P229 Integrated analysis of dermal blister fluid proteomics and skin biopsy transcriptomics gives new insight into pathogenesis of systemic sclerosis
P229 Integrated analysis of dermal blister fluid proteomics and skin biopsy transcriptomics gives new insight into pathogenesis of systemic sclerosis
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P229 真皮水疱液蛋白质组学和皮肤活检转录组学的综合分析为系统性硬化症的发病机制提供了新的见解
DOI：
10.1093/rheumatology/keac133.228
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发表时间：
2022
期刊：
影响因子：
5.5
通讯作者：
Clark K
中科院分区：
文献类型：
--
作者：
Clark K
背景/目的吸疱液(BF)为分析SSC的真皮微环境提供了独特的机会。我们使用一种综合的方法来分析来自真皮间质液的蛋白质组数据和来自早期DcSSc和健康对照(HC)的皮肤活检的全基因组转录数据。在RNA测序(RNAseq)的背景下解释差异蛋白质表达为了解与SSc相关的潜在介质和途径提供了独特的见解。方法活检队列收集了21例早期dcSSc患者，平均皮肤评分(MRSS)为21(SD 11)和16 HC。采用真皮吸疱法获得前臂皮肤BF，在Olink平台(www.olink.com)上测定1192种蛋白质。同时对所有受试者进行4 mm穿孔活检，通过RNAseq进行全基因组转录图谱分析。使用WGCNA和R中的ClusterProfiler对临床特征、基因和蛋白质表达数据进行综合分析。该方法将单个分析物聚类为具有相似表达模式的模块。结果WGCNA从高炉中识别出6个模块，从皮肤中识别出30个模块。两个BF模块和皮肤中的三个模块相互之间呈正相关，并且它们都与早期dcSSc的诊断相关。皮肤的品红模块(385个基因)和蓝色模块(450个蛋白质)的相关性最强(分别为r=0.74p<；0.001和r=0.6p<；0.001)。通过询问每个模块，确定了与当前SSC发病机制概念一致的关键生物学过程，并且皮肤和水泡模块似乎在很大程度上是相辅相成的。洋红色模块的过程集中于血管生成、细胞外基质结构和内皮细胞增殖，与HC相比，COMP(FC=5.06p<；0.001)、COL4A4(FC=3.86p<；0.001)、COL11A1(FC=3.46p<；0.001)的倍数变化最大。来自皮肤转录的红色模块由与免疫激活相关的基因组成，而干扰素信号主要出现在粉色模块的基因中。在BF蛋白质组学中，蓝色模块由围绕细胞外基质组织、细胞黏附和对肿瘤坏死因子的反应的蛋白质组成，在dcSSc早期上调了SFRP1(FC=4.88，p=0.013)、TnC(FC=4.81，p=0.0018)和KLK4(FC=7.17，p<；0.001)。而代谢过程是绿松石模块的中心。在每种组织形态中，比较所有五个模块之间的重要Hallmark通路。值得注意的是，一些途径在皮肤基因表达和水疱液模块中都上调，包括NFKB信号、同种异体移植排斥反应和干扰素伽马反应。在皮肤基因中显著激活的途径包括G2M检查点、P53途径和异体代谢，而不是水泡蛋白模块。相反，只有水疱区的蛋白质分泌、未折叠蛋白反应和有丝分裂纺锤体通路显著增加。结论同时对皮肤基因和蛋白质表达的高维分析为深入了解SSC的分子病理机制提供了新的视角。这为开发一种新型的皮肤复合生物标志物提供了一个潜在的平台。发现：K.E.N.Clark：无。E.E.CSomor：公司任命；GSK.C.Campochiaro的员工：无。Y.Teo：公司任命；GSK.J.Freudenberg的员工：公司任命；GSK.K.内文的员工：公司任命；的员工
Background/AimsSuction blister fluid (BF) provides a unique opportunity to analyse the dermal microenvironment of SSc. We utilised an integrated approach to analyse proteomic data from dermal interstitial fluid and genome-wide transcriptomic data from skin biopsies from early dcSSc and healthy controls (HC). Interpreting differential protein expression in the context of RNA sequencing (RNAseq) provides unique insight into potential mediators and pathways relevant to SSc.MethodsThe BIOPSY cohort recruited 21 early dcSSc patients with mean skin score (MRSS) of 21 (sd 11) and 16 HCs. Forearm skin BF was obtained using the dermal suction blister method and assayed using the Olink platform (www.olink.com) (1192 proteins). Simultaneous 4mm punch biopsies were taken from all subjects, for genome-wide transcriptomic profiling by RNAseq. Integrated analysis of clinical characteristics, gene and protein expression data was conducted using WGCNA and clusterProfiler in R. This method clusters individual analytes into modules sharing similar expression patterns. Each module has been arbitrarily assigned as a colour.ResultsWGCNA identified 6 modules in BF, and 30 modules from skin. Two BF modules and three in the skin positively correlated with each other, and all of them correlated with a diagnosis of early dcSSc. The magenta module in the skin (385 genes) and blue module (450 proteins) in BF had the strongest correlation (r = 0.74, p < 0.001 and r = 0.6, p < 0.001 respectively). Interrogating each module, key biological processes consistent with current concepts of SSc pathogenesis were identified, and skin and blister modules largely seemed to complement each other. Magenta module processes focused on angiogenesis, extracellular matrix (ECM) structure, and endothelial cell proliferation, with the highest fold change in COMP (FC = 5.06, p<0.001), COL4A4 (FC = 3.86,p<0.001), COL11A1 (FC = 3.46, p<0.001) having the highest fold change compared to HC. The red module from the skin transcriptomics consisted of genes relating to immune activation, whereas the interferon signature was predominantly seen in the genes of the pink module. Within BF proteomics, the blue module consisted of proteins centred around ECM organisation, cell adhesion, and response to TNF, with upregulation of SFRP1 (FC = 4.88, p = 0.013), TNC(FC = 4.81, p=0.0018) and KLK4 (FC = 7.17, p<0.001) in early dcSSc. Whereas metabolic processes were the centre of the turquoise module. Significant Hallmark pathways were compared between all five modules in each tissue modality. Notably, some pathways were upregulated in both skin gene expression and blister fluid modules including NFKB signalling, allograft rejection, and interferon gamma response. Significantly activated pathways in skin gene but not the blister protein modules include G2M checkpoint, p53 pathway, and xenobiotic metabolism. Conversely, protein secretion, unfolded protein response and mitotic spindle pathways were significantly increased only in the blister modules.ConclusionSimultaneous high dimensional analysis of skin gene and protein expression gives new insight into the molecular pathology of SSc. This provides a potential platform for developing a novel composite biomarker for skin.DisclosureK.E.N. Clark:None.E. Csomor:Corporate appointments; employee of GSK.C. Campochiaro:None.Y. Teo:Corporate appointments; employee of GSK.J. Freudenberg:Corporate appointments; employee of GSK.K. Nevin:Corporate appointments; employee of …