Derivation of functional insulin-producing cell lines from primary mouse embryo culture

Li, Guo Dong; Luo, Ronghua; Zhang, Jiping; Yeo, Keng Suan; Xie, Fei; Tan, Eileen Khia Way; Caille, Dorothée; Que, Jianwen; Kon, Oi Lian; Salto-Tellez, Manuel; Meda, Paolo; Lim, Sai Kiang

doi:10.1016/j.scr.2008.07.004

Cited by 7 publications

(11 citation statements)

References 28 publications

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“…Although beta cells are the predominant cell type in mouse pancreatic islets, other types of cells, including alpha and delta cells, are also present. To confirm whether Smad2 and Smad3 are also differentially phosphorylated in response to activin proteins in beta cells, we investigated Smad2 and Smad3 phosphorylation in MEPI cells, an insulin-producing cell line directly derived from cultures of E6 mouse embryos [29], and in rat insulinoma INS-1, a widely used cell model for in vitro studies of GSIS [30]. In both cell types, Smad2 was phosphorylated to a higher level in response to activin A compared with activin B, while Smad3 was more strongly activated in response to activin B (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…They were then treated with activins at 100 ng/ml for 1 h in the same RPMI-based medium. MEPI and INS-1 cells were cultured as previously described [29,30]. Islets and cell monolayers were lysed in lysis buffer (50 mmol/l Tris-HCl, 0.15 mol/l NaCl, 1% Triton-X100, pH 7.4) supplemented with protease inhibitors (1 mmol/l PMSF and 1 μg/ml aprotinin) and phosphatase inhibitors (NaPPi, phosphoglycerol and NaO 4 Va).…”

Section: Animalsmentioning

confidence: 99%

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Differential regulation of mouse pancreatic islet insulin secretion and Smad proteins by activin ligands

Mezghenna

Mármol

et al. 2013

Diabetologia

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Aims/hypothesis Glucose-stimulated insulin secretion (GSIS) from pancreatic beta cells is regulated by paracrine factors, the identity and mechanisms of action of which are incompletely understood. Activins are expressed in pancreatic islets and have been implicated in the regulation of GSIS. Activins A and B signal through a common set of intracellular components, but it is unclear whether they display similar or distinct functions in glucose homeostasis. Methods We examined glucose homeostatic responses in mice lacking activin B and in pancreatic islets derived from these mutants. We compared the ability of activins A and B to regulate downstream signalling, ATP production and GSIS in islets and beta cells. Results Mice lacking activin B displayed elevated serum insulin levels and GSIS. Injection of a soluble activin B antagonist phenocopied these changes in wild-type mice. Isolated pancreatic islets from mutant mice showed enhanced GSIS, which could be rescued by exogenous activin B. Activin B negatively regulated GSIS and ATP production in wild-type islets, while activin A displayed the opposite effects. The downstream mediator Smad3 responded preferentially to activin B in pancreatic islets and beta cells, while Smad2 showed a preference for activin A, indicating distinct signalling effects of the two activins. In line with this, overexpression of Smad3, but not Smad2, decreased GSIS in pancreatic islets. Conclusions/interpretation These results reveal a tug-of-war between activin ligands in the regulation of insulin secretion by beta cells, and suggest that manipulation of activin signalling could be a useful strategy for the control of glucose homeostasis in diabetes and metabolic disease.

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Section: Resultsmentioning

confidence: 99%

Section: Animalsmentioning

confidence: 99%

Differential regulation of mouse pancreatic islet insulin secretion and Smad proteins by activin ligands

Mezghenna

Mármol

et al. 2013

Diabetologia

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“…We have previously generated more than a dozen functional insulin-producing cell lines from progenitor cells of early gastrulating mouse embryos (Li et al 2009b) and also from mouse ES cells (Li et al 2009a). Although these cell lines were derived from different precursors, they all displayed similar properties in terms of gene expression profile and responsiveness to glucose stimulation (Li et al 2009a,b, Chen et al 2010.…”

Section: Introductionmentioning

confidence: 99%

“…These stimuli trigger the closure of K ATP channels leading to the depolarization of the cell membrane and opening of the voltage-gated Ca 2C channels in these cells. In addition, these cells can be stably expanded to a large number with no loss in insulin content (Li et al 2009b). In vivo, MEPI-1 cells engraft and reverse streptozotocin (STZ)-induced hyperglycemia in SCID mice without producing teratomas (Li et al 2009b).…”

