【文献阅读031】肝脏T细胞单细胞图谱揭示了PSC患者中肝脏驻留初始样CD4+T细胞的增加

Background & Aims:Little is known about the composition of intrahepatic immune cells and their contribution to the pathogenesis of primary sclerosing cholangitis (PSC). Herein, we aimed to create an atlas of intrahepatic T cells and thereby perform an in-depth characterization of T cells in inflamed human liver.

Methods: Different single-cell RNA sequencing methods were combined with in silico analyses on intrahepatic and peripheral T cells from patients with PSC (n = 11) and healthy donors (HDs, n=4). Multi-parameter flow cytometry and functional in vitro experiments were conducted on samples from patients with PSC(n=24), controls with other liver diseases and HDs.

Results: We identified a population of intrahepatic naive-like CD4+ T cells, which was present in all liver diseases tested, but particularly expanded in PSC. This population had a transcriptome and T cell receptor repertoire similar to circulating naive T cells but expressed a set of genes associated with tissue residency. Their periductal location supported the concept of tissue-resident naive-like T cells in livers of patients with PSC. Trajectory inference suggested that these cells had the developmental propensity to acquire a T helper 17 (TH17) polarization state. Functional and chromatin accessibility experiments revealed that circulating naive T cells in patients with PSC were predisposed to polarize towards TH17 cells.

Conclusion: We report the first atlas of intrahepatic T cells in PSC, which led to the identification of a previously unrecognized population of tissue-resident naive-like T cells in the inflamed human liver and to the finding that naive CD4+ T cells in PSC harbour the propensity to develop into TH17 cells.

Lay summary: The composition of intrahepatic immune cells in primary sclerosing cholangitis (PSC) and their contribution to disease pathogenesis is widely unknown. We analysed intrahepatic T cells and identified a previously uncharacterized population of liver-resident CD4+ T cells which are expanded in the livers of patients with PSC compared to healthy liver tissue and other liver diseases. These cells are likely to contribute to the pathogenesis of PSC and could be targeted in novel therapeutic approaches.

Genome-wide association studies (GWAS) implicated T cells in the pathogenesis of PSC^7–9 and phenotypic changes of T cells were detected in livers and peripheral blood of patients with PSC.

Recent studies highlighted the power of single-cell transcriptomics to unravel the heterogeneity of liver-parenchymal cells,^10,11 whereas little is known about the composition and function of immune cells isolated from inflamed human livers. 

Although intrahepatic immune cells mostly comprise innate cells, liver-resident T cells have emerged as central mediators of liver injury.^12–14 

Tissue residency is a feature assigned to memory T cells, which is established during the immune response in nonlymphoid tissues.¹⁵ 

In contrast to memory T cells, naive T cells usually circulate between blood and lymph nodes and whether they are able to reside in non-lymphoid tissues such as the liver has never been substantiated in humans.¹⁶ 

Few studies have analysed the intrahepatic T cell compartment in PSC,^17–21 mostly due to the rarity of the disease and the limited availability of tissue specimens.

Despite an overall similarity, we observed selective expression of CD69, a hallmark of tissue residency, and CXCR4, which is associated with T cell homing towards the biliary epithelium,²⁹ in the intrahepatic naive-like CD4+ T cells (Fig. 3B). To specifically evaluate the expression of tissue residency genes in naivelike CD4+ T cells, we tested the accordance of these cells to literature-based gene profiles of CD69+ tissue-resident T cells.³⁰ The expression of a signature associated with tissue residency was seen in liver-derived naive-like CD4+ T cells (Fig. 3C).

naive T cells being primed in lymphoid tissues and polarized towards effector cells depending on the microenvironment¹⁶

A hallmark of naive T cells is their high expression of CCR7 and CD62L,^27,28 which were highly expressed by the naive-like CD4+ T cells.The expression of the ligands to these molecules, CCL21 and MAdCAM-1, respectively, has been reported within livers of patients with PSC.^19,39–41 Of note, the expression of other ligands of MAdCAM-1, e.g. integrins alpha-4 and beta-7, which were supposed to facilitate aberrant liver homing of gut-primed cells,¹⁷ was not detectable within these cells.

Interestingly, the identified population of intrahepatic naive-like CD4+ T cells expressed CD69 and CXCR4, genes required for activation and homing towards the biliary epithelium within the liver.²⁹

In this study, we report that intrahepatic naive-like CD4+ T cells are prone to develop a TH17 rather than a Foxp3+ TREG polarization state in patients with PSC. Of note, intrahepatic naive-like CD4+ T cells were present in all liver diseases analysed, but their abundance was highest in patients with PSC. TH17 cells have been implicated in the pathogenesis and disease progression of PSC and other inflammatory liver diseases, such as HCV and NASH.^20,42–45 Recent translational studies suggested that TH17 cells can be induced by PSC-derived intestinal microbiota and that targeting these TH17 cells improved cholangitis in mice.⁴⁴ The finding that epigenetic imprinting contributes to the propensity to develop into TH17 cells should spark interest into factors promoting chromatin accessibility, which may even be of microbial origin.

Considering the findings of altered intestinal^5,6 and also biliary microbiota⁴ in patients with PSC, and the increased propensity of T cells to differentiate into TH17 cells upon challenge by pathogens,^44–46 it is tempting to speculate that microbiota are involved in the activation and differentiation of intrahepatic naive-like CD4+ T cells.^44,45,47 Evidence for the differentiation of naive T cells within the liver has already been shown in mice.⁴⁸ Along this line, we have recently shown that the interplay of biliary epithelial cells and hepatic monocytes, responding to pathogens, promotes a TH17-polarizing microenvironment in the context of PSC.⁴⁶