Embryonic Pancreas Research Articles

A 3D-Printed Microdevice Encapsulates Vascularized Islets Composed of iPSC-derived β-like Cells and Microvascular Fragments for Type 1 Diabetes Treatment

Tranplantation of insulin-secreting cells provides a promising method for re-establishing the autonomous blood glucose control ability of type 1 diabetes (T1D) patients, but the low survival of the tranlplanted cells hinder the therapeutic efficacy. In this study, we 3D-printed an encapsulation system contraining β-like cells and microvascular fragments (MVF), to create a retrivable microdevice with vascularized islets in vivo for T1D therapy. The functional β-like cells were differentiated from the urine epithelial cell-derived induced pluripotent stem cells (UiPSCs). Single-cell RNA sequencing provided an integrative study and macroscopic developmental analyses of the entire process of differentiation, which revealed the developmental trajectory of differentiation in vitro follows the developmental pattern of embryonic pancreas in vivo. The MVF were isolated from the epididymal fat pad. The microdevice with a groove structure were rapidly fabricated by the digital light processing (DLP)-3D printing technology. The β-like cells and MVF were uniformly distributed in the device. After subcutaneous transplantation into C57BL/6 mice, the microdevice have less collagen accumulation and low immune cell infiltration. Moerover, the microdevice encapsulated vascularized islets reduced hyperglycemica in 33% of the treated mice for up to 100 days without immunosuppressants, and the humanized C-peptide was also detected in the serum of the mice. In summary, we described the microdevice-protecd vasculared islets for long-term treatment of T1D, with high safety and potential clinical transformative value, and may therefore provide a translatable solution to advance the research progress of β cell replacement therapy for T1D.

Constitutive Pleiotrophin Deletion Results in a Phenotype with an Altered Pancreatic Morphology and Function in Old Mice.

Pleiotrophin (PTN) is crucial for embryonic development and pancreas organogenesis as it regulates metainflammation, metabolic homeostasis, thermogenesis, and glucose tolerance. Pleiotrophin deletion is associated with a lipodystrophic phenotype in which adipose tissue plasticity is altered in late life. This study explored the impact of pleiotrophin deletion on pancreatic morphology and function in later life. We analyzed glucose tolerance and circulating parameters on female wild-type (Ptn+/+) and knock-out (Ptn-/-) mice. At 9 and 15 months, we conducted morphometric analyses of pancreatic islets and evaluated the levels of insulin, glucagon, somatostatin, glucose transporter 2 (GLUT2), vesicle-associated membrane protein 2 (VAMP2), and synaptosome-associated protein 25 (SNAP25) via immunofluorescence. The effect of PTN on glucose-stimulated insulin secretion (GSIS) was evaluated in INS1E cells and isolated islets. Ptn-/- mice showed hyperinsulinemia, impaired glucose tolerance, and increased homeostatic model assessment for insulin resistance (HOMA-IR) with age. While Ptn+/+ islets enlarge with age, in Ptn-/- mice, the median size decreased, and insulin content increased. Vesicle transport and exocytosis proteins were significantly increased in 9-month-old Ptn-/- islets. Islets from Ptn-/- mice showed impaired GSIS and decreased cell membrane localization of GLUT2 whereas, PTN increased GSIS in INS1E cells. Ptn deletion accelerated age-related changes in the endocrine pancreas, affecting islet number and size, and altering VAMP2 and SNAP25 levels and GLUT2 localization leading to impaired GSIS and insulin accumulation in islets.

