Spatial Transcriptomic Analysis of Early and Late Stages of Hidradenitis Suppurativa
Background: Hidradenitis suppurativa (HS) is a chronic inflammatory skin disease characterized by recurrent inflammatory nodules, abscess formation, and sinus tract development. Current biologic therapies provide limited clinical benefit, underscoring the need for deeper understanding of the immune mechanisms driving HS pathogenesis. Objective: We sought to comprehensively characterize the immune landscape of early- and late-stage HS. Methods: Skin biopsies from a total of 15 patients with early and late-stage HS were analyzed using spatial transcriptomics, with 6 samples from these patients further analyzed using single-cell RNA sequencing (scRNA-seq). Gene expression profiles were compared to the epidermal epithelium of acne conglobata in spatial transcriptomics and to psoriasis samples in scRNA-seq as controls. Results: Spatial transcriptomics revealed extensive dermal immune activation from the early stage of HS, including a prominent B-cell signature (immunoglobulins, CD19, MS4A1), neutrophilic infiltration (CD177, MPO, ELANE), and robust IL-23/IL-17 axis, with enriched expression of T17-inducing transcripts (TGFB3, RUNX1, BATF) and T17 cytokines (IL17A, IL17F, IL21). ScRNA-seq confirmed marked plasma cell and B-cell expansion in HS and identified potentially multifunctional B-cells expressing pro-inflammatory cytokines (IL7, IL16) in addition to their role in antigen presentation and antibody production. Furthermore, T-cells were identified as the primary producers of the B-cell chemoattractant CXCL13. Conclusions: HS exhibits early and sustained immune activation characterized by B-cell enrichment, IL-23/IL-17 axis activation, and neutrophilic inflammation extending into the deep dermis. These findings highlight the aggressive nature of HS from its onset, suggesting that early therapeutic targeting of the IL-23/IL-17 axis and B-cell-mediated inflammation may prevent disease progression. Overall design: FFPE tissue sections (5 µm) were prepared on BOND Plus slides, followed by deparaffinization, rehydration, and antigen retrieval using Tris-EDTA and proteinase K treatment. The tissues were hybridized overnight with GeoMx WTA RNA probes (18,677 genes) and stained with morphology markers (panCK, CD45, CD31, and SYTO 13). ROIs were selected and categorized into four regions: epithelium epidermis (above dermis), epithelium dermis (sinus tracts), dermis superficial inflammation (immune cells near epidermis), and dermis deep inflammation (immune cells near epithelium dermis). Using the DSP instrument, ROIs were segmented into AOIs, and indexed oligonucleotides were UV-released and collected. Libraries were constructed following the NanoString GeoMx-NGS protocol, using Seq Code primers and AMPure XP bead purification. Sequencing was performed on Illumina NovaSeq 6000 (2 × 151 bp paired-end).