Comprehensive Gene Expression Profiling of Peyer’s Patch M Cells, Villous M-Like Cells, and Intestinal Epithelial Cells

Abstract

4 of both GALT and nasopharynx-associated lymphoid tissue, which act as a major inductive site for Ag-specific mucosal immune responses (1, 2). Recently, we also identified M cells in the small intestinal villous epithelium, at effector sites far from the FAE, suggesting that Ag sampling via villous M cells may be responsible for induction of systemic Ag-specific immune responses, such as IgG production via the oral route (3). Still missing, however, were a characterization of the shared and distinctive traits of Peyer's patches (PPs) and villous M cells and a better understanding of the immunological nature of each.Recent comprehensive gene expression analyses using microdissected FAE or whole cells dissociated from the FAE identified genes specifically expressed by PP M cells (4 -6). Similar data, however, have not been available for villous M cells, in part because sufficient numbers of M cells are difficult to isolate from the surrounding intestinal epithelial cells (IECs). In mice, lectin Ulex europaeus agglutinin-1 (UEA-1) possessing affinity for ␣ (1, 2) fucose has been routinely used for the detection of such M cells (3, 7). UEA-1, however, does not alone suffice to identify M cells because it also reacts to goblet cells (3). Our laboratory has recently succeeded in distinguishing M cells from goblet cells by developing a mAb (NKM 16-2-4 mAb) that specifically reacts to murine PP and villous M cells but not goblet cells and IECs (8). Furthermore, our recent separate studies have demonstrated that oral administration of cholera toxin (CT) as mucosal adjuvant resulted in the induction of NKM 16-2-4 mAb ϩ and UEA-1 ϩ M-like cells, which have pocket structure and Ag uptake ability, in the duodenal villous epithelium (Terahara et al., submitted for publication). These recent advances in our understanding of M cells allowed us to define gene expression profiles capable of distinguishing PP M cells, CT-induced villous M-like cells, and IECs. Materials and Methods AnimalsBALB/c mice were purchased from Japan SLC. These mice were maintained under specific pathogen-free conditions in horizontal flow cabinets in our experimental animal facility at the University of Tokyo. Following a previously established protocol (9, 10), CT (List Biologic Laboratories) was dissolved in PBS (20 g/mouse) and then orally administered to BALB/c mice. Two days after CT administration, mice were used for experiments. All animal experiments were approved by the Animal Care and Use Committee of University of Tokyo. Lectins and Abs for the detection of M cellsThe following fluorescence-conjugated lectins and Abs were used for the identification of PP and villous M cells by FACS and histochemistry: PEconjugated UEA-1 (Biogenesis), rhodamine-conjugated UEA-1 (Vector

A single-cell survey of the small intestinal epithelium

“…We annotated a cluster of 378 cells (Fig. 5a, Methods ) as differentiated M cells based on known marker gene expression 37 (Extended Data Fig. 8b–d), and used it to construct in vitro M cell-specific signatures (Extended Data Fig.…”

“…d. Expression of M cell marker genes 4–6 in each of the organoid cell clusters. Violin plots show the distribution of expression levels (log 2 (TPM+1)) for each of 10 previously reported M cell marker genes 5 (columns), in the cells (dots) in each of 13 clusters, including mature M cells (red), identified by k -NN clustering of the 5,434 scRNA-seq profiles from organoids. e,f.…”

Crohn’s disease specific pancreatic antibodies: clinical and pathophysiological challenges

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Defining a transcriptional fingerprint of murine splenic B-cell development

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