Project description:House dust mite/HDM atopy patch test/APT elicits positive reactions in the majority of atopic dermatitis/AD and healthy individuals. Experimental systems for new-onset/chronic AD are needed to support rapid therapeutic development, particularly since animal models representing AD pathology in humans are lacking. HDM APT historically simulated AD, but its suitability to model the emerging AD skin phenotype as Th2/Th22 polarized with Th1 and Th17 components is unknown. To assess whether HDM APT tissues reproduce acute or chronic AD, positive HDM APT (n=14) were compared with nonlesional, acute (<72hrs; n=10), and chronic phase AD biopsies (n=8), allergic contact reactions (to nickel [n=10] and fragrance [n=3]) using arrays. Overall design: 14 patients with positive dust mite reactions, were compared to 8 AD patients (acute and chronic lesions). 8 patients out of 14 patients with positive dust mite reactions did not have positive reactions to nickel or fragrance. 5 patients out of 14 patients with positive dust mite reactions had positive reactions to nickel. 1 patient out of 14 patients with positive dust mite reactions had positive reaction to fragrance. 5 other patients with positive nickel reactions and 2 other patients with fragrance reaction but not dustmite reactions were included in the study. The study considered a total of 29 patients (8 Atopic Dermatitis and 21 dust mite/nickel/fragrance positive reactions). Lesional and Non Lesional skin for AD patients were analyzed. Atopic patch skin and non lesional skin for dust mite/nickel/fragrance patients were analyzed. For patients with more than one positive patch reaction a sample from each atopy patch test reaction was obtained. We are submiting here only the samples from dust mite atopic patch skin (14) and the non lesional skin samples related to patients with positive dust mite reaction only (8). The other samples have been submited in: series accession no. GSM815426, GSM815427, GSE36842 and GSE36842.
Project description:The PI3K-AKT pathway is known to regulate cytokines in dust mite-induced pediatric asthma. However, the underlying molecular steps involved are not clear. In order to clarify further the molecular steps, this study investigated the expression of certain genes and the involvement of miRNAs in the PI3K-AKT pathway, which might affect the resultant cytokine-secretion. In-vivo and in-vitro ELISA, qRT-PCR, western-blot and microarrays analyses were used in this study. A down-expression of miRNA-27b-3p in dust mite induced asthma group (group D) was found by microarray analysis. This was confirmed by qRT-PCR that found the miRNA-27b-3p transcripts that regulated the expression of SYK and EGFR were also significantly decreased (p < 0.01) in group D. The transcript levels of the SYK and PI3K genes were higher, while those of EGFR were lower in the former group. Meanwhile, we found significant differences in plasma concentrations of some cytokines between the dust mite-induced asthma subjects and the healthy controls. On the other hand, this correlated with the finding that the transcripts of SYK and its downstream PI3K were decreased in HBE transfected with miRNA-27b-3p, but were increased in HBE transfected with the inhibitor in vitro. Our results indicate that the differential expression of the miRNAs in dust mite-induced pediatric asthma may regulate their target gene SYK and may have an impact on the PI3K-AKT pathway associated with the production of cytokines. These findings should add new insight into the pathogenesis of pediatric asthma. Overall design: Twelve pairs of gender- and age-matched Group D dust mite-induced asthma children and Group N normal control children were included in the initial microarray-based discovery analysis.
Project description:Allergic asthmatic, allergy only, asthma only (no allergy), and non-allergic non-asthmatic (control) subjects underwent bronchoscopy with instillation of saline, lipopolysaccharide (LPS), and house dust mite antigen in separate subsegmental bronchi. Airway epithelial cells were collected four hours later (three samples per subject). RNA was extracted from these cells for microarray analysis. Keywords: gene expression arrays (two-dye: sample against common "universal" reference RNA) Overall design: There are four main phenotypic groups: 1. control (no allergy or asthma) 2. allergy only (no asthma) 3. asthma only (no allergy) 4. allergy and asthma and three exposures: saline, house dust mite antigen (HDM), and LPS. Samples from the different exposures were all collected at the same time: four hours after instillation. The hybridizations were carried out in two main "batches": samples in batch 1 were processed in mid 2004, samples in batch 2 about a year later in 2005. There is a clear "batch effect": differences between expression profiles from the two batches (likely caused by technical differences between hybridization and scanning methods). This should be considered when analyzing the data.
Project description:Response to allergen was studied in epithelial cells derived from allergic pantients and from healthy controls. Cells were cultured after isolation from a nasal biopsy. Cells were exposed to Housed dust mite or vessel (saline) Microarray data was analysed using bioinformatics and biostaistics. We conclude that a marked difference in basal expression and in response to hous dust mite exists Keywords: cellular response to allergen Overall design: We used 5 monotypic house dust mite allergic patients and 5 non-allergic healthy controls, the cells obtained from the biopsies were cultured in two wells, for two different conditions.