Transcription profiling of IL-4-expressing CD4 T cells in the lung and lung-draining lymph nodes
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ABSTRACT: Using single cell RNA-sequencing and whole-genome targeted transcription profiling, this study reveals that helminth infection causes the expansion of distinct but clonally related IL-4-competent CD4+ T cells in the lung and lung-draining lymphoid tissues providing insight into tissue-specific mechanisms of type-2 cytokine regulation and the lineage commitment of T-helper 2 (Th2) and T follicular helper (Tfh) cells.
Project description:Using bulk-, single cell-, ATAC-, and TCR repertoire sequencing, this study reveals that helminth infection causes the expansion of distinct IL-4-competent CD4+ T cells in the lung and lung-draining lymphoid tissues, providing insight into tissue-specific mechanisms of type-2 cytokine regulation and the lineage commitment of T-helper 2 (Th2) and T follicular helper (Tfh) cells. IL-4 competent CD4+ T cells were stratified into three groups: lymphoid-restricted T follicular helper (Tfh) cells; nonlymphoid resident T-helper 2 (Th2) cells; and transitional cells that share identity with both tissue compartments. Differences in transcriptome, locus accessibility, and the enrichment of distinct transcription factor motifs suggested that IL-4 competency was likely achieved via distinct mechanisms among lymphoid- and lung-resident CD4+IL-4+ populations. These findings reveal a complex relationship between lymphoid- and nonlymphoid-resident CD4+ T cell populations in settings of type-2 immunity.
Project description:We report the transcriptional profiles of human lung and lung draining lymph node T cell subsets that were sorted from paired samples. Naïve, central memory, effector memory and tissue resident memory CD4 and CD8 T cell subsets were sorted by FACS. Samples then underwent RNA sequencing to compare the similarities and differences between these T cell subsets and across tissues.
Project description:Germinal center (GC) like B cells can be generated in unstructured non-ectopic T and B cell infiltrates in the in inflamed lung. We used single cell RNA sequencing (scRNA-seq) for gene expression and B cell receptor (BCR) to compare antigen-specific B cells from lung to those from the lung-draining lymph node.
Project description:RNAseq was performed on human memory B cells isolated from lung, lung-draining lymph nodes and PBMCs to identify differentially expressed genes underpinning tissue localisation.
Project description:Follicular helper T (Tfh) cells have been implicated in controlling rejection after allogeneic kidney transplantation, but the precise subsets, origins and functions of Tfh cells in this process have not been fully characterized. Here we show that a subset of effector Tfh cell marked by previous IL- 21 production is potently induced during allogeneic kidney transplantation and is inhibited by immunosuppressive agents. Single-cell RNAseq revealed that these lymph node effector Tfh cells have transcriptional and clonal overlap with IL-21 producing kidney infiltrating Tfh cells, implicating common origins and developmental trajectories. To investigate the precise functions of IL-21 producing effector Tfh cells in lymph nodes and allografts, we used a mouse model to selectively eliminate these cells and assessed allogeneic B cell clonal dynamics using a single B cell culture system. We found that IL-21 producing effector Tfh cells were essential for transplant rejection by regulating donor-specific germinal center B cell clonal dynamics both systemically in the draining lymph node and locally within kidney grafts. Thus, IL-21 producing effector Tfh cells have multifaceted roles in antibody-mediated rejection after kidney transplantation by promoting B cell alloimmunity.
Project description:Topical (epicutaneous, e.c.) application of the adjuvant CpG ODN during immunization leads to a robust immune response compared to when subcutaneous (s.c.) administration. Dendritic cells are hematopoietically derived cells that are important in cross-presenting to and activating CD8 T cells. Dermal dendritic cells are one of the two major dendritic cell subsets found in the skin which mobilize from the skin to draining lymph nodes to present to T cells upon activation. Dermal dendritic cells are found in skin draining lymph nodes around 24 hours post immunization. To determine how the immune system respond differently between e.c. versus s.c. administration of CpG ODN, we evaluated changes in the skin draining lymph node environment upon the two routes of adjuvant application. Expression chemokines and chemokine receptors were assessed with real-time qPCR. To determine the changes in the skin draining lymph node environment (cytokine and cytokine receptor levels) upon immunization via real time RT-PCR.
Project description:Antigen presenting dendritic cells (DCs) and monocytes capture and transport antigens from barrier tissues for presentation to antigen-specific T cells in the draining-lymph nodes (LNs). While DCs enter LNs through afferent lymphatics in a CCR7-dependent manner, how exactly antigen-carrying monocytes reach LNs is less clear since monocytes do not express CCR7 and can also enter LNs via the bloodstream. In steady state, and following injection of several PAMPs, scRNA-seq data on LN mononuclear phagocytes identified LN resident versus migratory type 1 and type 2 conventional (c)DCs despite downregulation of DC subset-defining transcripts, such as Xcr1, Clec9a, H2-Ab1, Sirpa, and Clec10a on migratory cDCs. Migratory cDCs gained expression of transcripts controlling cellular migration such as Ccr7, Ccl17, Ccl22, and Ccl5, while migratory monocytes expressed Ccr5 without Ccr7. Using two tracking methods and a gating strategy that clearly distinguishes migratory CD88hiCD26lo monocytes from CD88-CD26hi cDCs, we found that both captured antigens in the lung and migrated to lung-draining LNs. Using global and mixed-chimeric Ccl5-, Ccr2-, Ccr5-, Ccr7-, and Batf3-deficient mice, we found that CCR5+ monocytes follow CCL5-secreting migratory cDCs to reach the draining LN via lymphatic vessels. In a model of asthma, such recruited monocytes regulated the induction of type 2 immunity. Overall, our data suggest that CCL5-secreting migratory cDCs lay down the chemokine trail for CCR5+ antigen-presenting monocytes to reach draining lymph nodes and regulate adaptive immunity.
Project description:To determine the influence of primary tumors on pre-metastatic lymph nodes, we have employed whole genome microarray expression profiling as a discovery platform to identify gene signatures of B cells from tumor-draining lymph nodes, compared with normal lymph nodes. We subcutaneously inoculated C57BL/6 mice with the 4T1 mammary carcinoma. Two weeks later, tumor-draining lymph nodes were dissociated and B cells (CD19+) were sorted. Lymph nodes B cells from normal mice without tumor bearing were set as controls.