Serine-Protease Inhibition During Islet Isolation Increases Islet Yield From Human Pancreases With Prolonged Ischemia1

Lakey,; Lm, Helms; Kin, Tatsuya; Gs, Korbutt; Rv, Rajotte; Am, Shapiro; Gl, Warnock

doi:10.1097/00007890-200108270-00003

Cited by 78 publications

(65 citation statements)

References 16 publications

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“…Spectrophotometric analysis was then performed to measure the absorbance of NADH, for processing. In this study, human pancreata were selected with the criteria of having more than 8 h (8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19) providing quantification of ATP, ADP, and AMP. Values are reported per gram of wet weight.…”

Section: Methodsmentioning

confidence: 99%

Short-Term Storage of the Ischemically Damaged Human Pancreas by the Two-Layer Method Prior to Islet Isolation

et al. 2004

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A two-layer cold storage method (TLM) allows sufficient oxygen delivery to pancreata during preservation and resuscitates the viability of ischemically damaged pancreata in the canine pancreas transplant model. In this study, we applied a short-term preservation of the TLM to human pancreata after prolonged cold ischemia prior to islet isolation, and investigated the mechanisms of resuscitation of the ischemically damaged human pancreas by the TLM. Human pancreata were procured from cadaveric donors and preserved by the TLM for 3.2 ± 0.5 h after 11.1 ± 0.9 h of cold storage in UW (TLM group), or by cold UW alone for 11.0 ± 0.3 h (UW group). Islet isolations of all pancreata were performed using the Edmonton protocol. Islet recovery and in vitro functional viability of isolated islets were significantly increased in the TLM group compared with the UW group. According to the criteria of the Edmonton protocol, 10/14 cases (71%) in the TLM group were transplanted to patients with type I diabetes mellitus compared with only 5/21 cases (24%) in the UW group. In the metabolic assessment of human pancreata, levels of energetic parameters (ATP, total adenylates, and energy charge) were significantly increased, and malondialdehyde (MDA) levels were significantly decreased after the TLM preservation. There was no observable change in the incidence or degree of mitochondrial injury after the TLM preservation. Additional short-term storage by the TLM resuscitates the ischemically damaged human pancreas by regenerating the energetic status and prevents further damage by oxidative stress, ultimately leading to improvements of islet recovery and in vitro function. Use of the TLM following prolonged storage in UW provides an excellent adjunctive protocol for treating human pancreata for the rigors of the islet isolation process.

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Section: Methodsmentioning

confidence: 99%

Short-Term Storage of the Ischemically Damaged Human Pancreas by the Two-Layer Method Prior to Islet Isolation

et al. 2004

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“…The aim of the present study is to compare the use of the two commercially available, clinical grade iron oxUnlike for other types of organ transplantation, tests allowing the detection of early rejection after islet transide nanoparticles, ferucarbotran (Resovist) and ferumoxide (Feridex), in terms of iron uptake, toxicity, inplantation are lacking (9,19). Alteration of glycemic control occurs too late in the graft destruction process to sulin response, and MRI imaging ability, both in vitro and in vivo.…”

Section: Introductionmentioning

confidence: 99%

Assessment of Human Islet Labeling with Clinical Grade Iron Nanoparticles Prior to Transplantation for Graft Monitoring by MRI

