장 상피세포의 소장 내 위치별 특화 및 CD4+CD8αα+상피내 림프구의 분화에서의 역할

DoctorTissue-resident T cells reside in the tissue of residence for long period without recirculation. During the long-term residency, tissue-resident T cells receive various signals such as cytokines or antigen stimulation from their niche. These tissue specific signals contribute to the tissue adaptation of resident T cells, which involves acquisition of integrin or effector functions particularly required for the tissue. The small intestinal epithelium also harbors heterogeneous resident T cell population called intraepithelial lymphocytes (IELs). Among IELs, CD4+ IELs expressing CD8αα homodimer are defined as CD4+CD8αα+ (double positive, DP) IELs. Previous reports have shown that DP IELs are derived from tissue adaptation and maturation of conventional CD4+ T cells or induced regulatory CD4+ T cells (Tregs) reaching the epithelium. Interestingly, these CD4+ T cells experience lineage redirection involving loss of and acquisition of master regulators for CD4 and CD8 lineage, respectively, upon arrival to the epithelium. The lineage reprogramming is attributed to the gut-specific environment such as microbiota, transforming growth factor (TGF)-β, and retinoic acid (RA). In addition, recent reports showed that clonal selection of T cell receptor (TCR) preceded in early stage of the maturation of CD4+ T cells into DP IELs. It implies that the major histocompatibility complex class II (MHC II)-mediated TCR engagement occurs in the epithelium for the differentiation of DP IELs. Given that intestinal epithelial cells (IECs) constitutively express MHC II in normal condition, the involvement of IECs during the tissue adaptation of CD4+ T cells into DP IELs is highly plausible. However, the role of IECs in the process is not clearly understood. Here, regional specialization of IECs within the small intestine was identified by comparing transcriptome of IECs obtained from duodenum, jejunum, and ileum, separately. In particular, gene signature related to antigen presentation via MHC II was markedly enhanced toward distal small intestine. Accordingly, the importance of epithelial MHC II in DP IEL differentiation was investigated using IEC-specific MHC II-deficient mice. Ablation of epithelial MHC II resulted in significant reduction of DP IELs throughout the small intestine, suggesting the essential role of epithelial MHC II in DP IEL generation. Notably, coculture of gut organoids with OVA-specific TCR transgenic CD4+ T cells directly showed that cognate antigen presentation by MHC II+ organoids induced differentiation of DP cells with the aid of TGF-β and RA. Moreover, an unexpected T cell coinhibitory molecule, programmed cell death ligand-1 (PD-L1) on IECs was also found during the screening of coreceptor ligand capable of supporting DP generation in concert with MHC II. IEC-specific PD-L1-deficient mice revealed significant reduction of DP IELs in the ileum, indicating the important role of epithelial PD-L1 in differentiation of DP IELs in the distal small intestine. The requirement of PD-L1 in DP IEL differentiation was confirmed by analyzing DP IELs in PD-L1-/- mice or RAG-1-/- recipients reconstituted with splenic T cells and administered PD-L1 antagonist during the reconstitution period. Interestingly, all of epithelial MHC II and PD-L1, and frequency of DP IEL were increased toward the distal small intestine where abundant microbiota existed. According to references showing importance of microbiota in both expression of epithelial MHC II and differentiation of DP IELs, I also checked epithelial PD-L1 in germ-free (GF) or antibiotics-treated mice. Significant reduction of all of them indicated the essential role of microbiota not only for epithelial MHC II and DP IELs but also for epithelial expression of PD-L1. Furthermore, several distinct papers also reported microbiota-dependent production of IFN-γ by gut-immune cells, and the essential role of IFN-γ in both expression of epithelial MHC II and differentiation of DP IELs. Therefore, epithelial PD-L1 was also analyzed in IFN-γR-/- mice or mice receiving IFN-γ-neutralizing antibodies, revealing IFN-γ-dependency of epithelial expression of PD-L1. Collectively, these results suggest that the microbiota and IFN-γ contribute to the regional difference of epithelial expression of MHC II and PD-L1, as well as differentiation of DP IELs. In addition, series of in vitro experiments using plate-coated PD-L1 or beads coated with PD-L1 further revealed that PD-L1 ligation to CD4+ T cells promoted lineage redirection for DP generation by inducing downmodulation of CD4 master regulator, T helper–inducing POZ/Krüppel-like factor (ThPOK), via the canonical Src homology 2 domain-containing tyrosine phosphatase (SHP) pathway. In summary, I report that IECs constitute a favorable epithelial niche for differentiation of DP IELs by expressing MHC II and PD-L1, particularly in the distal small intestine. The regional specification within the small intestine was attributed to the microbiota and microbiota-dependent IFN-γ milieu. IECs support DP IEL differentiation by providing cognate TCR stimulation and coreceptor PD-1 stimulation through expression of MHC II and PD-L1, respectively. Intracellularly, CD4+ T cell intrinsic PD-1 signal suppressed the expression of CD4 master regulator ThPOK, via the canonical SHP pathway, driving lineage redirection which is critical for the generation of DP IELs. This study provides insights how tissue cells provide a niche to resident T cells for their tissue adaptation, especially as atypical antigen presenting cells