyr p 5 exhibited the highest level of correlation with rTyr p 21 (r = 0.950). rDer f 21 exhibited a high inhibitory effect against rTyr p 5 (93.0%), rTyr p 21 (92.1%), rBlo t 5 (87.0%), and rBlo t 21 (70.3%), implying cross-reactivity across mite species. rDer p 23 effectively inhibited a IgE binding to rDer f 23.
Not only ability of IgE responses but also capability to induce inflammatory cytokine production is important properties of allergens. In this study, IL-6 and IL-8 production in BEAS-2B cells (bronchial epithelial cell-line) were examined by challenging of the recombinant mite allergens. rDer f 4, rDer f 5, rDer f 10 and rDer f 21 induced IL-6 or IL-8 secretions in BEAS-2B. In particular, rDer f 21 could trigger IL-6 and IL-8 production in BEAS-2B through TLR4-dependent signaling.
In summary, Der f 1 and Der f 2 was shown to be the most potent allergens. Der f 3, Der f 4, Der 5, Der f 7, Der f 21 and Der f 23 were found to be the major allergens sensitized more than 50% of HDM allergy patients. Der f 21 showed cross-reactivity to T. putrescentiae and B. tropicalis group 5 and 21 allergens. In addition, Der f 21 was found to trigger of IL-6 and IL-8 production in a bronchial epithelial cell line through a TLR4.
The information on allergenicity and IgE reactivities of various mite allergens shown in this study will be useful for development of a component-resolved diagnosis and effective treatment to HDM allergy.