et al. 2010

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Ex vivo labeling of islets with superparamagnetic iron oxide (SPIO) nanoparticles allows posttransplant MRI imaging of the graft. In the present study, we compare two clinical grade SPIOs (ferucarbotran and ferumoxide) in terms of toxicity, islet cellular uptake, and MRI imaging. Human islets (80-90% purity) were incubated for 24 h with various concentrations of SPIOs (14-280 µg/ml of iron). Static incubations were performed, comparing insulin response to basal (2.8 mM) or high glucose stimulation (16.7 mM), with or without cAMP stimulation. Insulin and Perl's (assessment of iron content) staining were performed. Electronic microscopy analysis was performed. Labeled islets were used for in vitro or in vivo imaging in MRI 1.5T. Liver section after organ removal was performed in the same plane as MRI imaging to get a correlation between histology and radiology. Postlabeling islet viability (80 ± 10%) and function (in vitro static incubation and in vivo engraftment of human islets in nude mice) were similar in both groups. Iron uptake assessed by electron microscopy showed iron inclusions within the islets with ferucarbotran, but not with ferumoxide. MRI imaging (1.5T) of phantoms and of human islets transplanted in rats, demonstrated a strong signal with ferucarbotran, but only a weak signal with ferumoxide. Signal persisted for >8 weeks in the absence of rejection. An excellent correlation was observed between radiologic images and histology. The hepatic clearance of intraportally injected ferucarbotran was faster than that of ferumoxide, generating less background. A rapid signal decrease was observed in rejecting xenogeneic islets. According to the present data, ferucarbotran is the most appropriate of available clinical grade SPIOs for human islet imaging.Key words: Islet imaging; Islet transplantation; Iron nanoparticles; Magnetic resonance; Imaging INTRODUCTIONone of the most promising strategies (11,12,23,(29)(30)(31).The aim of the present study is to compare the use of the two commercially available, clinical grade iron oxUnlike for other types of organ transplantation, tests allowing the detection of early rejection after islet transide nanoparticles, ferucarbotran (Resovist) and ferumoxide (Feridex), in terms of iron uptake, toxicity, inplantation are lacking (9,19). Alteration of glycemic control occurs too late in the graft destruction process to sulin response, and MRI imaging ability, both in vitro and in vivo. enable graft salvage and islet graft biopsy (i.e., liver biopsy) is not sensitive enough, due to sampling issues MATERIALS AND METHODS (3,28). New strategies are currently under development, Animal Selection including immune monitoring, molecular monitoring (2), and islet graft imaging. Magnetic resonance imaging Male Lewis rats (Janvier, Le Genest, France; 300-500 g) and athymic nu/nu (nude) mice (Janvier, Le Gen-(MRI) after ex vivo islet labeling currently appears to be est, France) were used. All experiments were performed CMRL 1066-based medium, as described above, with SPIOs at iron co...

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“…Briefly, human pancreata were perfused of TLM storage upon arrival at the islet isolation laborawith Liberase-HI purified enzyme blend and 0.4 mM tory after cold storage in UW solution (7-11 h) during Pefabloc (Roche Molecular Biochemicals, Summerville, transportation. Additional preservation by the TLM was NJ) under controlled conditions for 10 min (20). Panrandomly assigned before islet isolations.…”

Section: Introductionmentioning

confidence: 99%

Ameliorating Injury during Preservation and Isolation of Human Islets Using the Two-Layer Method with Perfluorocarbon and UW Solution

et al. 2006

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This study assessed the effects of a two-layer method (TLM), using perfluorocarbon and UW solution, on the quality of human pancreata following storage and islet yield/function after isolation. In part A, TLM was applied immediately after procurement and the energetic profile was compared to a group treated with UW solution only (control) throughout 24-h storage. In part B, cadaveric human pancreata were procured and subjected to a TLM after cold storage in UW solution (TLM group) or UW solution (control group). Energetics, lipid peroxidation, and islet recovery/function were assessed after preservation at 4°C. In part A, after 9-h storage, the energetic profile (ATP, ATP/ADP, energy charge) for the TLM group was superior to controls. In part B, TLM treatment resulted in consistently greater ATP, ATP/ADP, and energy charge values than with storage in UW solution alone (p < 0.05). UW treatment resulted in 40% greater peroxidative damage than in the TLM group (p < 0.05). Islet recovery and functional viability were 30-40% higher following TLM treatment (p < 0.05). These data support the hypothesis that islet viability and yields can be significantly improved using a brief period of TLM treatment following conventional UW storage; reduced energetic and oxidative stress are implicated as potential mechanisms.

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Serine-Protease Inhibition During Islet Isolation Increases Islet Yield From Human Pancreases With Prolonged Ischemia1

Abstract: Inhibition of intrinsic protease activity within pancreases after prolonged cold storage improves isolation of viable islets.

Cited by 78 publications

References 16 publications

Short-Term Storage of the Ischemically Damaged Human Pancreas by the Two-Layer Method Prior to Islet Isolation

Short-Term Storage of the Ischemically Damaged Human Pancreas by the Two-Layer Method Prior to Islet Isolation

Assessment of Human Islet Labeling with Clinical Grade Iron Nanoparticles Prior to Transplantation for Graft Monitoring by MRI

Ameliorating Injury during Preservation and Isolation of Human Islets Using the Two-Layer Method with Perfluorocarbon and UW Solution

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