The Mesenchyme Controls the Timing of Pancreatic β-Cell Differentiation

Duvillié, Bertrand; Attali, Myriam; Bounacer, Ali; Ravassard, Philippe; Basmaciogullari, A; Scharfmann, Raphaël

doi:10.2337/diabetes.55.03.06.db05-0839

Cited by 98 publications

(76 citation statements)

References 38 publications

(38 reference statements)

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“…Such a point fits well with recent data where it was shown that knock-down of VEGF signaling in the embryonic pancreas does not seem to perturb the development of NGN3-positive endocrine progenitors but completely represses their differentiation into beta cells. 6 It is also supported by recent data indicating that while the expression of Ngn3 can be turned on in the embryonic pancreas using ␥-secretase inhibitors, insulin-positive cells did not develop under such conditions (30). Thus, Ngn3 induction is necessary but not sufficient for beta cells differentiation and the cellular context and additional signals seem necessary to allow NGN3-positive cells to develop into beta cells.…”

Section: Discussionsupporting

confidence: 69%

“…Plasmids were linearized and used as templates for synthesizing sense or antisense riboprobes using T7 or SP6 RNA polymerase (Roche Diagnostics, Meylan, France), in the presence of digoxygenin-UTP (Roche Diagnostics). In situ hybridization was done as described previously (30), and colorimetric revelation was performed with 5-bromo-4-chloro-3-indolyl phosphate (Promega, Charbonnière, France) and nitro blue tetrazolium (Roche Diagnostics) to obtain a blue precipitate. Photographs were digitized using a Hamamatsu (Middlesex, NJ) C5810 cooled 3CCD camera.…”

Section: Methodsmentioning

confidence: 99%

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Glucose Is Necessary for Embryonic Pancreatic Endocrine Cell Differentiation

Guillemain

Filhoulaud

Xavier

et al. 2007

Journal of Biological Chemistry

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Mature pancreatic cells develop during embryonic life from endodermal progenitors, and this developmental process depends on activation of a hierarchy of transcription factors. While information is available on mesodermal signals controlling pancreas development, little is known about environmental factors, such as the levels of nutrients including glucose, that may control this process. Here, we studied the effects of glucose on pancreatic cells development. We used an in vitro model where both endocrine and acinar cells develop from early pancreatic and duodenal homeobox-1 (PDX1)-positive embryonic pancreatic progenitors. We first showed that glucose does not have a major effect on global pancreatic cell proliferation, survival, and acinar cell development. On the other hand, glucose controlled both alpha and beta cell development. Specifically, the surface occupied by insulin-positive cells was 20-fold higher in pancreases cultured in presence than in absence of glucose, and this effect was dose-dependent over the range 0.5-10 mM. Glucose did not appear to control beta cell development by activating the proliferation of early progenitors or beta cells themselves but instead tightly regulated cell differentiation. Thus, glucose did not modify the pattern of expression of Neurogenin3, the earliest marker of endocrine progenitor cells, but was necessary for the expression of the transcription factor NeuroD, a direct target of Neurogenin3 known to be important for proper pancreatic endocrine cell development. We conclude that glucose interferes with the pancreatic endocrine cells development by regulating the transition between Ngn3 and upstream NeuroD.The mature mammalian pancreas contains two types of tissues: endocrine islet cells that produce hormones such as insulin (beta cells) and glucagon (alpha cells) and exocrine tissue whose acinar cells produce enzymes (e.g. amylase) that are secreted via the pancreatic ducts into the intestine. The pancreas originates from the dorsal and ventral regions of the foregut endoderm directly posterior to the stomach (1). Research conducted in the last few years has shed light on the processes controlling pancreatic endocrine-cell development. Studies of genetically engineered mice have identified a hierarchy of transcription factors regulating pancreas organogenesis and islet-cell differentiation (2, 3). The endodermal region committed to a pancreatic fate expresses the transcription factor Pancreatic and duodenal homeobox-1 (Pdx-1) (4, 5). The basic helix-loop-helix factor Neurogenin3 (Ngn3) is then expressed in epithelial pancreatic progenitor cells prior to endocrine differentiation (6). Ngn3 is necessary for pancreatic endocrine cell development, and Ngn3-deficient mice lack pancreatic endocrine cells (7). Lineage tracing experiments have also provided direct evidence that NGN3-expressing cells are islet progenitors (8). Thus, Ngn3 is a valuable marker for monitoring pancreatic endocrine-cell differentiation. NGN3 controls the expression of another member of the basic ...

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Section: Discussionsupporting

confidence: 69%

Section: Methodsmentioning

confidence: 99%

Glucose Is Necessary for Embryonic Pancreatic Endocrine Cell Differentiation

Guillemain

Filhoulaud

Xavier

et al. 2007

Journal of Biological Chemistry

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“…In earlier work, we showed that ␤-cells developed from embryonic pancreatic epithelium cultured in collagen gel without mesenchyme (20), whereas ␤-cells developed poorly when cultured with mesenchyme (21). This inhibitory effect of mesenchyme was due to delayed Ngn3 induction and required a functional Notch pathway (24). A working hypothesis was that hypoxia occurred in collagen gel, activating the Notch pathway and inhibiting endocrine differentiation, a mechanism recently established for other cell types (29).…”

Section: Discussionmentioning

confidence: 99%

“…Paraffin sections (4 m thick) or cryosections (14 m thick) were prepared. A Ngn3 probe (726 bp) was used (23) and in situ hybridization was done as previously described (24). No signal was obtained when a sense riboprobe was used.…”

