Reconstituting pancreas development from purified progenitor cells reveals genes essential for islet differentiation.
Ontology highlight
ABSTRACT:
SUBMITTER: Sugiyama T
PROVIDER: S-EPMC3732989 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:As one of the key nutrient sensors, insulin signaling plays an important role in integrating environmental energy cues with organism growth. In adult organisms, relative insufficiency of insulin signaling induces compensatory expansion of insulin-secreting pancreatic beta (?) cells. However, little is known about how insulin signaling feedback might influence neogenesis of ? cells during embryonic development. Using genetic approaches and a unique cell transplantation system in developing zebrafish, we have uncovered a novel role for insulin signaling in the negative regulation of pancreatic progenitor cell differentiation. Blocking insulin signaling in the pancreatic progenitors hastened the expression of the essential ? cell genes insulin and pdx1, and promoted ? cell fate at the expense of alpha cell fate. In addition, loss of insulin signaling promoted ? cell regeneration and destabilization of alpha cell character. These data indicate that insulin signaling constitutes a tunable mechanism for ? cell compensatory plasticity during early development. Moreover, using a novel blastomere-to-larva transplantation strategy, we found that loss of insulin signaling in endoderm-committed blastomeres drove their differentiation into ? cells. Furthermore, the extent of this differentiation was dependent on the function of the ? cell mass in the host. Altogether, our results indicate that modulation of insulin signaling will be crucial for the development of ? cell restoration therapies for diabetics; further clarification of the mechanisms of insulin signaling in ? cell progenitors will reveal therapeutic targets for both in vivo and in vitro ? cell generation.
| S-EPMC4950988 | biostudies-literature
Project description:Development of systems that reconstitute hallmark features of human pancreatic intraepithelial neoplasia (PanINs), the precursor to pancreatic ductal adenocarcinoma, could generate new strategies for early diagnosis and intervention. However, human cell-based PanIN models with defined mutations are unavailable. Here, we report that genetic modification of primary human pancreatic cells leads to development of lesions resembling native human PanINs. Primary human pancreas duct cells harbouring oncogenic KRAS and induced mutations in CDKN2A, SMAD4 and TP53 expand in vitro as epithelial spheres. After pancreatic transplantation, mutant clones form lesions histologically similar to native PanINs, including prominent stromal responses. Gene expression profiling reveals molecular similarities of mutant clones with native PanINs, and identifies potential PanIN biomarker candidates including Neuromedin U, a circulating peptide hormone. Prospective reconstitution of human PanIN development from primary cells provides experimental opportunities to investigate pancreas cancer development, progression and early-stage detection.
| S-EPMC5344977 | biostudies-literature
Project description:Isl1 is a LIM homeobox transcription factor showing conserved expression in the developing and mature vertebrate pancreas. So far, functions of pancreatic Isl1 have mainly been studied in the mouse, where Isl1 has independent functions during formation of exocrine and endocrine tissues. Here, we take advantage of a recently described isl1 mutation in zebrafish to address pancreatic isl1 functions in a non-mammalian system. Isl1 in zebrafish, as in mouse, shows transient expression in mesenchyme flanking the pancreatic endoderm, and continuous expression in all endocrine cells. In isl1 mutants, endocrine cells are specified in normal numbers but more than half of these cells fail to establish expression of endocrine hormones. By using a lineage tracking approach that highlights cells leaving cell cycle early in development, we show that isl1 functions are different in first and second wave endocrine cells. In isl1 mutants, early forming first wave cells show virtually no glucagon expression and a reduced number of cells expressing insulin and somatostatin, while in the later born second wave cells somatostatin expressing cells are strongly reduced and insulin and glucagon positive cells form in normal numbers. Isl1 mutant zebrafish also display a smaller exocrine pancreas. We find that isl1 expression in the pancreatic mesenchyme overlaps with that of the related genes isl2a and isl2b and that pancreatic expression of isl-genes is independent of each other. As a combined block of two or three isl1/2 genes results in a dose-dependent reduction of exocrine tissue, our data suggest that all three genes cooperatively contribute to non-cell autonomous exocrine pancreas extension. The normal expression of the pancreas mesenchyme markers meis3, fgf10 and fgf24 in isl1/2 depleted embryos suggests that this activity is independent of isl-gene function in pancreatic mesenchyme formation as was found in mouse. This indicates species-specific differences in the requirement for isl-genes in pancreatic mesenchyme formation. Overall, our data reveal a novel interaction of isl1 and isl2 genes in exocrine pancreas expansion and cell type specific requirements during endocrine cell maturation.
