Adult rat liver cells transdifferentiated with lentiviral IPF1 vectors reverse diabetes in mice: an ex vivo gene therapy approach

Fodor, Adriana; Harel, Chava; Fodor, Lucian; Armoni, Michal; Salmon, Patrick; Trono, Didier; Karnieli, Eddy

doi:10.1007/s00125-006-0509-8

Cited by 39 publications

(23 citation statements)

References 48 publications

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“…The target cells in these methods mainly focus on hepatic and skeletal muscle cells, but hepatic and skeletal muscle cells are not endocrinal cells, and are quite different from β cells; they usually need to be genetically modified in response to glucose and processing from pro-insulin to insulin. There is a considerable amount of endocrinal cells in gastrointestinal tracts, which are considered ideal target cells in the gene therapy of diabetes [14,15] . Considering these facts, we attempted to transfer the exogenous human insulin gene by gastrointestinal tracts.…”

Section: Introductionmentioning

confidence: 99%

Expression of human insulin gene wrapped with chitosan nanoparticles in NIH3T3 cells and diabetic rats

Niu

Xie

et al. 2008

Acta Pharmacol Sin

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Aim: To study the expression of human insulin gene wrapped with chitosan nanoparticles in NIH3T3 cells and diabetic rats. Methods: pCMV.Ins, an expression plasmid of the human insulin gene, was constructed. In total, 100 μg pCMV.Ins wrapped with chitosan nanoparticles (chitosan-pCMV.Ins) was transfected to NIH3T3 cells and diabetes rats through lavage and coloclysis, respectively. The transfected cells were grown in Dulbecco's modified Eagle's medium, containing G418, for 72 h after transfection. The clones were selected and continued to grow in G418 medium for 24 d. The expression of human insulin was detected by immunohistochemistry. Human insulin in the culture medium of transfected cells was measured. Fasting blood glucose and plasma human insulin of diabetic rats were measured for 5 d after transfection. RT-PCR and Western blotting were performed to confirm the expression of the human insulin gene in diabetic rats. Results: Approximately 10% of NIH3T3 cells transfected by chitosan-pCMV.Ins expressed human insulin. Human insulin in the culture medium of NIH3T3 cells transfected by chitosan-pCMV.Ins significantly increased compared with that of the control group (P<0.01). Fasting blood glucose levels of the lavage group and the coloclysis group decreased significantly in 5 d (P<0.01) in comparison, while plasma insulin levels were much higher (P<0.01). The human insulin gene mRNA and human insulin were only detected in the lavage and the coloclysis groups. Conclusion: The human insulin gene can be transfected and expressed successfully by chitosanpCMV.Ins in NIH3T3 cells and diabetes rats, which indicates that chitosan is a promising, non-viral vector for gene expression.

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Section: Introductionmentioning

confidence: 99%

Expression of human insulin gene wrapped with chitosan nanoparticles in NIH3T3 cells and diabetic rats

Niu

Xie

et al. 2008

Acta Pharmacol Sin

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“…Our work provides potential novel regulatory elements that should be included when screening for mutations that affect HNF1␣ expression. NKX6.1 is uniquely expressed in mature beta cells of the pancreas (32) but is not expressed in the liver (57). This novel regulatory network may provide potential new targets for diagnosis and MODY and possibly implicate a new gene (NKX6.1) involved in this disease.…”

Section: Nkx61 Regulates Hnf1␣ In Beta Cellsmentioning

confidence: 99%

Distinct Regulation of Hepatic Nuclear Factor 1α by NKX6.1 in Pancreatic Beta Cells

