Synaptotagmin 13 orchestrates pancreatic endocrine cell egression and islet morphogenesis

Epithelial cell egression is important for organ development, but also drives cancer metastasis. Better understandings of pancreatic epithelial morphogenetic programs generating islets of Langerhans aid to diabetes therapy. Here we identify the Ca2+-independent atypical Synaptotagmin 13 (Syt13) as a key driver of endocrine cell egression and islet formation. We detected upregulation of Syt13 in endocrine precursors that correlates with increased expression of unique cytoskeletal components. High-resolution imaging reveals a previously unidentified apical-basal to front-rear repolarization during endocrine cell egression. Strikingly, Syt13 interacts with acetylated tubulin and phosphatidylinositol phospholipids and localizes to the leading-edge of egressing cells. Knockout of Syt13 impairs endocrine cell egression and skews the α- to-β-cell ratio. Mechanistically, Syt13 regulates endocytosis to remodel the basement membrane and cell-matrix adhesion at the leading-edge of egressing endocrine cells. Altogether, these findings implicate an unexpected role of Syt13 in regulating cell polarity to orchestrate endocrine cell egression and islet morphogenesis.

3D Atlas of the Pituitary Gland of the Model Fish Medaka (Oryzias latipes)

In vertebrates, the anterior pituitary plays a crucial role in regulating several essential physiological processes via the secretion of at least seven peptide hormones by different endocrine cell types. Comparative and comprehensive knowledge of the spatial distribution of those endocrine cell types is required to better understand their physiological functions. Using medaka as a model and several combinations of multi-color fluorescence in situ hybridization, we present the first 3D atlas revealing the gland-wide distribution of seven endocrine cell populations: lactotropes, thyrotropes, Lh and Fsh gonadotropes, somatotropes, and pomca-expressing cells (corticotropes and melanotropes) in the anterior pituitary of a teleost fish. By combining in situ hybridization and immunofluorescence techniques, we deciphered the location of corticotropes and melanotropes within the pomca-expressing cell population. The 3D localization approach reveals sexual dimorphism of tshba-, pomca-, and lhb-expressing cells in the adult medaka pituitary. Finally, we show the existence of bi-hormonal cells co-expressing lhb-fshb, fshb-tshba and lhb-sl using single-cell transcriptomics analysis and in situ hybridization. This study offers a solid basis for future comparative studies of the teleost pituitary and its functional plasticity.

H3K4 Trimethylation is Required for Postnatal Pancreatic Endocrine Cell Functional Maturation

During pancreas development, endocrine progenitors differentiate into the islet-cell subtypes, which undergo further functional maturation in postnatal islet development. In islet b-cells, genes involved in glucose-stimulated insulin secretion are activated and glucose exposure increases the insulin response as b-cells mature. Here, we investigated the role of H3K4 trimethylation in endocrine cell differentiation and functional maturation by disrupting TrxG complex histone methyltransferase activity in mouse endocrine progenitors. In the embryo, genetic inactivation of TrxG component <i>Dpy30</i> in NEUROG3+ cells did not affect the number of endocrine progenitors or endocrine cell differentiation. H3K4 trimethylation was progressively lost in postnatal islets and the mice displayed elevated non-fasting and fasting glycemia, as well as impaired glucose tolerance by postnatal day 24. Although postnatal endocrine cell proportions were equivalent to controls, islet RNA-sequencing revealed a downregulation of genes involved in glucose-stimulated insulin secretion and an upregulation of immature b-cell genes. Comparison of histone modification enrichment profiles in NEUROG3+ endocrine progenitors and mature islets suggested that genes downregulated by loss of H3K4 trimethylation more frequently acquire active histone modifications during maturation. Taken together, these findings suggest that H3K4 trimethylation is required for the activation of genes involved in the functional maturation of pancreatic islet endocrine cells.