9271 Cholinergic Signaling in the Mouse Embryonic Pancreas Targets Early Endocrine Cells

Abstract Disclosure: C. Duarte: None. J.K. Panzer: None. N. Borowsky: None. A. Caicedo: None. The parasympathetic vagus nerve affects pancreas function by activating local intrapancreatic neurons that release acetylcholine (ACh). These neurons are derived from progenitors of the vagal neural crest and colonize the pancreas during embryonic development. The specific mechanisms by which ACh is released from these neurons influence embryonic development of the islet and, specifically, beta cells remain largely unknown. The purpose of this study was to identify cholinergic signaling components and assess early physiological responses to ACh during murine development of endocrine lineage cells. We extracted pancreatic rudiments from mice at embryonic stages E15 to E18 and stained them for neuronal and endocrine markers, along with components associated with cholinergic signaling. We further performed Ca2+ imaging of the pancreatic rudiments from NGN3CRE-GCaMP3 mice to record responses to ACh in cells of the endocrine lineage. At E15, we observed a significantly higher concentration of glucagon+ cells compared to insulin+ cells, consistent with the earlier differentiation of alpha cells in the developing pancreas. Neuronal body clusters were identified in close proximity to endocrine lineage cells from E15 to E18, with the neuronal network becoming more dispersed and interconnected. At E15, wide distribution of the presynaptic vesicular acetylcholine transporter (VAChT) was observed later becoming localized around insulin+ and glucagon+ cells at E18. Postsynaptic acetylcholinesterase (AChE) colocalized with glucagon+ cells from E15 to E18 and with insulin+ cells by E18. ACh elicited Ca2+ responses in individual endocrine cells as early as E15. Our results show that cholinergic nerve terminals target early endocrine cells and elicit responses to ACh, indicating that local cholinergic neurons innervate and activate endocrine progenitors in the embryonic pancreas. Our current studies are aimed at determining how cholinergic signaling contributes to beta cell differentiation and function. We expect this knowledge to guide future protocols for deriving beta cells from stem cells. Presentation: 6/2/2024

8715 Cholinergic Signaling in the Mouse Embryonic Pancreas Targets Early Endocrine Cells

Abstract Disclosure: C. Duarte: None. J.K. Panzer: None. N. Borowsky: None. A. Caicedo: None. The parasympathetic vagus nerve affects pancreas function by activating local intrapancreatic neurons that release acetylcholine (ACh). These neurons are derived from progenitors of the vagal neural crest and colonize the pancreas during embryonic development. The specific mechanisms by which ACh is released from these neurons influence embryonic development of the islet and, specifically, beta cells remain largely unknown. The purpose of this study was to identify cholinergic signaling components and assess early physiological responses to ACh during murine development of endocrine lineage cells. We extracted pancreatic rudiments from mice at embryonic stages E15 to E18 and stained them for neuronal and endocrine markers, along with components associated with cholinergic signaling. We further performed Ca2+ imaging of the pancreatic rudiments from NGN3CRE-GCaMP3 mice to record responses to ACh in cells of the endocrine lineage. At E15, we observed a significantly higher concentration of glucagon+ cells compared to insulin+ cells, consistent with the earlier differentiation of alpha cells in the developing pancreas. Neuronal body clusters were identified in close proximity to endocrine lineage cells from E15 to E18, with the neuronal network becoming more dispersed and interconnected. At E15, wide distribution of the presynaptic vesicular acetylcholine transporter (VAChT) was observed later becoming localized around insulin+ and glucagon+ cells at E18. Postsynaptic acetylcholinesterase (AChE) colocalized with glucagon+ cells from E15 to E18 and with insulin+ cells by E18. ACh elicited Ca2+ responses in individual endocrine cells as early as E15. Our results show that cholinergic nerve terminals target early endocrine cells and elicit responses to ACh, indicating that local cholinergic neurons innervate and activate endocrine progenitors in the embryonic pancreas. Our current studies are aimed at determining how cholinergic signaling contributes to beta cell differentiation and function. We expect this knowledge to guide future protocols for deriving beta cells from stem cells. Presentation: 6/2/2024

3D model of mouse embryonic pancreas and endocrine compartment using stem cell-derived mesoderm and pancreatic progenitors

The developing mouse pancreas is surrounded by mesoderm compartments providing signals that induce pancreas formation. Most pancreatic organoid protocols lack this mesoderm niche and only partially capture the pancreatic cell repertoire. This work aims to generate pancreatic aggregates by differentiating mouse embryonic stem cells (mESCs) into mesoderm progenitors (MPs) and pancreas progenitors (PPs), without using Matrigel. First, mESCs were differentiated into epiblast stem cells (EpiSCs) to enhance the PP differentiation rate. Next, PPs and MPs aggregated together giving rise to various pancreatic cell types, including endocrine, acinar, and ductal cells, and to endothelial cells. Single-cell RNA sequencing analysis revealed a larger endocrine population within the PP+ MP aggregates, as compared to PPs alone or PPs in Matrigel aggregates. The PP+ MP aggregate gene expression signatures and its endocrine population percentage closely resembled those of the endocrine population found in the mouse embryonic pancreas, which holds promise for studying pancreas development.