Donelan

Koya

et al. 2010

Journal of Biological Chemistry

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Hepatic nuclear factor 1␣ (HNF1␣) is a key regulator of development and function in pancreatic beta cells and is specifically involved in regulation of glycolysis and glucose-stimulated insulin secretion. Abnormal expression of HNF1␣ leads to development of MODY3 (maturity-onset diabetes of the young 3). We report that NK6 homeodomain 1 (NKX6.1) binds to a cis-regulatory element in the HNF1␣ promoter and is a major regulator of this gene in beta cells. We identified an NKX6.1 recognition sequence in the distal region of the HNF1␣ promoter and demonstrated specific binding of NKX6.1 in beta cells by electrophoretic mobility shift and chromatin immunoprecipitation assays. Site-directed mutagenesis of the NKX6.1 core-binding sequence eliminated NKX6.1-mediated activation and substantially decreased activity of the HNF1␣ promoter in beta cells. Overexpression or small interfering RNA-mediated knockdown of the Nkx6.1 gene resulted in increased or diminished HNF1␣ gene expression, respectively, in beta cells. We conclude that NKX6.1 is a novel regulator of HNF1␣ in pancreatic beta cells. This novel regulatory mechanism for HNF1␣ in beta cells may provide new molecular targets for the diagnosis of MODY3. Hepatic nuclear factor 1␣ (HNF1␣)2 is a key transcription factor involved in glucose-stimulated insulin secretion (GSIS) and is the major factor involved in most cases of maturity-onset diabetes of the young (MODY), which accounts for ϳ1% of worldwide diabetes cases (1, 2). Precise cellular concentrations of HNF1␣ are required to maintain normal beta cell function, and both underexpression and overexpression have been shown to lead to diabetes (3-9).HNF1␣ is an important gene involved in the developmental regulation of the liver, pancreas, kidneys, stomach, and intestines (10 -14). This transcription factor is essential for control of mature cellular phenotype in these tissues. In pancreatic beta cells, HNF1␣ is involved in regulating the transcription of several genes that are involved in glycolysis and GSIS such as insulin (15), the Glut2 glucose transporter (16), pyruvate kinase (17), aldolase B (18), HNF3␥ (19), and HNF4␥ (19). Although the regulation of HNF1␣ has been well characterized in hepatocytes, its regulation in beta cells has not been studied in detail, and this gene is assumed to be regulated based on mechanistic studies from hepatocytes (20).HNF1␣ has an HNF4␣-binding site in its proximal promoter that has been shown to be sufficient for its activation in hepatocytes (21, 22), but evidence exists to suggest that the mechanism for regulation of HNF1␣ may different in beta cells. First, HNF1␣ is expressed as three isoforms (A, B, and C) that have tissue-specific distribution ratios (14). HNF1␣A is the predominant form in hepatocytes, whereas HNF1␣B is predominant in pancreatic islets. It has been shown that HNF1␣B and HNF1␣C have greater transactivation potential than HNF1␣A (23). Second, the major regulator of this gene, HNF4␣, is predominantly expressed from the P1 promoter in hepatocytes, whereas in th...

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“…Ferber et al [2000] first reported that transient adenovirus-mediated expression of pancreatic and duodenal homeobox gene 1 (Pdx-1) in hepatocytes activated the expression of endogenous insulin and ameliorated hyperglycemia in diabetic mice treated with streptozotocin (STZ) [Ber et al, 2003]. Recently, reprogramming of a range of liver cell types into pancreatic cells in vitro by forced expression of Pdx-1 has also been reported [Yang et al, 2002;Zalzman et al, 2003Zalzman et al, , 2005Cao et al, 2004;Li et al, 2005;Sapir et al, 2005;Tang et al, 2006a,b;Fodor et al, 2007].…”

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confidence: 92%

Conversion of immortal liver progenitor cells into pancreatic endocrine progenitor cells by persistent expression of Pdx‐1

Jin

et al. 2007

J of Cellular Biochemistry

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The conversion of expandable liver progenitor cells into pancreatic beta cells would provide a renewable cell source for diabetes cell therapy. Previously, we reported the establishment of liver epithelial progenitor cells (LEPCs). In this work, LEPCs were modified into EGFP/Pdx-1 LEPCs, cells with stable expression of both Pdx-1 and EGFP. Unlike previous work, with persistent expression of Pdx-1, EGFP/Pdx-1 LEPCs acquired the phenotype of pancreatic endocrine progenitor cells rather than giving rise to insulin-producing cells directly. EGFP/Pdx-1 LEPCs proliferated vigorously and expressed the crucial transcription factors involved in beta cell development, including Ngn3, NeuroD, Nkx2.2, Nkx6.1, Pax4, Pax6, Isl1, MafA and endogenous Pdx-1, but did not secrete insulin. When cultured in high glucose/low serum medium supplemented with cytokines, EGFP/Pdx-1 LEPCs stopped proliferating and gave rise to functional beta cells without any evidence of exocrine or other islet cell lineage differentiation. When transplanted into diabetic SCID mice, EGFP/Pdx-1 LEPCs ameliorated hyperglycemia by secreting insulin in a glucose regulated manner. Considering the limited availability of beta cells, we propose that our experiments will provide a framework for utilizing the immortal liver progenitor cells as a renewable cell source for the generation of functional pancreatic beta cells.

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Adult rat liver cells transdifferentiated with lentiviral IPF1 vectors reverse diabetes in mice: an ex vivo gene therapy approach

Cited by 39 publications

References 48 publications

Expression of human insulin gene wrapped with chitosan nanoparticles in NIH3T3 cells and diabetic rats

Expression of human insulin gene wrapped with chitosan nanoparticles in NIH3T3 cells and diabetic rats

Distinct Regulation of Hepatic Nuclear Factor 1α by NKX6.1 in Pancreatic Beta Cells

Conversion of immortal liver progenitor cells into pancreatic endocrine progenitor cells by persistent expression of Pdx‐1

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