Pulmonary Endocrine Cells In Vivo and In Vitro

Dense-core secretory granule (DCG) biogenesis is a prerequisite step for the sorting, processing, and secretion of neuropeptides and hormones in (neuro)endocrine cells. Previously, chromogranin A (CgA) has been shown to play a key role in the regulation of DCG biogenesis in vitro and in vivo. However, the underlying mechanism of CgA-mediated DCG biogenesis has not been explored. In this study, we have uncovered a novel mechanism for the regulation of CgA-mediated DCG biogenesis. Transfection of CgA into endocrine 6T3 cells lacking CgA and DCGs not only recovered DCG formation and regulated secretion but also prevented granule protein degradation. Genetic profiling of CgA-expressing 6T3 versus CgA- and DCG-deficient 6T3 cells, followed by real-time reverse transcription-polymerase chain reaction and Western blotting analyses, revealed that a serine protease inhibitor, protease nexin-1 (PN-1), was significantly up-regulated in CgA-expressing 6T3 cells. Overexpression of PN-1 in CgA-deficient 6T3 cells prevented degradation of DCG proteins at the Golgi apparatus, enhanced DCG biogenesis, and recovered regulated secretion. Moreover, depletion of PN-1 by antisense RNAs in CgA-expressing 6T3 cells resulted in the specific degradation of DCG proteins. We conclude that CgA increases DCG biogenesis in endocrine cells by up-regulating PN-1 expression to stabilize granule proteins against degradation.

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Genetic regulation of murine pituitary development

Journal of Molecular Endocrinology ◽

10.1530/jme-14-0237 ◽

2015 ◽

Vol 54 (2) ◽

pp. R55-R73 ◽

Cited By ~ 26

Author(s):

Karine Rizzoti

Keyword(s):

Endocrine Cells ◽

Molecular Mechanisms ◽

Genetic Regulation ◽

Three Dimensional ◽

Significant Progress ◽

Pituitary Development ◽

Tumour Formation ◽

Ventral Diencephalon

Significant progress has been made recently in unravelling the embryonic events leading to pituitary morphogenesis, bothin vivoandin vitro. This includes dissection of the molecular mechanisms controlling patterning of the ventral diencephalon that regulate formation of the pituitary anlagen or Rathke's pouch. There is also a better characterisation of processes that underlie maintenance of pituitary progenitors, specification of endocrine lineages and the three-dimensional organisation of newly differentiated endocrine cells. Furthermore, a population of adult pituitary stem cells (SCs), originating from embryonic progenitors, have been described and shown to have not only regenerative potential, but also the capacity to induce tumour formation. Finally, the successful recapitulationin vitroof embryonic events leading to generation of endocrine cells from embryonic SCs, and their subsequent transplantation, represents exciting advances towards the use of regenerative medicine to treat endocrine deficits. In this review, an up-to-date description of pituitary morphogenesis will be provided and discussed with particular reference to pituitary SC studies.

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Mitchell-Riley syndrome iPSCs exhibit reduced pancreatic endoderm differentiation due to a mutation in RFX6

Development ◽

10.1242/dev.194878 ◽

2020 ◽

Vol 147 (21) ◽

pp. dev194878

Author(s):

Jamie Trott ◽

Yunus Alpagu ◽

Ee Kim Tan ◽

Mohammad Shboul ◽

Yousif Dawood ◽

...

Keyword(s):

Endocrine Cells ◽

Neonatal Diabetes ◽

Pancreas Head ◽

Recessive Mutations ◽

Pancreatic Endocrine Cells ◽

Pancreas Body ◽

Pancreatic Hypoplasia

ABSTRACTMitchell-Riley syndrome (MRS) is caused by recessive mutations in the regulatory factor X6 gene (RFX6) and is characterised by pancreatic hypoplasia and neonatal diabetes. To determine why individuals with MRS specifically lack pancreatic endocrine cells, we micro-CT imaged a 12-week-old foetus homozygous for the nonsense mutation RFX6 c.1129C>T, which revealed loss of the pancreas body and tail. From this foetus, we derived iPSCs and show that differentiation of these cells in vitro proceeds normally until generation of pancreatic endoderm, which is significantly reduced. We additionally generated an RFX6HA reporter allele by gene targeting in wild-type H9 cells to precisely define RFX6 expression and in parallel performed in situ hybridisation for RFX6 in the dorsal pancreatic bud of a Carnegie stage 14 human embryo. Both in vitro and in vivo, we find that RFX6 specifically labels a subset of PDX1-expressing pancreatic endoderm. In summary, RFX6 is essential for efficient differentiation of pancreatic endoderm, and its absence in individuals with MRS specifically impairs formation of endocrine cells of the pancreas head and tail.