A 3D atlas of the human developing pancreas to explore progenitor proliferation and differentiation

Aims/hypothesisRodent pancreas development has been described in great detail. On the other hand, there are still gaps in our understanding of the developmental trajectories of pancreatic cells during human ontogenesis. Here, our aim was to map the spatial and chronological dynamics of human pancreatic cell differentiation and proliferation by using 3D imaging of cleared human embryonic and fetal pancreases.MethodsWe combined tissue clearing with light-sheet fluorescence imaging in human embryonic and fetal pancreases during the first trimester of pregnancy. In addition, we validated an explant culture system enabling in vitro proliferation of pancreatic progenitors to determine the mitogenic effect of candidate molecules.ResultsWe detected the first insulin-positive cells as early as five post-conceptional weeks, two weeks earlier than previously observed. We observed few insulin-positive clusters at five post-conceptional weeks (mean ± SD 9.25±5.65) with a sharp increase to 11 post-conceptional weeks (4307±152.34). We identified a central niche as the location of onset of the earliest insulin cell production and detected extra-pancreatic loci within the adjacent developing gut. Conversely, proliferating pancreatic progenitors were located in the periphery of the epithelium, suggesting the existence of two separated pancreatic niches for differentiation and proliferation. Additionally, we observed that the proliferation ratio of progenitors ranged between 20% and 30%, while for insulin-positive cells it was 1%. We next unveiled a mitogenic effect of the platelet-derived growth factor AA isoform (PDGFAA) in progenitors acting through the pancreatic mesenchyme by increasing threefold the number of proliferating progenitors.Conclusions/interpretationThis work presents a first 3D atlas of the human developing pancreas, charting both endocrine and proliferating cells across early development.Graphical

Mettl3-Mediated m6A Methylation Controls Pancreatic Bipotent Progenitor Fate and Islet Formation.

The important role of m6A RNA modification on β cell function has been established, yet how it regulates pancreas development and endocrine differentiation remains unknown. Here, we generated transgenic mice lacking RNA methyltransferase-like 3 (Mettl3) specifically in Pdx1+ pancreatic progenitor cells and found the mutant mice developed hyperglycemia and hypo-insulinemia at 2 weeks of age, with atrophic pancreas, reduced islet mass and abnormal increase in duct formation. At E15.5, Mettl3 deletion caused a significant loss of Ngn3+ endocrine progenitor cells, which was accompanied by increased Sox9+ duct precursor cells. We identified histone deacetylase 1(Hdac1) as the critical direct m6A target in bipotent progenitor, whose degeneration caused abnormal activation of Wnt/Notch signaling pathway and blocked endocrine differentiation. This transformation could be manipulated in embryonic pancreas culture in vitro through the regulation of the axis of Mettl3-Hdac1-Wnt/Notch signaling. Our finding that Mettl3 determines endocrine lineage through modulating Hdac1 activity during the transition of bipotent progenitor might help in the development of targeted endocrine cell protocols for diabetes treatment.

Deciphering early human pancreas development at the single-cell level

Understanding pancreas development can provide clues for better treatments of pancreatic diseases. However, the molecular heterogeneity and developmental trajectory of the early human pancreas are poorly explored. Here, we performed large-scale single-cell RNA sequencing and single-cell assay for transposase accessible chromatin sequencing of human embryonic pancreas tissue obtained from first-trimester embryos. We unraveled the molecular heterogeneity, developmental trajectories and regulatory networks of the major cell types. The results reveal that dorsal pancreatic multipotent cells in humans exhibit different gene expression patterns than ventral multipotent cells. Pancreato-biliary progenitors that generate ventral multipotent cells in humans were identified. Notch and MAPK signals from mesenchymal cells regulate the differentiation of multipotent cells into trunk and duct cells. Notably, we identified endocrine progenitor subclusters with different differentiation potentials. Although the developmental trajectories are largely conserved between humans and mice, some distinct gene expression patterns have also been identified. Overall, we provide a comprehensive landscape of early human pancreas development to understand its lineage transitions and molecular complexity.