Single-Cell RNA Sequencing Analysis of Chicken Anterior Pituitary: A Bird’s-Eye View on Vertebrate Pituitary

It is well-established that anterior pituitary contains multiple endocrine cell populations, and each of them can secrete one/two hormone(s) to regulate vital physiological processes of vertebrates. However, the gene expression profiles of each pituitary cell population remains poorly characterized in most vertebrate groups. Here we analyzed the transcriptome of each cell population in adult chicken anterior pituitaries using single-cell RNA sequencing technology. The results showed that: (1) four out of five known endocrine cell clusters have been identified and designated as the lactotrophs, thyrotrophs, corticotrophs, and gonadotrophs, respectively. Somatotrophs were not analyzed in the current study. Each cell cluster can express at least one known endocrine hormone, and novel marker genes (e.g., CD24 and HSPB1 in lactotrophs, NPBWR2 and NDRG1 in corticotrophs; DIO2 and SOUL in thyrotrophs, C5H11ORF96 and HPGDS in gonadotrophs) are identified. Interestingly, gonadotrophs were shown to abundantly express five peptide hormones: FSH, LH, GRP, CART and RLN3; (2) four non-endocrine/secretory cell types, including endothelial cells (expressing IGFBP7 and CFD) and folliculo-stellate cells (FS-cells, expressing S100A6 and S100A10), were identified in chicken anterior pituitaries. Among them, FS-cells can express many growth factors, peptides (e.g., WNT5A, HBEGF, Activins, VEGFC, NPY, and BMP4), and progenitor/stem cell-associated genes (e.g., Notch signaling components, CDH1), implying that the FS-cell cluster may act as a paracrine/autocrine signaling center and enrich pituitary progenitor/stem cells; (3) sexually dimorphic expression of many genes were identified in most cell clusters, including gonadotrophs and lactotrophs. Taken together, our data provides a bird’s-eye view on the diverse aspects of anterior pituitaries, including cell composition, heterogeneity, cell-to-cell communication, and gene expression profiles, which facilitates our comprehensive understanding of vertebrate pituitary biology.

3D atlas of the pituitary gland of the model fish medaka

In vertebrates, the anterior pituitary plays a crucial role in regulating several essential physiological processes via the secretion of at least seven peptide hormones by different endocrine cell types. Comparative and comprehensive knowledge of the spatial distribution of those endocrine cell types is required to better understand their role during the animal life. Using medaka as the model and several combinations of multi-color fluorescence in situ hybridization, we present the first 3D atlas revealing the gland-wide distribution of seven endocrine cell populations: lactotropes, thyrotropes, Lh and Fsh gonadotropes, somatotropes, and pomca-expressing cells (corticotropes and melanotropes) in the anterior pituitary of a teleost fish. By combining in situ hybridization and immunofluorescence techniques, we deciphered the location of corticotropes and melanotropes within the pomca-expressing cell population. The 3D localization approach reveals sexual dimorphism of tshba- and pomca-expressing cells in the adult medaka pituitary. Finally, we show the existence of bi-hormonal cells co-expressing lhb-fshb, fshb-tshba and lhb-sl using single-cell transcriptomics analysis and in situ hybridization. This study offers a solid basis for future comparative studies of the teleost pituitary and its developmental plasticity.

A Gastric Intramucosal Mixed Adenocarcinoma-Neuroendocrine Tumor

Gastric mixed adenocarcinoma-neuroendocrine tumor (NET) is a rare composite tumor, and a limited number of studies have reported on it. A 77-year-old man was admitted to our hospital because of acute cholecystitis. He underwent a cholecystectomy. Esophagogastroduodenoscopy during his admission revealed a slightly elevated tumor, and biopsy demonstrated a well-differentiated tubular adenocarcinoma. The tumor was resected completely by endoscopic submucosal dissection. Histological findings showed that it measured 9 mm in diameter, was located within the mucosa, and consisted of well-differentiated tubular adenocarcinoma and a NET G1. The NET was covered with adenocarcinoma and both components exhibited histological continuity. The NET and a part of the adenocarcinoma component showed a positive reaction for chromogranin A and synaptophysin. Neither enterochromaffin-like cell hyperplasia nor endocrine cell micronest surrounded the tumor. The diagnosis was gastric mixed adenocarcinoma-NET. The histological continuity between the two components can be likened to the same histogenesis.

