Large scale isolation, growth, and function of porcine neonatal islet cells.

Korbutt, Greg; Elliott, John F.; Ao, Ziliang; Smith, Dean K.; Warnock, Garth L.; Rajotte, Ray V.

doi:10.1172/jci118649

Cited by 355 publications

(531 citation statements)

References 45 publications

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“…The isolation method for NPIs has been described [26]. Briefly, pancreases from Landrace-Yorkshire neonatal pigs (1-3 days old, 1.5-2.0 kg body weight) were removed and digested in 2.5 mg/ml of collagenase (Sigma).…”

Section: Pigmentioning

confidence: 99%

“…The medium was replaced every second day. After this culture period, the NPIs are routinely composed of 35% endocrine and 57% ductal endocrine precursor (CK7-positive) cells [26].…”

Section: Pigmentioning

confidence: 99%

“…Cell preparations were assessed for cell number by measuring DNA content at time 0 and day 6 of culture. We have previously shown that human islet cells contain 6.6 pg DNA/cell [31] whereas NPI cells have 7.1 pg DNA/cell [26]. Cellular insulin content and cellular composition of the NPI cultures were also characterised at these timepoints.…”

Section: Cell Expansion and Characterisationmentioning

confidence: 99%

“…To determine cell number, aliquots of the cell suspension were measured in triplicate for total cellular DNA content using a fluorometric assay (Hoefer DyNa Quant 200; Amersham Pharmacia Biotech, San Francisco, CA, USA) as described previously [26]. Insulin content was measured after cellular extraction by RIA (porcine) or by ELISA (human) [26].…”

Section: Cell Characterisationmentioning

confidence: 99%

“…We have also tested the effects of sirolimus on neonatal porcine islets (NPIs). NPIs constitute a potential source of cells for clinical transplantation because of their inherent ability for proliferation and differentiation [26]. Recently, we provided the first evidence that allogeneic NPIs can reverse diabetes in a large animal model [27,28].…”

mentioning

confidence: 99%

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The impact of the mTOR inhibitor sirolimus on the proliferation and function of pancreatic islets and ductal cells

et al. 2006

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Aims/hypothesis The Edmonton Protocol for islet transplantation has provided hope for type 1 diabetic patients. However, this protocol requires lifelong immunosuppression, specifically sirolimus, a cellular antiproliferate. The effect of sirolimus on human pancreatic ductal cells (HDCs) is not known. This may be important since HDCs are believed to be islet precursors. Since neonatal porcine islets (NPIs), which contain many ductal precursor cells, could be a potential clinical source of islets, we also tested the effects of sirolimus on this tissue. Methods HDCs (n=4), NPIs (n=9) and human islets (n=5) were cultured with and without sirolimus (20 ng/ml) for 6 days. Results HDCs and NPIs cultured with sirolimus showed a 50 and 28% decrease, respectively, in cell number relative to control (p<0.05). Control cultures expanded 1.65-and 2.44-fold relative to time 0. Decreases in cell number of sirolimus-treated HDCs were not due to apoptosis as measured by TUNEL staining. No functional effects on human islets or NPIs were observed following static incubation with high glucose. Treatment of syngeneically transplanted and naïve BALC/c mice with sirolimus resulted in altered OGTT profiles with prolonged elevation of hyperglycaemia and weight gain. There was no difference in graft and organ insulin content between treatment groups. Conclusions/interpretation Our results indicate that sirolimus decreases ductal cell numbers in culture and alters glucose-stimulated insulin secretion in vivo. The administration of sirolimus to islet transplant recipients is likely to impair graft function as a result of decreasing ductal neogenesis and induction of insulin resistance.

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Section: Pigmentioning

confidence: 99%

“…The medium was replaced every second day. After this culture period, the NPIs are routinely composed of 35% endocrine and 57% ductal endocrine precursor (CK7-positive) cells [26].…”

Section: Pigmentioning

confidence: 99%

Section: Cell Expansion and Characterisationmentioning

confidence: 99%

Section: Cell Characterisationmentioning

confidence: 99%

mentioning

confidence: 99%

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The impact of the mTOR inhibitor sirolimus on the proliferation and function of pancreatic islets and ductal cells

et al. 2006

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Improved Human Pancreatic Islet Isolation for a Prospective Cohort Study of Islet Transplantation vs Best Medical Therapy in Type 1 Diabetes Mellitus

et al. 2005

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Islet Transplantation

Ryan

Lakey

Paty

et al. 2003

International Textbook of Diabetes Mellitus

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The discovery of insulin dramatically changed the outcome for patients with type 1 diabetes and the acute lethal complications of diabetes like diabetic ketoacidosis could be effectively treated. However, the improved survival allowed the development of the secondary complications of diabetes. Typically the patient with type 1 diabetes develops retinopathy, nephropathy, neuropathy, or vascular disease over time. It was not until 1993 that definitive proof for the value of good glycemic control was established. Controlling glycemia with exogenous insulin remains a challenge. Given the nonphysiological subcutaneous route of insulin administration, its inherent delay in absorption, the variability of serum levels obtained, and the systemic versus portal venous delivery, it is perhaps remarkable how well the glucose levels are actually controlled. Many efforts have been made to generate a closed‐loop system for glucose control, including the Biostator® and insulin infusion pumps. The goal of any closed‐loop system is to have reliable glucose sensing linked to appropriate insulin delivery on a continuous basis. Mechanical closed‐loop systems may in the future achieve this but have been fraught with problems despite early promise. Transplantation of islets of Langerhans or of the whole pancreas can achieve a physiological closed‐loop system today.

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Large scale isolation, growth, and function of porcine neonatal islet cells.

Cited by 355 publications

References 45 publications

The impact of the mTOR inhibitor sirolimus on the proliferation and function of pancreatic islets and ductal cells

The impact of the mTOR inhibitor sirolimus on the proliferation and function of pancreatic islets and ductal cells

Improved Human Pancreatic Islet Isolation for a Prospective Cohort Study of Islet Transplantation vs Best Medical Therapy in Type 1 Diabetes Mellitus

Islet Transplantation

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