290-LB: Single-Cell Analysis of Endocrine Development Identifies Novel Roles for GLIS3 in Pancreatic ß-Cell Fate

The Krüppel-like zinc finger transcription factor GLIS3 has been identified as a critical regulator of pancreatic beta cell development and function. Mice lacking a functional Glis3 gene fail to produce enough insulin and suffer from severe hyperglycemia soon after birth. This phenotype matches what has been observed in humans with genetic mutations in GLIS3 and who have been diagnosed with permanent neonatal diabetes within the first month of life. Additional and ongoing work has begun to identify some of the transcriptional mechanisms utilized by GLIS3, yet many questions remain unanswered. One such question is whether GLIS3 has roles in endocrine development prior to the formation of functional β-cells. To address this question, I have utilized single cell RNA-sequencing of mouse embryonic pancreas, at embryonic days (e)15.5 and e18.5, of both wild type (WT) and Glis3 knockout (KO) embryos. Analysis of this data has identified the expected developmental populations, from bipotent progenitor cells to pro-endocrine cells and ultimately hormone positive β-, α-, δ-, and ε-cells. Interestingly, there is little difference in gene expression between the WT and Glis3 KO pancreas in most of the populations observed (pro-endocrine, FEV+, α-/δ-/ε-cells). However, there are significant differences in gene expression between WT β-cells and KO β-cells. Surprisingly, most of the differences are in genes upregulated in the KO cells, with only a small number of genes downregulated. These upregulated genes are normally downregulated over the course of development from FEV+ to β-cells. These data indicate that GLIS3 plays a unique role in developing β-cells in that it primarily represses genes during the course of development but is also critical for the transcriptional activation of specific targets, particularly Ins2. Understanding how GLIS3 regulates β-cell generation, and how GLIS3 itself is regulated, could potentially provide new therapeutic avenues for the treatment of diabetes. Disclosure D. W. Scoville: None. S. Grimm: None. A. M. Jetten: None. Funding National Institutes of Health (Z01-ES-101585)

Rab11 is essential to pancreas morphogenesis, lumen formation and endocrine mass

The molecular links between tissue-level morphogenesis and the differentiation of cell lineages in the pancreas remain elusive despite a decade of studies. We previously showed that in pancreas both processes depend on proper lumenogenesis. The Rab GTPase Rab11 is essential for epithelial lumen formation in vitro, however few studies have addressed its functions in vivo and none have tested its requirement in pancreas. Here, we show that Rab11 is critical for proper pancreas development. Co-deletion of the Rab11 isoforms Rab11A and Rab11B in the developing pancreatic epithelium (Rab11pancDKO) results in ∼50% neonatal lethality and surviving adult Rab11pancDKO mice exhibit defective endocrine function. Loss of both Rab11A and Rab11B in the embryonic pancreas results in morphogenetic defects of the epithelium, including defective lumen formation and lumen interconnection. In contrast to wildtype cells, Rab11pancDKO cells initiate the formation of multiple ectopic lumens, resulting in a failure to coordinate a single apical membrane initiation site (AMIS) between groups of cells. This results in an inability to form ducts with continuous lumens. Here, we show that these defects are due to failures in vesicle trafficking, as apical and junctional components remain trapped within Rab11pancDKO cells. Together, these observations suggest that Rab11 directly regulates epithelial lumen formation and morphogenesis. Our report links intracellular trafficking to organ morphogenesis in vivo and presents a novel framework for decoding pancreatic development.

