Glucose-regulated insulin production in hepatocytes1

Alam, Tausif; Sollinger, Hans W.

doi:10.1097/00007890-200212270-00024

Cited by 55 publications

(45 citation statements)

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“…Our results differ from another study in which primary rat hepatocytes transduced with a different insulin transgene construct showed a significant increase in insulin secretion only 4 h after incubation in 27.5 mM glucose. 21 Although the reasons for these kinetic differences are unclear, it may be related to the known strength of the human metallothionein IIA promoter in driving expression of transgenes in liver. 22 High promoter strength is consistent with distinct cytoplasmic immunostaining for human insulin in hepatocytes electroporated with p3MTChins (Figure 4), whereas rat hepatocytes transduced with a rat albumin promoter-insulin expression construct failed to show insulin immunostaining and also responded slowly to glucose stimulation.…”

Section: Hepatocyte-mediated Therapy Of Murine Diabetes Nkf Chen Et Almentioning

confidence: 99%

Plasmid-electroporated primary hepatocytes acquire quasi-physiological secretion of human insulin and restore euglycemia in diabetic mice

et al. 2005

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We describe the durable correction of streptozotocin-induced murine diabetes by in vivo implantation of primary mouse hepatocytes electroporated ex vivo with a human proinsulin cDNA plasmid construct controlled by glucose and zinc regulatory elements. Transfected hepatocytes increased insulin transgene transcription and secretion within 10-20 min of exposure to 25 mM glucose or 60 mM zinc. Insulin release did not occur from secretory granules. Electroporated Rosa26 hepatocytes (B8 Â 10 5 viable cells) were implanted in C57BL/6J diabetic mice in one of three sites: unresected liver, regenerating liver or mesentery. Control diabetic mice were implanted with untransfected hepatocytes. At 30 days after implantation, 8/15 control mice were alive, while 19/19 treated mice were alive. The ratio of body weight on day 30/nadir body weight was significantly higher for all treated groups compared with controls. All eight surviving control mice were hyperglycemic 30 days postimplantation, while 16/19 treated diabetic mice remained normoglycemic. Treated mice had lower mean glucose values (Pp0.001) without fasting hypoglycemia and better glucose tolerance (Pp0.0003) than untreated controls. All (6/6) diabetic mice implanted in regenerating liver and 71% (5/7) implanted in unresected liver were alive 77 days after implantation. Engrafted hepatocytes were identified, mainly around central veins, by staining for b-galactosidase activity and with anti-human insulin antibody. Gene Therapy (2005) 12, 655-667.

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Section: Hepatocyte-mediated Therapy Of Murine Diabetes Nkf Chen Et Almentioning

confidence: 99%

Plasmid-electroporated primary hepatocytes acquire quasi-physiological secretion of human insulin and restore euglycemia in diabetic mice

et al. 2005

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“…This has stimulated efforts to expand existing pancreatic islets or grow new ones. As beta cells have an extremely low replication rate [1], transplantation of non-beta cells that produce insulin has become a reasonable alternative [2][3][4][5][6][7][8][9][10][11][12].…”

Section: Introductionmentioning

confidence: 99%

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

et al. 2006

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Aims/hypothesis We examined a clinical model of ex vivo transdifferentiation of primary adult hepatocytes to insulinsecreting cells for the treatment of type 1 diabetes. Materials and methods Isolated rat hepatocytes were transduced in primary culture with a human lentivirus containing pancreatic duodenal homeobox 1 (PDX1, now known as insulin promoter factor 1, homeodomain transcription factor [IPF1]). Insulin expression and secretion of the newly engineered cells were assessed in vitro by RT-PCR, in situ hybridisation, immunostaining and radioimmunoassay. PDX1-transduced hepatocytes were further studied in vivo by injecting them under the renal capsule of diabetic SCID mice. Results Isolated rat hepatocytes were efficiently transduced with the lentiviral vector, as assessed by green fluorescent reporter gene expression. The transduced cells exhibited insulin at both mRNA (RT-PCR, in situ hybridisation) and protein levels (immunostaining and radioimmunoassay). Moreover, insulin secretion by the engineered cells was dependent on glucose and sulfonylurea. Other beta cell genes, including those encoding solute carrier family 2 (facilitated glucose transporter), member 2 (Slc2a2), glucokinase (Gck), ATP-binding cassette, sub-family C (CFTR/ MRP), member 8 (Abcc8), the potassium inwardly-rectifying channel, subfamily J, member 11 (Kcnj11) and proprotein convertase subtilisin/kexin type 1 (Pcsk1) were also expressed. The PDX1-transduced hepatocytes expressed several pancreatic transcription factors related to early pancreatic endocrine development (endogenous Pdx1, 6 transdifferentiated liver cells under the renal capsule of seven streptozotocin-induced diabetic SCID mice resulted in significant reduction of nonfasting blood glucose levels from 30.7 ± 1.3 to 8.7 ± 3.7 mmol/l (mean ± SEM, p=0.01), in 6 to 8 weeks. Removal of the graft resulted in severe hyperglycaemia. Conclusions/interpretation Ex vivo lentiviral-mediated PDX1 expression in isolated adult liver cells represents a potential model for type 1 diabetes mellitus therapy.

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“…To regulate insulin transgene expression, investigators have made use of glucoseresponsive promoters such as the insulin promoter, glucose 6-phosphatase promoter, fructose 1,6-bisphosphatase promoter 11 or glucose-inducible response elements such as those based on S14 gene, a gene involved in lipogenesis that is regulated by glucose, fatty acids and thyroid hormone as well as other factors. 12 This last step of according glucose responsiveness has been the most difficult to achieve primarily because in the normal b-cell glucose-regulated insulin secretion is biphasic 13 and is attained through an immediate first phase insulin secretory response that is dependant on the release of preformed insulin secretory granules, a step that has been impossible to replicate with insulin transgene therapy, and a more sustained second phase that is regulated both at the transcriptional and post-transcriptional levels. Furthermore, insulin secretion must be rapidly turned off when blood glucose drops to prevent the occurrence of hypoglycemia.…”

Section: Prospectsmentioning

confidence: 99%

Gene Therapy Progress and Prospects: Gene therapy for diabetes mellitus

Yechoor

Chan

2004

Gene Ther

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Diabetes mellitus has long been targeted, as yet unsuccessfully, as being curable with gene therapy. The main hurdles have not only been vector-related toxicity but also the lack of physiological regulation of the expressed insulin. Recent advances in understanding the developmental biology of b-cells and the transcriptional cascade that drives it have enabled both in vivo and ex vivo gene therapy combined with cell therapy to be used in animal models of diabetes with success. The associated developments in the stem cell biology and immunology have opened up further opportunities for gene therapy to be applied to target autoimmune diabetes.

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Glucose-regulated insulin production in hepatocytes1

Abstract: These studies demonstrate in vivo glucose-regulated insulin secretion from an autologous non-beta cell leading to fasting euglycemia and an improved glucose tolerance, thereby supporting the feasibility of hepatocyte-based insulin gene-therapy for treatment of type 1 diabetes mellitus.

Cited by 55 publications

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Plasmid-electroporated primary hepatocytes acquire quasi-physiological secretion of human insulin and restore euglycemia in diabetic mice

Plasmid-electroporated primary hepatocytes acquire quasi-physiological secretion of human insulin and restore euglycemia in diabetic mice

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

Gene Therapy Progress and Prospects: Gene therapy for diabetes mellitus

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