Transcriptomic analyses of cutaneous immune network in patiens with atopic dermatitis challenged with house dust mite (HDM) patch test

Our study investigates the activation and transcriptional programming of primary skin cells (FACS-purified Langerhans cells and single cell analysis of the whole biopsy) from patients with atopic dermatitis (n=22, nr = rensponding, nnr=nonresponding), exposed to a control or HDM patch test. 6 mm biopsies were taken from control and HDM patch test site 48h post application. Patients with moderate to severe atopic dermatitis eczema severity scores (EASI) median = 17.70, IQR:10.20 – 30.90, max = 51.40) under the care of a dermatologist in a tertiary referral centre were recruited to the study. 48h post application of a patch test to buttock skin, 11 out of the 28 patients were HDM-reactive, while In 12 patients, HDM did not induce a visible response, creating “HDM-non-reactive” group. 4 patients reacted to both HDM and control patch test "irritant", and one showed redness which was not patch test localised "undefined". 6 mm biopsies were taken from all patch test sites, and processed for flow cytometry or single cell RNA-seq analysis. CD207, CD1a, HLA-DR+ Langerhans cells (LCs)were FACS-sorted into RLT buffer and processed for bulk RNA-seq. Non-LCs cells were sorted and cryopreserved for further analysis with Constellation 10X. Freshly dissociated whole skin biopsies from 6 biopsies were suspended in RNAse-out buffer and processed on ice to the co-encapsulation of single cells with genetically-encoded beads (Drop-seq).