USP7 controls NGN3 stability and pancreatic endocrine lineage development

Understanding the factors and mechanisms involved in beta-cell development will guide therapeutic efforts to generate fully functional beta cells for diabetes. Neurogenin 3 (NGN3) is the key transcription factor that marks endocrine progenitors and drives beta-cell differentiation. Here we screen for binding partners of NGN3 and identify the deubiquitylating enzyme USP7 as a key regulator of NGN3 stability. Mechanistically, USP7 interacts with, deubiquitinates and stabilizes NGN3. In vivo, conditional knockout of Usp7 in the mouse embryonic pancreas causes a dramatic reduction in islet formation and hyperglycemia in adult mice, due to impaired NGN3-mediated endocrine specification during pancreatic development. Furthermore, pharmacological inhibition of USP7 during endocrine specification in human iPSC models of beta-cell differentiation decreases NGN3 expressing progenitor cell numbers and impairs beta cell differentiation. Thus, the USP7-NGN3 axis is an essential mechanism for driving endocrine development and beta-cell differentiation, which can be therapeutically exploited.

Focal adhesion kinase-mediated signaling controls the onset of pancreatic cell differentiation.

ABSTRACTSignals from the endothelium play a pivotal role in pancreatic lineage commitment. As such, the fate of the epithelial cells relies heavily on the spatiotemporal recruitment of the endothelial cells to the embryonic pancreas. Although it is known that VEGFA secreted by the epithelium recruits the endothelial cells to the specific domains within the developing pancreas, the mechanism that controls the timing of such recruitment is poorly understood. Here, we have assessed the role of focal adhesion kinase (FAK) in mouse pancreatic development based on our observation that the presence of the enzymatically active form of FAK (pFAK) in the epithelial cells is inversely correlated with vessel recruitment. To study the role of FAK in the pancreas, we conditionally deleted the gene encoding focal adhesion kinase in the developing mouse pancreas. We found that homozygous deletion of Fak (Ptk2) during embryogenesis resulted in ectopic epithelial expression of VEGFA, abnormal endothelial recruitment and a delay in endocrine and acinar cell differentiation. The heterozygous mutants were born with no pancreatic phenotype but displayed gradual acinar atrophy due to cell polarity defects in exocrine cells. Together, our findings imply a role for FAK in controlling the timing of pancreatic lineage commitment and/or differentiation in the embryonic pancreas by preventing endothelial recruitment to the embryonic pancreatic epithelium.

A novel approach to describing the pancreas and submandibular gland: Can they be classified as primary and secondary tissue organs?

Our aim is to examine the relationship between the submandibular gland (SMG) and the pancreas by discussing similarities and differences in their embryology, histology, physiology, and pathology, and to introduce the question of classifying them as primary or secondary organs. From an embryonic standpoint, SMG and pancreas originate from different germ layers, yet they share a variety of similar elements in their morphogenesis, branching process, and mesenchymal molecular interaction. The histological and anatomical comparison between these two organs reveals parallels in the basic function of their exocrine physiology. With both the SMG and pancreas playing a significant role in digestive processes, there are also common aspects in their exocrine function. Furthermore, recent research has unraveled an intricate novel system of hormonal interaction between the salivary glands and pancreas which regulates pancreatic cellular differentiation and injury repair mechanism. Lastly, there are analogous features in the pathological mechanisms and inflammatory processes in the course of chronic disease in both organs. By defining this close relationship between the SMG and the pancreas, we aim to provide alternative insights for scholars and physicians in the shared characteristics of basic function of these organs, and possible pathological consequences of their dysregulation.

1450-P: Identifying Hidden Sources of Tissue Plasticity in Pancreatic Islets with a Novel Multicolor Flow Cytometry Approach