Section: Introductionmentioning

confidence: 99%

“…In addition, these cells can be stably expanded to a large number with no loss in insulin content (Li et al 2009b). In vivo, MEPI-1 cells engraft and reverse streptozotocin (STZ)-induced hyperglycemia in SCID mice without producing teratomas (Li et al 2009b). However, transplantation of these cells in STZ-induced immune-competent diabetic mice only transiently and partially corrected hyperglycemia, apparently due to acute immune rejection (unpublished data).…”

Section: Introductionmentioning

confidence: 99%

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Correction of Hyperglycemia in Type 1 Diabetic Models by Transplantation of Encapsulated Insulin-producing Cells Derived from Mouse Embryo Progenitor

Shao¹,

Gao²,

Xie³

et al. 2011

Journal of Endocrinology

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Shortage of cadaveric pancreata and requirement of immune suppression are two major obstacles in transplantation therapy of type 1 diabetes. Here, we investigate whether i.p. transplantation of alginate-encapsulated insulin-producing cells from the embryo-derived mouse embryo progenitorderived insulin-producing-1 (MEPI-1) line could lower hyperglycemia in immune-competent, allogeneic diabetic mice. Within days after transplantation, hyperglycemia was reversed followed by about 2 . 5 months of normo-to moderate hypoglycemia before relapsing. Mice transplanted with unencapsulated MEPI cells relapsed within 2 weeks. Removal of the transplanted capsules by washing of the peritoneal cavity caused an immediate relapse of hyperglycemia that could be reversed with a second transplantation. The removed capsules had fibrotic overgrowth but remained permeable to 70 kDa dextrans and displayed glucose-stimulated insulin secretion.Following transplantation, the number of cells in capsules increased initially, before decreasing to below the starting cell number at 75 days. Histological examination showed that beyond day 40 post-transplantation, encapsulated cell clusters exhibited proliferating cells with a necrotic core. Blood glucose, insulin levels, and oral glucose tolerance test in the transplanted animals correlated directly with the number of viable cells remaining in the capsules. Our study demonstrated that encapsulation could effectively protect MEPI cells from the host immune system without compromising their ability to correct hyperglycemia in immune-competent diabetic mice for 2 . 5 months, thereby providing proof that immunoisolation of expansible but immune-incompatible stem cell-derived surrogate b-cells by encapsulation is a viable diabetes therapy.

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Generating mESC-derived insulin-producing cell lines through an intermediate lineage-restricted progenitor line

Luo

Zhang

et al. 2009

Stem Cell Research

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Generating surrogate insulin-producing cells from embryonic stem cells (ESCs) through in vitro replication of successive steps during pancreatic development has been challenging . Here we describe a novel reproducible protocol to establish homogeneous and scalable insulin-producing cell lines from mouse (m) ESCs via differentiation of the previously described lineage-restricted clonal mESC-derived E-RoSH cells. Unlike their parental mESCs, E-RoSH cells expressed high levels of mesodermal and endodermal genes. Nutrient depletion in the presence of nicotinamide inhibited proliferation of E-RoSH cells and induced differentiation into heterogeneous cultures comprising vascular-like structures that produced detectable levels of insulin and C-peptide in an equimolar ratio. Limiting dilution of these cultures resulted in the isolation of eight independent insulin-producing cell lines in five experiments. All these lines were cloned and shown to be amenable to repeated cycles of freeze and thaw and to replicate for months with a doubling time of 3-4 days. Under such conditions, the cultured cells exhibited genomic, structural, biochemical, and pharmacological properties of pancreatic beta cells, including storage of an equimolar ratio of insulin and C-peptide in granules and release of the contents of these organelles through a glucose-sensitive machinery. After transplantation, these cells reversed hyperglycemia in streptozotocin-treated SCID mice and did not form teratomas.

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Derivation of functional insulin-producing cell lines from primary mouse embryo culture

Cited by 7 publications

References 28 publications

Differential regulation of mouse pancreatic islet insulin secretion and Smad proteins by activin ligands

Differential regulation of mouse pancreatic islet insulin secretion and Smad proteins by activin ligands

Correction of Hyperglycemia in Type 1 Diabetic Models by Transplantation of Encapsulated Insulin-producing Cells Derived from Mouse Embryo Progenitor

Generating mESC-derived insulin-producing cell lines through an intermediate lineage-restricted progenitor line

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