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TGF-beta 1 influences the relative development of the exocrine and endocrine pancreas in vitro

Development ◽

10.1242/dev.120.12.3451 ◽

1994 ◽

Vol 120 (12) ◽

pp. 3451-3462 ◽

Cited By ~ 8

Author(s):

F. Sanvito ◽

P.L. Herrera ◽

J. Huarte ◽

A. Nichols ◽

R. Montesano ◽

...

Keyword(s):

Endocrine Cells ◽

Three Dimensional ◽

Collagen Gel ◽

Tgf Beta ◽

Collagen Gels ◽

Paracrine Factors ◽

Fold Family ◽

Acinar Tissue

Pancreatic rudiments from E12.5 mouse embryos undergo extensive development and differentiation when cultured in three-dimensional gels of extracellular matrix proteins for up to 12 days. Whereas collagen gels promote the formation of numerous exocrine acini and relatively small clusters of endocrine cells, in basement membrane (EHS) matrices the development of endocrine cells is dramatically favoured over that of acinar tissue. Buds embedded in a collagen gel contiguous to an EHS gel also fail to develop acini, suggesting the involvement of diffusible factor(s). Addition of cytokines to cultures of pancreatic buds in collagen gels modifies the relative proportions of the epithelial components of the gland. In the presence of EGF the proportion of the tissue occupied by ducts overrides that of acinar structures, whereas the endocrine portion of the tissue is not significantly modified. TGF-beta 1 partially mimicks the effect of EHS matrix in inhibiting the development of acinar tissue without decreasing the amount of ducts and mesenchyme; TGF-beta 1 also promotes the development of endocrine cells, in particular of insulin-containing beta cells and of cells expressing genes of the PP-fold family. These results show that cytokines can modulate the development of the pancreas and suggest a role for TGF-beta 1 in regulating the balance between the acinar and endocrine portions of the gland in vivo. More generally, they are compatible with the notion that, during organogenesis, cytokines act as paracrine factors responsible for the development and maintenance of appropriate proportions of different tissue constituents.

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Identification of rat pancreatic duct cells by their expression of cytokeratins 7, 19, and 20 in vivo and after isolation and culture.

Journal of Histochemistry & Cytochemistry ◽

10.1177/43.3.7532655 ◽

1995 ◽

Vol 43 (3) ◽

pp. 245-253 ◽

Cited By ~ 45

Author(s):

L Bouwens ◽

F Braet ◽

H Heimberg

Keyword(s):

Endocrine Cells ◽

Pancreatic Tissue ◽

Immunocytochemical Staining ◽

Experimental Conditions ◽

Partial Pancreatectomy ◽

Tissue Sections ◽

Duct Cells ◽

Centroacinar Cells

Cells from the excretory ducts of the pancreas are thought to be capable of differentiating into exocrine and endocrine cells. To study this in rat models, markers must be found to identify the cells under different experimental conditions. We tested antibodies to different cytokeratins (CKs) by immunocytochemical staining on pancreatic tissue sections from normal rats, after partial pancreatectomy, and after isolation and culture of duct fragments. Monoclonal antibodies to human CK7, CK19, and CK20 were found to react specifically on rat pancreas tissue, as shown by Western blotting. CK20 and CK19 were immunocytochemically detected only in cells of the ductal system, from centroacinar cells to main ducts. CK7 was expressed by islets of Langerhans and by duct cells from main, inter-, and intralobular ducts, but not by centroacinar and terminal duct cells. CKs 7, 19, and 20 were also expressed in proliferating duct cells during tissue regeneration and after isolation and different periods of culture. We conclude that CKs 7, 19, and 20 are very useful markers to study the differentiation of rat duct cells under experimental conditions in vivo and in vitro.

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Insulin-Producing Endocrine Cells Differentiated In Vitro From Human Embryonic Stem Cells Function in Macroencapsulation Devices In Vivo

Stem Cells Translational Medicine ◽

10.5966/sctm.2015-0079 ◽

2015 ◽

Vol 4 (10) ◽

pp. 1214-1222 ◽

Cited By ~ 126

Author(s):

Alan D. Agulnick ◽

Dana M. Ambruzs ◽

Mark A. Moorman ◽

Anindita Bhoumik ◽

Rosemary M. Cesario ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Human Embryonic Stem Cells ◽

Endocrine Cells ◽

Embryonic Stem

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In vitro and in vivo expression of Galalpha-(1,3)Gal on porcine islet cells is age dependent