| S-EPMC3657195 | biostudies-literature
Project description:Aldh1 expression is known to mark candidate progenitor populations in adult and embryonic mouse pancreas, and Aldh1 enzymatic activity has been identified as a potent regulator of pancreatic endocrine differentiation in zebrafish. However, the location and identity of Aldh1-expressing cells in zebrafish pancreas remain unknown. In this study we demonstrate that Aldh1-expressing cells are located immediately adjacent to 2F11-positive pancreatic ductal epithelial cells, and that their abundance dramatically increases during zebrafish secondary islet formation. These cells also express neurod, a marker of endocrine progenitor cells, but do not express markers of more mature endocrine cells such as pax6b or insulin. Using formal cre/lox-based lineage tracing, we further show that Aldh1-expressing pancreatic epithelial cells are the direct progeny of pancreatic notch-responsive progenitor cells, identifying them as a critical intermediate between multi-lineage progenitors and mature endocrine cells. Pharmacologic manipulation of Aldh1 enzymatic activity accelerates cell entry into the Aldh1-expressing endocrine progenitor pool, and also leads to the premature maturation of these cells, as evidenced by accelerated pax6b expression. Together, these findings suggest that Aldh1-expressing cells act as both participants and regulators of endocrine differentiation during zebrafish secondary islet formation.
| S-EPMC3798260 | biostudies-literature
Project description:Mutations in the human Shwachman-Bodian-Diamond syndrome (SBDS) gene cause defective ribosome assembly and are associated with exocrine pancreatic insufficiency, chronic neutropenia and skeletal defects. However, the mechanism underlying these phenotypes remains unclear. Here we show that knockdown of the zebrafish sbds ortholog fully recapitulates the spectrum of developmental abnormalities observed in the human syndrome, and further implicate impaired proliferation of ptf1a-expressing pancreatic progenitor cells as the basis for the observed pancreatic phenotype. It is thought that diseases of ribosome assembly share a p53-dependent mechanism. However, loss of p53 did not rescue the developmental defects associated with loss of zebrafish sbds. To clarify the molecular mechanisms underlying the observed organogenesis defects, we performed transcriptional profiling to identify candidate downstream mediators of the sbds phenotype. Among transcripts displaying differential expression, functional group analysis revealed marked enrichment of genes related to ribosome biogenesis, rRNA processing and translational initiation. Among these, ribosomal protein L3 (rpl3) and pescadillo (pes) were selected for additional analysis. Similar to knockdown of sbds, knockdown or mutation of either rpl3 or pes resulted in impaired expansion of pancreatic progenitor cells. The pancreatic phenotypes observed in rpl3- and pes-deficient embryos were also independent of p53. Together, these data suggest novel p53-independent roles for ribosomal biogenesis genes in zebrafish pancreas development.
| S-EPMC3413166 | biostudies-literature
Project description:Development of systems that reconstitute hallmark features of human pancreatic intraepithelial neoplasia (PanINs), the precursor to pancreatic ductal adenocarcinoma, could generate new strategies for early diagnosis and intervention. However, human cell-based PanIN models with defined mutations are unavailable. Here, we report that genetic modification of primary human pancreatic cells leads to development of lesions resembling native human PanINs. Primary human pancreas duct cells harbouring oncogenic KRAS and induced mutations in CDKN2A, SMAD4 and TP53 expand in vitro as epithelial spheres. After pancreatic transplantation, mutant clones form lesions histologically similar to native PanINs, including prominent stromal responses. Gene expression profiling reveals molecular similarities of mutant clones with native PanINs, and identifies potential PanIN biomarker candidates including Neuromedin U, a circulating peptide hormone. Prospective reconstitution of human PanIN development from primary cells provides experimental opportunities to investigate pancreas cancer development, progression and early-stage detection.
