小儿急性肝衰竭患者肝组织的转录分析
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Background and Aims: Many patients with indeterminate pediatric acute liver failure (PALF) have evidence of T-cell driven immune injury, however the precise inflammatory pathways have not been well defined. We have thus characterized the hepatic cytokine and transcriptional signatures of PALF patients. Approach and Results: Retrospective review was performed on 22 children presenting with indeterminate (IND-PALF; n=17) or other diagnoses (DX-PALF; n=6) with available archived liver tissue. Specimens were stained for CD8 T-cells and scored as dense, moderate, or minimal. Measurement of immune analytes and RNA-sequencing (RNA-seq) were performed on whole liver tissue. Immune analyte data was analyzed by principal component analysis, and RNA-seq was analyzed by unsupervised hierarchical clustering, differential gene expression, and gene set enrichment analysis. Most IND-PALF patients (94%) had dense/moderate CD8 staining and were characterized by Th1 immune analytes including TNFα, IFNγ, IL-1β, IL-12, CXCL9, and CXCL12. Transcriptional analyses identified 2 transcriptional PALF phenotypes. Most patients in Group 1 (91%) had IND-PALF and dense/moderate CD8 staining. This group was characterized by increased expression of genes and cell-subset specific signatures related to innate inflammation, T-cell activation, and antigen stimulation. Group 1 also expressed significantly higher levels of gene signatures for T-regulatory cells, macrophages, Th1 cells, T effector memory cells, cytotoxic T-cells, and activated dendritic cells (p-adj <0.05). In contrast, patients in Group 2 exhibited increased expression for genes involved in metabolic processes. Conclusions: IND-PALF patients have evidence of a Th1 mediated inflammatory response driven by IFNγ. Transcriptional analyses suggest a complex immune network may regulate an immune-driven PALF phenotype with less evidence of metabolic processes. These findings provide novel insight into mechanisms of hepatic injury in PALF, areas for future research, and potential therapeutic targets.
研究背景与目的：多数不明原因儿童急性肝衰竭（indeterminate pediatric acute liver failure, PALF）患者存在T细胞介导的免疫损伤证据，但确切的炎症通路尚未明确。本研究旨在解析PALF患者的肝脏细胞因子与转录特征。 研究方法与结果：本研究对22例符合入组标准的儿童开展回顾性分析，其中17例为不明原因PALF（IND-PALF）、6例为其他病因PALF（DX-PALF），所有入组者均留存有可用的肝脏组织存档标本。对标本进行CD8 T细胞染色，并依据染色密度分为致密、中度及微弱三个评分等级。对全肝组织开展免疫分析物检测及RNA测序（RNA-seq）：采用主成分分析对免疫分析物数据进行统计分析，针对RNA-seq数据则通过无监督层次聚类、差异基因表达分析及基因集富集分析进行解析。 94%的IND-PALF患者呈现致密/中度CD8染色，且其免疫表型以Th1型细胞因子为主，包括TNFα、IFNγ、IL-1β、IL-12、CXCL9及CXCL12。转录组分析共鉴定出两类PALF相关转录表型：第1组中91%的患者为IND-PALF，且存在致密/中度CD8染色；该组患者的基因表达及细胞亚群特征与先天炎症、T细胞活化及抗原刺激相关通路显著上调有关，同时其调节性T细胞、巨噬细胞、Th1细胞、效应记忆T细胞、细胞毒性T细胞及活化树突状细胞的基因特征表达水平均显著升高（校正后P值<0.05）。与之相反，第2组患者的基因表达则以代谢相关通路上调为主要特征。 研究结论：IND-PALF患者存在IFNγ介导的Th1型炎症反应，该过程由T细胞驱动。转录组分析提示，免疫介导的PALF表型可能受复杂免疫网络调控，且代谢通路激活的证据相对较少。本研究结果为PALF的肝损伤机制提供了全新视角，可为后续研究方向及潜在治疗靶点提供参考。