Journal of Endocrinology ◽

10.1677/joe.0.1770127 ◽

2003 ◽

Vol 177 (1) ◽

pp. 127-135 ◽

Cited By ~ 79

Author(s):

GR Rayat ◽

RV Rajotte ◽

BJ Hering ◽

TM Binette ◽

GS Korbutt

Keyword(s):

Beta Cells ◽

Endocrine Cells ◽

Islet Cells ◽

Ductal Cells ◽

Porcine Islet ◽

In Vivo Expression ◽

Neonatal Porcine Islets ◽

Mature Beta Cells

The expression of Galalpha-(1,3)Gal (alphaGal) on porcine islet cells remains controversial. Several groups have reported that porcine islet endocrine cells do not express alphaGal while we have shown in neonatal porcine islets (NPI) that beta cells do express this antigen. We hypothesize that endocrine cells expressing alphaGal on NPI are less mature cells that may have originated from ductal cells and that expression of this antigen disappears as they develop into fully mature beta cells. Thus, we further examined alphaGal expression on various porcine islet cell preparations and correlated this with the proportion of cytokeratin 7 (CK7)-positive ductal cells. In vitro and in vivo expression of alphaGal and CK7 was significantly (P<0.05) higher in less mature NPI cells compared with matured NPI and adult porcine islet cells while the reverse was observed in the proportion of beta cells. Moreover, a significantly higher proportion of CK7-positive cells was detected in the Gal-expressing population compared with non-expressing cells. In contrast, a higher proportion of beta cells was observed in the Gal-negative population compared with the Gal-positive population. These data showed a reduced expression of alphaGal and CK7 as porcine islet cells mature into beta cells suggesting a possible role for alphaGal in the maturation of pancreatic endocrine beta cells.

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Abstract 19676: A Molecular Switch Regulating Heart and Pancreatic Development

Circulation ◽

10.1161/circ.130.suppl_2.19676 ◽

2014 ◽

Vol 130 (suppl_2) ◽

Author(s):

Weijia Luo ◽

Kuo-chan Weng ◽

Wei Yu ◽

Donna Dike ◽

Yu Liu ◽

...

Keyword(s):

Cell Fate ◽

Endocrine Cells ◽

Molecular Mechanisms ◽

Ectopic Expression ◽

Regulatory Elements ◽

Initiation Site ◽

Molecular Switch ◽

Transcription Program

The mesendoderm is thought to be a population of the intermediate and transient common progenitors prior to the emergence of the me soderm (Me) and the en doderm (En) in vitro ; however, it is unclear whether it exists in vivo , and if so, what are the molecular mechanisms that establish the subsequent Me or En fate of the mesendoderm. Here we show that Mesp1 transiently marks a subset of the epiblast at the p rimitive s treak (PS) initiation site at gastrulation in vivo . It is also evidenced by the cell fate mapping that the progeny of Mesp1 + progenitors gives rise to both the Me and the definitive En, but neither the primitive En nor the ectoderm, suggesting that Mesp1 + cells are the bio-potent mesendodermal progenitors. Mesp1 -fated dorsal foregut En (derived from the definitive En) subsequently contributes exclusively to the Pdx1 + /Foxa2 + progenitors in the p ancreatic b ud (PB), and ultimately to the endocrine cells in the pancreas. RNA-seq and ch romatin- i mmuno p recipitation of Mesp1 -fated cells show that Mesp1 directly binds to the endogenous regulatory elements of mesendodermal and endodermal modulators (e.g. Foxa2 ). Conditional ablation of Foxa2 in the Mesp1 -fated cells results in aberrant PB formation and postnatal death. In situ hybridization shows that ablation of Mesp1 abolishes the majority of mesendodermal modulators while it leads to ectopic expression of endodermal transcription factors in the distal portion of the PS, indicating that transient Mesp1 expression maintains bi-potency of the mesendoderm by inhibiting the endodermal transcription program. We generated chimera embryos by injecting into WT blastocysts the Mesp1 Cre/+ ; Rosa26-EYFP (control) or the Mesp1 Cre/Cre ; Rosa26-EYFP (mutant) ES cell lines (n=3/group). We found that, the mutant ESCs fail to contribute to the cardiac mesoderm, but instead they adopt the endodermal fate evidenced by their increased contribution to the pancreatic endocrine cells, indicating that the inhibition of the endodermal transcription program by Mesp1 in the mesendoderm progenitors is a cell-autonomous effect. Our results demonstrate that Mesp1 marks the bi-potent mesendoderm in vivo and functions as a critical molecular switch between the Me and the En formation via an orchestrating dual molecular mechanism.

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