The SNAG Domain of Insm1 Regulates Pancreatic Endocrine Cell Differentiation and Represses Beta- to Delta-Cell Transdifferentiation

The allocation and specification of pancreatic endocrine lineages are tightly regulated by transcription factors. Disturbances in differentiation of these lineages contribute to the development of various metabolic diseases, including diabetes. The Insulinoma-associated protein 1 (<i>Insm1</i>), which encodes a protein containing one SNAG domain and five zinc fingers, plays essential roles in pancreatic endocrine cell differentiation and in mature beta-cell function. In the present study, we compared the differentiation of pancreatic endocrine cells between Insm1 null and Insm1 SNAG domain mutants (Insm1delSNAG) to explore the specific function of the SNAG domain of Insm1. We show that the delta-cell number is increased in Insm1delSNAG but not in Insm1 null mutants as compared to the control mice. We also show a less severe reduction of the beta-cell number in Insm1delSNAG as that in Insm1 null mutants. In addition, similar deficits are observed in alpha-, PP- and epsilon-cell in Insm1delSNAG and Insm1 null mutants. We further identified that the increased delta-cell number is due to beta- to delta-cell transdifferentiation. Mechanistically, the SNAG domain of Insm1 interacts with Lsd1, the demethylase of H3K4me1/2. Mutation in the SNAG domain of Insm1 results in impaired recruitment of Lsd1 and increased H3K4me1/2 levels at <i>H</i><i>hex</i> loci that are bound by Insm1, thereby promoting the transcriptional activity of the delta-cell-specific gene <i>Hhex</i>. Our study has identified a novel function of the SNAG domain of Insm1 in the regulation of pancreatic endocrine cells differentiation, particularly in the repression of beta- to delta-cell transdifferentiation.

The SNAG Domain of Insm1 Regulates Pancreatic Endocrine Cell Differentiation and Represses Beta- to Delta-Cell Transdifferentiation

The allocation and specification of pancreatic endocrine lineages are tightly regulated by transcription factors. Disturbances in differentiation of these lineages contribute to the development of various metabolic diseases, including diabetes. The Insulinoma-associated protein 1 (<i>Insm1</i>), which encodes a protein containing one SNAG domain and five zinc fingers, plays essential roles in pancreatic endocrine cell differentiation and in mature beta-cell function. In the present study, we compared the differentiation of pancreatic endocrine cells between Insm1 null and Insm1 SNAG domain mutants (Insm1delSNAG) to explore the specific function of the SNAG domain of Insm1. We show that the delta-cell number is increased in Insm1delSNAG but not in Insm1 null mutants as compared to the control mice. We also show a less severe reduction of the beta-cell number in Insm1delSNAG as that in Insm1 null mutants. In addition, similar deficits are observed in alpha-, PP- and epsilon-cell in Insm1delSNAG and Insm1 null mutants. We further identified that the increased delta-cell number is due to beta- to delta-cell transdifferentiation. Mechanistically, the SNAG domain of Insm1 interacts with Lsd1, the demethylase of H3K4me1/2. Mutation in the SNAG domain of Insm1 results in impaired recruitment of Lsd1 and increased H3K4me1/2 levels at <i>H</i><i>hex</i> loci that are bound by Insm1, thereby promoting the transcriptional activity of the delta-cell-specific gene <i>Hhex</i>. Our study has identified a novel function of the SNAG domain of Insm1 in the regulation of pancreatic endocrine cells differentiation, particularly in the repression of beta- to delta-cell transdifferentiation.