Identifying pancreatic islet cells that acquire a beta-cell like phenotype is important for developing new diabetes therapies. Our studies have revealed a novel serpinB13 protease inhibitor antibody, that augments cathepsin L protease activity, resulting in Notch1 receptor cleavage and mobilization of endocrine progenitor cells to become beta cells in the embryonic pancreas. It is unclear, however, whether a similar induction of additional beta cells occurs with exposure to serpinB13 monoclonal antibody (mAb) in later life. To explore the exact source (s) of new beta cells in response to serpinB13 mAb after birth we developed a novel, multicolor flow cytometry analysis approach to study pancreatic islets. Five-week old normal Balb/c mice were injected with serpinB13 mAb (or control IgG) , and sacrificed at 8 weeks for islet isolation. Islet cell suspensions were stained with a viability dye, and a panel of antibodies to CD31, CD45, cytokeratin 19, EpCAM, insulin (Ins) , glucagon (Glu) , somatostatin (SS) , c-peptide, and four transcription factors, Maf-A, Nkx6.1, NeuroD1, and Pdx-1. Exposure to the serpinB13 mAb caused a significant increase in the number of hormonal-negative (e.g., Ins-Glu-SS-c-peptide-) cells (p=0.0011) , and hormonal negative, but c-peptide positive, cells (p=0.002) . At least one of four transcription factors examined were positive in 5-10% and 10-30% of these populations, respectively. At the same time, there was a trend in the drop of Glu+ and SS+ cells. The average number of pancreatic islets, and the total islet cellularity per mouse, was augmented in animals receiving serpinB13 mAb. Although additional studies are necessary to characterize the hormonal negative cells and assess whether Glu+ and SS+ cells transdifferentiate to beta cells, these findings demonstrate that enhanced tissue plasticity and beta-cell output can be achieved in the pancreas following treatment with antibodies to protease inhibitors. Disclosure Y.Kryvalap: None. S.Z.Meng: None. J.Czyzyk: None.

Generation of human islet cell type-specific identity genesets

Generation of surrogate cells with stable functional identities is crucial for developing cell-based therapies. Efforts to produce insulin-secreting replacement cells to treat diabetes require reliable tools to assess islet cellular identity. Here, we conduct a thorough single-cell transcriptomics meta-analysis to identify robustly expressed markers used to build genesets describing the identity of human α-, β-, γ- and δ-cells. These genesets define islet cellular identities better than previously published genesets. We show their efficacy to outline cell identity changes and unravel some of their underlying genetic mechanisms, whether during embryonic pancreas development or in experimental setups aiming at developing glucose-responsive insulin-secreting cells, such as pluripotent stem-cell differentiation or in adult islet cell reprogramming protocols. These islet cell type-specific genesets represent valuable tools that accurately benchmark gain and loss in islet cell identity traits.

Mitofusin2 Promotes β Cell Maturation from Mouse Embryonic Stem Cells via Sirt3/Idh2 Activation.

β cell dysfunction is the leading cause of diabetes. Adult β cells have matured glucose-stimulated insulin secretion (GSIS), whereas fetal and neonatal β cells are insensitive to glucose and are functionally immature. However, how β cells mature and acquire robust GSIS is not fully understood. Here, we explored the potential regulatory proteins of β cell maturation process and the capacity for GSIS. Combined with the data from public databases, we found that the gene expression of Mitofusin2 (Mfn2) showed an increasing trend from mouse neonatal β cells to mature β cells. Moreover, its protein expression increased during mouse embryonic pancreas development and β cell differentiation from mouse embryonic stem cells. Knocking down Mfn2 reduced Urocortin3 (Ucn3) expression, GSIS, and ATP production in induced β cells, while overexpressing it had the opposite effect. However, neither Mfn2 knockdown nor overexpression affected the differentiation rate of insulin-positive cells. In immature and mature β cells, Mfn2 and its correlated genes were enriched in tricarboxylic acid (TCA) cycle-related pathways. The expressions of Sirtuin 3 (Sirt3) and isocitrate dehydrogenase 2 (NADP+) and mitochondrial (Idh2) were Mfn2-regulated during β cell differentiation. Inhibiting Idh2 or Sirt3 reduced cellular ATP content and insulin secretion levels that increased by Mfn2 overexpression. Thus, Mfn2 modulated the induced β cell GSIS by influencing the TCA cycle through Sirt3/Idh2 activation. We demonstrated that Mfn2 promoted embryonic stem cell-derived β cell maturation via the Sirt3/Idh2 pathway, providing new insights into β cell development. Our data contribute to understanding diabetes pathogenesis and offer potential new targets for β cell regeneration therapies.

