Single-cell Transcriptomics Combined With Interstitial Fluid Proteomics Defines Cell Type-Specific Immune Regulation in Atopic Dermatitis

Background: Atopic dermatitis (AD) is the most common chronic inflammatory skin disease, but its complex pathogenesis is only insufficiently understood, resulting in still limited treatment options. Objective: We sought to characterize AD on both transcriptomic and proteomic levels in humans. Methods: We used skin suction blistering, a painless and nonscarring procedure that can simultaneously sample skin cells and interstitial fluid. We then compared results with conventional biopsies. Results: Suction blistering captured epidermal and most immune cells equally well as biopsies, except for mast cells and nonmigratory CD163+ macrophages that were only present in biopsy isolates. Using single-cell RNA sequencing, we found comparable transcriptional profiles of key inflammatory pathways between blister and biopsy AD, but suction blistering was superior in cell-specific resolution for high-abundance transcripts (KRT1/KRT10, KRT16/KRT6A, S100A8/S100A9), which showed some background signals in biopsy isolates. Compared with healthy controls, we found characteristic upregulation of AD-typical cytokines such as IL13 and IL22 in Th2 and Th22 cells, respectively, but we also discovered these mediators in proliferating T cells and natural killer T cells, that also expressed the antimicrobial cytokine IL26. Overall, not T cells, but myeloid cells were most strongly enriched in AD, and we found dendritic cell (CLEC7A, amphiregulin/AREG, EREG) and macrophage products (CCL13) among the top upregulated proteins in AD blister fluid proteomic analyses. Conclusion: These data show that by using cutting-edge technology, suction blistering offers several advantages over conventional biopsies, including better transcriptomic resolution of skin cells, combined with proteomic information from interstitial fluid, unraveling novel inflammatory players that shape the cellular and proteomic microenvironment of AD. Comparison of skin cells obtained by skin suction blistering and conventional biopsy >>>Raw data are unvailable due to patient privacy concerns<<<

背景：特应性皮炎（Atopic dermatitis, AD）是最常见的慢性炎症性皮肤病，但其复杂的发病机制尚未被充分阐明，导致临床可用治疗手段依然有限。
目的：本研究旨在从转录组学与蛋白质组学水平对人类特应性皮炎进行特征解析。
方法：我们采用皮肤负压吸疱术（skin suction blistering）——一种无痛且无瘢痕的操作，可同时采集皮肤细胞与组织间液——并将所得结果与常规活检组织进行对比。
结果：与常规活检相比，负压吸疱术可同等有效地采集表皮细胞与多数免疫细胞，但肥大细胞及仅存在于活检分离样本中的非迁移性CD163+巨噬细胞除外。通过单细胞RNA测序（single-cell RNA sequencing），我们发现吸疱样本与活检样本中特应性皮炎的关键炎症通路转录谱具有可比性，但在高丰度转录本（KRT1/KRT10、KRT16/KRT6A、S100A8/S100A9）的细胞特异性解析度上，负压吸疱术更具优势——这类转录本在活检分离样本中会出现一定程度的背景信号。与健康对照组相比，我们在Th2细胞与Th22细胞中分别观察到特应性皮炎典型细胞因子IL13与IL22的特征性上调表达；同时还在增殖性T细胞及自然杀伤T细胞中发现了这类介质，此类细胞同时也表达抗菌细胞因子IL26。总体而言，特应性皮炎中富集程度最高的并非T细胞，而是髓系细胞；在特应性皮炎吸疱液的蛋白质组学分析中，我们发现树突状细胞相关产物（CLEC7A、双调蛋白（amphiregulin, AREG）、上皮调节蛋白（EREG））以及巨噬细胞产物CCL13位列上调蛋白前列。
结论：本研究数据表明，借助前沿技术，皮肤负压吸疱术相较常规活检具有多项优势，包括可实现皮肤细胞更高分辨率的转录组学解析，同时能获取组织间液的蛋白质组学信息，从而揭示了调控特应性皮炎细胞与蛋白质组微环境的新型炎症介质。
皮肤负压吸疱术与常规活检获取的皮肤细胞对比 >>>因患者隐私保护需求，原始数据暂未公开<<<