Chronic Disease Program, Ottawa Hospital Research Institute, Ottawa, ON, Canada.
Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada.
Front Immunol. 2024 Jun 3;15:1375485. doi: 10.3389/fimmu.2024.1375485. eCollection 2024.
The impact of chronic hepatic infection on antigen non-specific immune cells in circulation remains poorly understood. We reported lasting global hyperfunction of peripheral CD8 T cells in HCV-infected individuals with cirrhosis. Whether gene expression patterns in bulk CD8 T cells are associated with the severity of liver fibrosis in HCV infection is not known.
RNA sequencing of blood CD8 T cells from treatment naïve, HCV-infected individuals with minimal (Metavir F0-1 ≤ 7.0 kPa) or advanced fibrosis or cirrhosis (F4 ≥ 12.5 kPa), before and after direct-acting antiviral therapy, was performed. CD8 T cell function was assessed by flow cytometry.
In CD8 T cells from pre-DAA patients with advanced compared to minimal fibrosis, Gene Ontology analysis and Gene Set Enrichment Analysis identified differential gene expression related to cellular function and metabolism, including upregulated Hedgehog (Hh) signaling, IFN-α, -γ, TGF-β response genes, apoptosis, apical surface pathways, phospholipase signaling, phosphatidyl-choline/inositol activity, and second-messenger-mediated signaling. In contrast, genes in pathways associated with nuclear processes, RNA transport, cytoskeletal dynamics, cMyc/E2F regulation, oxidative phosphorylation, and mTOR signaling, were reduced. Hh signaling pathway was the top featured gene set upregulated in cirrhotics, wherein hallmark genes and ranked highly. Inhibition of Smo-dependent Hh signaling ablated the expression of IFN-γ and perforin in stimulated CD8 T cells from chronic HCV-infected patients with advanced compared to minimal fibrosis. CD8 T cell gene expression profiles post-DAA remained clustered with pre-DAA profiles and disparately between advanced and minimal fibrosis, suggesting a persistent perturbation of gene expression long after viral clearance.
This analysis of bulk CD8 T cell gene expression in chronic HCV infection suggests considerable reprogramming of the CD8 T cell pool in the cirrhotic state. Increased Hh signaling in cirrhosis may contribute to generalized CD8 T cell hyperfunction observed in chronic HCV infection. Understanding the lasting nature of immune cell dysfunction may help mitigate remaining clinical challenges after HCV clearance and more generally, improve long term outcomes for individuals with severe liver disease.
慢性肝感染对循环中抗原非特异性免疫细胞的影响仍知之甚少。我们报道了 HCV 感染伴肝硬化患者外周 CD8 T 细胞持续的整体高功能。在 HCV 感染中， bulk CD8 T 细胞的基因表达模式是否与肝纤维化的严重程度相关尚不清楚。
对未经治疗的、HCV 感染的个体的血液 CD8 T 细胞进行 RNA 测序，这些个体的纤维化程度为最小（Metavir F0-1 ≤ 7.0 kPa）或进展（F4 ≥ 12.5 kPa），在直接作用抗病毒治疗之前和之后进行。通过流式细胞术评估 CD8 T 细胞的功能。
与最小纤维化相比，在进展性纤维化的 pre-DAA 患者的 CD8 T 细胞中，通过基因本体论分析和基因集富集分析发现，与细胞功能和代谢相关的差异基因表达，包括 Hedgehog（Hh）信号上调、IFN-α、-γ、TGF-β反应基因、凋亡、顶端表面途径、磷脂酶信号、磷酸胆碱/肌醇活性和第二信使介导的信号。相反，与核过程、RNA 转运、细胞骨架动力学、cMyc/E2F 调节、氧化磷酸化和 mTOR 信号相关的途径中的基因减少。Hh 信号通路是在肝硬化患者中上调的最显著的基因集，其中标志性基因 和 排名很高。在慢性 HCV 感染进展性纤维化患者中，抑制 Smo 依赖性 Hh 信号可使刺激后的 CD8 T 细胞中 IFN-γ和穿孔素的表达减少。DAA 后 CD8 T 细胞的基因表达谱仍然与 DAA 前的基因表达谱聚类，并且在进展性纤维化和最小纤维化之间差异很大，这表明在病毒清除后很长一段时间内，基因表达仍然存在持续的干扰。
这项对慢性 HCV 感染中 bulk CD8 T 细胞基因表达的分析表明，在肝硬化状态下，CD8 T 细胞池发生了相当大的重编程。肝硬化中 Hh 信号的增加可能导致慢性 HCV 感染中观察到的普遍 CD8 T 细胞高功能。了解免疫细胞功能障碍的持久性可能有助于减轻 HCV 清除后仍然存在的临床挑战，更广泛地说，改善严重肝病患者的长期预后。
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