Spatial transcriptomics reveals differential inflammatory pathways in discoid lupus erythematosus and lichen planus

Here, we performed spatial transcriptomics using GeoMx® Digital Spatial Profiling, a highly multiplex method for detection of proteins and RNA in fixed tissue (Merritt et al., 2020) to broadly profile immune response pathways in keratinocytes and immune cells between patients with DLE and LP. Specifically, we used the Cancer Transcriptome Atlas (1868 genes) on archived biopsies from patients with DLE (n=9) and LP (n=10) and control skin (n=4). First, tissues were initially stained with morphological markers CD45 and pan-cytokeratin (Pan-CK) to identify regions of interest (ROIs) for subsequent analysis. Cell-masking was used to select mRNA in either keratinocytes (Pan-CK+) or CD45+ immune cells, but no other cells that may have been present in the geometric ROI. A total of 311 segments (CD45+ cells or PanCK+ cells) from 153 ROIs across 23 unique patient samples for a total of 95,823 cells were included in this analysis.本研究采用GeoMx®数字空间分析（GeoMx® Digital Spatial Profiling）技术开展空间转录组学（spatial transcriptomics）分析——该技术是一种可在固定组织中同时检测蛋白与RNA的高多重检测方法（Merritt等，2020），旨在全面解析盘状红斑狼疮（DLE）与扁平苔藓（LP）患者的角质形成细胞（keratinocytes）及免疫细胞内的免疫应答通路。具体而言，本研究针对存档的活检组织样本（含9例DLE患者、10例LP患者及4例皮肤对照样本），采用包含1868个基因的癌症转录组图谱（Cancer Transcriptome Atlas）开展检测。首先，研究人员借助形态学标志物CD45与广谱细胞角蛋白（pan-cytokeratin, Pan-CK）对组织进行染色，以明确后续分析所需的感兴趣区域（regions of interest, ROIs）。随后通过细胞掩膜技术（cell-masking），仅选取几何感兴趣区域内角质形成细胞（Pan-CK+）或CD45+免疫细胞中的mRNA，排除区域内其他可能存在的细胞类型。本次分析共纳入来自23例独立患者样本的153个ROIs中的311个细胞片段（CD45+细胞或PanCK+细胞），总计95823个细胞。