Gene expression profiling of HUH7-ins: Lack of a granulogenic function for Chromogranin A.

Lutherborrow, Mark; Appavoo, Mathiyalagan; Simpson, Ann M.; Tuch, Bernard E.

doi:10.4161/isl.1.1.8994

Cited by 4 publications

(7 citation statements)

References 17 publications

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“…Melligen cells maintained a β-cell-like phenotype, as is seen in the Huh7ins cells, 4 , 29 by developing secretory granules of similar appearance and size to those of pancreatic β-cells together with a regulated insulin secretory mechanism. This has undoubtedly arisen from the expression of insulin against a background of endogenous expression of pancreatic cell transcription factors and β-cell proteins that are present in the dedifferentiated parental Huh7 cell line.…”

Section: Discussionmentioning

confidence: 74%

Reversal of diabetes following transplantation of an insulin-secreting human liver cell line: Melligen cells

Lawandi

Tao

Ren

et al. 2015

Molecular Therapy - Methods & Clinical Development

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As an alternative to the transplantation of islets, a human liver cell line has been genetically engineered to reverse type 1 diabetes (TID). The initial liver cell line (Huh7ins) commenced secretion of insulin in response to a glucose concentration of 2.5 mmol/l. After transfection of the Huh7ins cells with human islet glucokinase, the resultant Melligen cells secreted insulin in response to glucose within the physiological range; commencing at 4.25 mmol/l. Melligen cells exhibited increased glucokinase enzymatic activity in response to physiological glucose concentrations, as compared with Huh7ins cells. When transplanted into diabetic immunoincompetent mice, Melligen cells restored normoglycemia. Quantitative real-time polymerase chain reaction (qRT-PCR) revealed that both cell lines expressed a range of β-cell transcription factors and pancreatic hormones. Exposure of Melligen and Huh7ins cells to proinflammatory cytokines (TNF-α, IL-1β, and IFN-γ) affected neither their viability nor their ability to secrete insulin to glucose. Gene expression (microarray and qRT-PCR) analyses indicated the survival of Melligen cells in the presence of known β-cell cytotoxins was associated with the expression of NF-κB and antiapoptotic genes (such as BIRC3). This study describes the successful generation of an artificial β-cell line, which, if encapsulated to avoid allograft rejection, may offer a clinically applicable cure for T1D.

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Section: Discussionmentioning

confidence: 74%

Reversal of diabetes following transplantation of an insulin-secreting human liver cell line: Melligen cells

Lawandi

Tao

Ren

et al. 2015

Molecular Therapy - Methods & Clinical Development

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“…Further support came from complementary experiments, which showed that over-expression of chromogranin A in non-secretory cells resulted in the formation of secretory vesicles-like structures [38]. On the other hand, deletion of the chromogranin A gene did not affect the synthesis of proteins targeted to the secretory granules or their biogenesis [41] and, in other cell types, such as pancreatic β-cells, insulin vesicle biogenesis was completely independent from both chromogranin A and B [42, 43]. Moreover, chromogranin A ablation resulted in the compensatory expression of other granins.…”

Section: Regulated Exocytosismentioning

confidence: 99%

Multiple roles for the actin cytoskeleton during regulated exocytosis

Porat-Shliom

Milberg

Masedunskas

et al. 2012

Cell. Mol. Life Sci.

167

162

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Regulated exocytosis is the main mechanism utilized by specialized secretory cells to deliver molecules to the cell surface by virtue of membranous containers (i.e. secretory vesicles). The process involves a series of highly coordinated and sequential steps, which include the biogenesis of the vesicles, their delivery to the cell periphery, their fusion with the plasma membrane and the release of their content into the extracellular space. Each of these steps is regulated by the actin cytoskeleton. In this review, we summarize the current knowledge regarding the involvement of actin and its associated molecules during each of the exocytic steps in vertebrates, and suggest that the overall role of the actin cytoskeleton during regulated exocytosis is linked to the architecture and the physiology of the secretory cells under examination. Specifically, in neurons, neuroendocrine, endocrine, and hematopoietic cells, which contain small secretory vesicles that undergo rapid exocytosis (on the order of milliseconds), the actin cytoskeleton plays a role in pre-fusion events, where it acts primarily as a functional barrier and facilitates docking. In exocrine and other secretory cells, which contain large secretory vesicles that undergo slow exocytosis (seconds to minutes), the actin cytoskeleton plays a role in post-fusion events, where it regulates the dynamics of the fusion pore, facilitates the integration of the vesicles into the plasma membrane, provides structural support, and promotes the expulsion of large cargo molecules.

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“…The Huh7 parent cell line, from which the insulinsecreting Huh7ins cells were derived, represents an ideal candidate for the engineering of an artificial β-cell. These cells possess several characteristics inherent to β-cells but not intrinsic to primary hepatocytes, such as the expression of β-cell transcription factors Neurod1 [7], Pdx1, Nkx2-2, Nkx6-1, Neurog3 and Pax 6 [29]. Importantly, however, the process of transfection with insulin resulted in the formation of insulin secretory granules and the development of a regulated insulin secretory pathway [7] as was observed in the rodent H4IIEins/ND cells.…”

Section: Insulin Trafficking In a Glucose Responsive Engineered Humanmentioning

confidence: 99%

“…Importantly, however, the process of transfection with insulin resulted in the formation of insulin secretory granules and the development of a regulated insulin secretory pathway [7] as was observed in the rodent H4IIEins/ND cells. Results of a mechanistic microarray analysis comparing Huh and Huh7ins cells following insulin transfection indicated that the formation of secretory granules and the development of a regulated secretory pathway was likely related to a protein interaction or posttranslational effect in combination with increased gene expression of secretory granule proteins such as chromgranin A [29]..…”

Section: Insulin Trafficking In a Glucose Responsive Engineered Humanmentioning

confidence: 99%

“…Huh7ins cells are more akin to pancreatic β-cells than HEPG2/ins/g cells. They express a range of β-cell transcription factors [7,29] and possess storage granules that cleave proinsulin to biologically active diarginyl insulin, due to the expression of the proconvertases PC1 and PC2 [7]. As Huh7ins cells also rapidly secrete insulin in a tightly regulated manner in response to glucose, the Huh7ins cells were able to reverse chemically induced diabetes when transplanted into an animal model [7], which HEPG2ins/g cells [3] failed to achieve [Tuch, unpublished results].…”

Section: Introductionmentioning

confidence: 99%

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Insulin Trafficking in a Glucose Responsive Engineered Human Liver Cell Line is Regulated by the Interaction of ATP-Sensitive Potassium Channels and Voltage-Gated Calcium Channels

Simpson¹,

Swan²,

Liu³

et al. 2013

Gene Therapy - Tools and Potential Applications

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Gene expression profiling of HUH7-ins: Lack of a granulogenic function for Chromogranin A.

Cited by 4 publications

References 17 publications

Reversal of diabetes following transplantation of an insulin-secreting human liver cell line: Melligen cells

Reversal of diabetes following transplantation of an insulin-secreting human liver cell line: Melligen cells

Multiple roles for the actin cytoskeleton during regulated exocytosis

Insulin Trafficking in a Glucose Responsive Engineered Human Liver Cell Line is Regulated by the Interaction of ATP-Sensitive Potassium Channels and Voltage-Gated Calcium Channels

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