Section: Methodsmentioning

confidence: 99%

Control of β-Cell Differentiation by the Pancreatic Mesenchyme

et al. 2007

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The importance of mesenchymal-epithelial interactions for normal development of the pancreas was recognized in the early 1960s, and mesenchymal signals have been shown to control the proliferation of early pancreatic progenitor cells. The mechanisms by which the mesenchyme coordinates cell proliferation and differentiation to produce the normal number of differentiated pancreatic cells are not fully understood. Here, we demonstrate that the mesenchyme positively controls the final number of ␤-cells that develop from early pancreatic progenitor cells. In vitro, the number of ␤-cells that developed from rat embryonic pancreatic epithelia was larger in cultures with mesenchyme than without mesenchyme. The effect of mesenchyme was not due to an increase in ␤-cell proliferation but was due to increased proliferation of early pancreatic duodenal homeobox-1 (PDX1)-positive progenitor cells, as confirmed by bromodeoxyuridine incorporation. Consequently, the window during which early PDX1؉ pancreatic progenitor cells differentiated into endocrine progenitor cells expressing Ngn3 was extended. Fibroblast growth factor 10 mimicked mesenchyme effects on proliferation of early PDX1؉ progenitor cells and induction of Ngn3 expression. Taken together, our results indicate that expansion of early PDX1؉ pancreatic progenitor cells represents a way to increase the final number of ␤-cells developing from early embryonic pancreas. Diabetes 56:1248-1258, 2007 E pithelium-mesenchyme interactions play a crucial role during organogenesis. They are mediated at least in part by soluble factors produced by the mesenchyme and acting on the epithelium (1). Evidence points to a crucial role for epithelialmesenchymal interactions in cell proliferation and differentiation during pancreatic development (2). However, the mechanisms by which the mesenchyme coordinates cell proliferation and differentiation to produce a normal number of differentiated pancreatic cells are not fully understood.The mature pancreas contains endocrine islets composed of cells producing hormones, such as insulin (␤-cells) and glucagon (␣-cells), and exocrine tissue composed of acinar cells producing enzymes (e.g., carboxypeptidase-A) secreted into the intestine. The pancreas originates from the dorsal and ventral regions of the foregut endoderm. Recently, important findings have shed light on the processes controlling pancreatic endocrine cell development. Studies of genetically engineered mice identified a hierarchy of transcription factors regulating pancreas organogenesis and islet-cell differentiation (3-5). The endodermal region committed to a pancreatic fate first expresses transcription factor pancreatic duodenal homeobox-1 (Pdx1). Pdx1 is detected in mouse embryos on embryonic day 8.5 (E8.5) (E9 in rats) in early pancreatic progenitors. During adulthood, Pdx1 expression becomes largely confined to ␤-cells, where it activates insulin gene transcription (6). Disruption of the Pdx1 gene in mice or human leads to pancreatic agenesis (7,8). These data indicate that P...

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“…37 The authors hypothesized that this phenomenon was due to signals from remaining mesenchyme acting downstream of Ngn3 expression to prevent beta cell differentiation. We have shown previously that mesenchymal cells do not survive culture in PEGCol hydrogels, 22 both explaining this discrepancy and highlighting a benefit of culturing within a synthetic PEG hydrogel platform.…”

Section: Mason and Mahoneymentioning

confidence: 99%

Inhibition of Gamma-Secretase Activity Promotes Differentiation of Embryonic Pancreatic Precursor Cells into Functional Islet-like Clusters in Poly(Ethylene Glycol) Hydrogel Culture

Mason

Mahoney

2010

Tissue Engineering Part A

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We assessed the ability of a gamma-secretase inhibitor to promote the in vitro differentiation of induced embryonic pancreatic precursor cell aggregates into functional islet-like clusters when encapsulated within a three-dimensional hydrogel. Undifferentiated pancreatic precursor cells were isolated from E.15 rat embryos, dissociated into single cells, and aggregated in suspension-rotation culture. Aggregates were photoencapsulated into poly(ethylene glycol) hydrogels with entrapped collagen type 1 and cultured for 14 days with or without a gamma-secretase inhibitor. Gene expression, proinsulin content, and C-peptide release were measured to determine differentiation and maturation of encapsulated precursor cell aggregates. In the control medium, scattered breakthrough beta cell differentiation was observed; however, cells remained largely insulin negative. Upon addition of a gamma-secretase inhibitor the majority of cells in clusters became insulin positive, and insulin per DNA and glucose-stimulated insulin release measurements for these cultures were comparable with those for adult rat islets. Cluster counts after culture day 14 were 88% of those initially encapsulated, demonstrating excellent cluster survival in hydrogel culture. These results indicate that concerted differentiation of pancreatic precursor cell aggregates into functionally mature islet-like clusters can be achieved in poly(ethylene glycol)-based hydrogel cultures by blocking cell contact-mediated Notch signaling with a gamma-secretase inhibitor.

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The Mesenchyme Controls the Timing of Pancreatic β-Cell Differentiation

Cited by 98 publications

References 38 publications

Glucose Is Necessary for Embryonic Pancreatic Endocrine Cell Differentiation

Glucose Is Necessary for Embryonic Pancreatic Endocrine Cell Differentiation

Control of β-Cell Differentiation by the Pancreatic Mesenchyme

Inhibition of Gamma-Secretase Activity Promotes Differentiation of Embryonic Pancreatic Precursor Cells into Functional Islet-like Clusters in Poly(Ethylene Glycol) Hydrogel Culture

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