Development of a 3D atlas of the embryonic pancreas for topological and quantitative analysis of heterologous cell interactions.

ABSTRACTGenerating comprehensive image maps, while preserving spatial three-dimensional (3D) context, is essential in order to locate and assess quantitatively specific cellular features and cell-cell interactions during organ development. Despite recent advances in 3D imaging approaches, our current knowledge of the spatial organization of distinct cell types in the embryonic pancreatic tissue is still largely based on two-dimensional histological sections. Here, we present a light-sheet fluorescence microscopy approach to image the pancreas in three dimensions and map tissue interactions at key time points in the mouse embryo. We demonstrate the utility of the approach by providing volumetric data, 3D distribution of three main cellular components (epithelial, mesenchymal and endothelial cells) within the developing pancreas, and quantification of their relative cellular abundance within the tissue. Interestingly, our 3D images show that endocrine cells are constantly and increasingly in contact with endothelial cells forming small vessels, whereas the interactions with mesenchymal cells decrease over time. These findings suggest distinct cell-cell interaction requirements for early endocrine cell specification and late differentiation. Lastly, we combine our image data in an open-source online repository (referred to as the Pancreas Embryonic Cell Atlas).

Lineage Tracing Reveals the Dynamic Contribution of Id2+ Progenitor Cells to Branching Morphogenesis.

Branching morphogenesis is an important process in shaping the arborized structures of several organs. However, the driving force that directs this process from progenitor pools remains incompletely understood. In this lineage tracing study, we investigated the role of Id2+ embryonic progenitor cells in branching organs such as the pancreas, kidney, mammary gland, thyroid gland, and salivary gland. We found that a subset of Id2+ distal progenitor cells in the embryonic pancreas and kidney can give rise to multiple lineages of progeny cells during branching morphogenesis. Id2-labeled cells also supported the postnatal development of the mammary glands. However, Id2+ cells did not contribute to the development of the salivary and thyroid glands. We found the Id2+ cells located in the tip progenitor pools of pancreas and kidney have self-renewal potential and contribute descendants to multiple epithelial cell lineages. Our findings enrich the current model of distal progenitor pools driving branching morphogenesis and provide a new marker to investigate the regularity of branching in these organs.

Stem/progenitor cells in normal physiology and disease of the pancreas

Though embryonic pancreas progenitors are well characterised, the existence of stem/progenitor cells in the postnatal mammalian pancreas has been long debated, mainly due to contradicting results on regeneration after injury or disease in mice. Despite these controversies, sequencing advancements combined with lineage tracing and organoid technologies indicate that homeostatic and trigger-induced regenerative responses in mice could occur. The presence of putative progenitor cells in the adult pancreas has been proposed during homeostasis and upon different stress challenges such as inflammation, tissue damage and oncogenic stress. More recently, single cell transcriptomics has revealed a remarkable heterogeneity in all pancreas cell types, with some cells showing the signature of potential progenitors. In this review we provide an overview on embryonic and putative adult pancreas progenitors in homeostasis and disease, with special emphasis on in vitro culture systems and scRNA-seq technology as tools to address the progenitor nature of different pancreatic cells.

REST is a major negative regulator of endocrine differentiation during pancreas organogenesis.

Multiple transcription factors have been shown to promote pancreatic β-cell differentiation, yet much less is known about negative regulators. Earlier epigenomic studies suggested that the transcriptional repressor REST could be a suppressor of endocrinogenesis in the embryonic pancreas. However, pancreatic Rest knockout mice failed to show abnormal numbers of endocrine cells, suggesting that REST is not a major regulator of endocrine differentiation. Using a different conditional allele that enables profound REST inactivation, we observed a marked increase in pancreatic endocrine cell formation. REST inhibition also promoted endocrinogenesis in zebrafish and mouse early postnatal ducts and induced β-cell-specific genes in human adult duct-derived organoids. We also defined genomic sites that are bound and repressed by REST in the embryonic pancreas. Our findings show that REST-dependent inhibition ensures a balanced production of endocrine cells from embryonic pancreatic progenitors.