2017-03-14 | GSE88997 | GEO
Project description:In vitro differentiation of pluripotent cells into β cells is a promising alternative to cadaveric-islet transplantation as a cure for type 1 diabetes (T1D). During the directed differentiation of human embryonic stem cells (hESCS) by exogenous factors, numerous genes that affect the differentiation process are turned on and off autonomously. Manipulating these reactions could increase the efficiency of differentiation and provide a more complete control over the final composition of cell populations. To uncover in vitro autonomous responses, we performed single-cell RNA sequencing on hESCs as they differentiate in spherical clusters. We observed that endocrine cells and their progenitors exist beside one another in separate compartments that activate distinct genetic pathways. WNT pathway inhibition in the endocrine domain of the differentiating clusters reveals a necessary role for the WNT inhibitor APC during islet formation in vivo. Accordingly, WNT inhibition in vitro causes an increase in the proportion of differentiated endocrine cells.
| S-EPMC6933053 | biostudies-literature
Project description:It is well known that iron overload can result in pancreatic iron deposition, beta-cell destruction, and diabetes in humans. Recent studies in animals have extended the link between iron status and pancreatic function by showing that iron depletion confers protection against beta-cell dysfunction and diabetes. The aim of the present study was to identify genes in the pancreas that are differentially expressed in response to iron deficiency or overload. Weanling male Sprague-Dawley rats (n?=?6/group) were fed iron-deficient, iron-adequate, or iron-overloaded diets for 3 weeks to alter their iron status. Total RNA was isolated from the pancreases and pooled within each group for microarray analyses in which gene expression levels were compared to those in iron-adequate controls. In iron-deficient pancreas, a total of 66 genes were found to be differentially regulated (10 up, 56 down), whereas in iron-overloaded pancreas, 164 genes were affected (82 up, 82 down). The most up-regulated transcript in iron-deficient pancreas was arachidonate 15-lipoxygenase (Alox15), which has been implicated in the development of diabetes. In iron-overloaded pancreas, the most upregulated transcripts were Reg1a, Reg3a, and Reg3b belonging to the regenerating islet-derived gene (Reg) family. Reg expression has been observed in response to pancreatic stress and is thought to facilitate pancreatic regeneration. Subsequent qRT-PCR validation indicated that Alox15 mRNA levels were 4 times higher in iron-deficient than in iron-adequate pancreas and that Reg1a, Reg3a, and Reg3b mRNA levels were 17-36 times higher in iron-overloaded pancreas. The elevated Alox15 mRNA levels in iron-deficient pancreas were associated with 8-fold higher levels of Alox15 protein as indicated by Western blotting. Overall, these data raise the possibility that Reg expression may serve as a biomarker for iron-related pancreatic stress, and that iron deficiency may adversely affect the risk of developing diabetes through up-regulation of Alox15.
| S-EPMC3897611 | biostudies-literature
Project description:The tissue dynamics that govern maintenance and regeneration of the pancreas remain largely unknown. In particular, the presence and nature of a cellular hierarchy remains a topic of debate. Previous lineage tracing strategies in the pancreas relied on specific marker genes for clonal labeling, which left other populations untested and failed to account for potential widespread phenotypical plasticity. Here we employed a tracing system that depends on replication-induced clonal marks. We found that, in homeostasis, steady acinar replacement events characterize tissue dynamics, to which all acinar cells have an equal ability to contribute. Similarly, regeneration following pancreatitis was best characterized by an acinar self-replication model because no evidence of a cellular hierarchy was detected. In particular, rapid regeneration in the pancreas was found to be driven by an accelerated rate of acinar fission-like events. These results provide a comprehensive and quantitative model of cell dynamics in the exocrine pancreas.
| S-EPMC8577826 | biostudies-literature
Project description:Neurogenin 3 (Ngn3) is key for endocrine cell specification in the embryonic pancreas and induction of a neuroendocrine cell differentiation program by misexpression in adult pancreatic duct cells. We identify the gene encoding IA1, a zinc-finger transcription factor, as a direct target of Ngn3 and show that it forms a novel branch in the Ngn3-dependent endocrinogenic transcription factor network. During embryonic development of the pancreas, IA1 and Ngn3 exhibit nearly identical spatio-temporal expression patterns. However, embryos lacking Ngn3 fail to express IA1 in the pancreas. Upon ectopic expression in adult pancreatic duct cells Ngn3 binds to chromatin in the IA1 promoter region and activates transcription. Consistent with this direct effect, IA1 expression is normal in embryos mutant for NeuroD1, Arx, Pax4 and Pax6, regulators operating downstream of Ngn3. IA1 is an effector of Ngn3 function as inhibition of IA1 expression in embryonic pancreas decreases the formation of insulin- and glucagon-positive cells by 40%, while its ectopic expression amplifies neuroendocrine cell differentiation by Ngn3 in adult duct cells. IA1 is therefore a novel Ngn3-regulated factor required for normal differentiation of pancreatic endocrine cells.
| S-EPMC1422151 | biostudies-literature