21 The influence of donor age on pancreas and islet quality before and after human islet isolation.
Rebecca Vaughan1, Sarah Cross1, Anju Abraham1, Reshma Ramracheya2, Elisa Maillard-Pedracini3, Paul Johnson1
1Academic Paediatric Surgery Unit, Nuffield Department of Surgical Sciences, John Radcliffe Hospital, OXFORD, United Kingdom. 2Oxford Centre for Diabetes, Endocrinology, and Metabolism (OCDEM), Oxford, United Kingdom. 3CEED, Strasbourg, France

Abstract

Aim of Study
Type 1 diabetes affects millions of children worldwide. Transplantation / regenerative strategies hold the key to a potential cure for this debilitating disease, and is highly relevant to paediatric surgeons. Islet transplantation has been successful in selected adults, but several challenges remain before implementation in children. These include the considerable variability in islet isolation outcomes influenced by donor factors. This study aimed to characterise the influence of donor age on pancreas and islet quality. 

Methods
Human pancreases retrieved with appropriate consent/ethical approval were categorised into 2 groups: ‘young’ (age 32 (25-39); n=15) & ‘old’ (age 53 (46-60); n=15), controlling for BMI and Cold Ischaemia Time (CIT). Pancreas & isolated islet samples (pre & post 37°C culture ) were collected. Islet viability, cytotoxicity and function were assessed using FDA-EtBr-labelling, adenylate-kinase release & static GSIS. Pancreatic oxidative stress, hypoxia & apoptosis were assessed using caspase-3, TUNEL, HSP-70, lactate & SOD assays. Results are mean±SEM.

Main Results
No significant differences in islet yield (194±36 vs 229±28 IEQx103), viability (79±2 vs 75±2%), purity (71±4 vs 66±3%) or insulin content (33±6 vs 35±7 ng/islet) were found. However, superior insulin secretion was observed in 'young' islets(12.2±6.1 vs 2.6±0.5) and significantly increased cytotoxicity in 'old' cultured islets (280±72% at 72h; p<0.01). No differences were seen in active caspase-3 (0.9±0.1 vs 1.2±0.3 ng/ml), HSP-70 (19.7±10.8 vs 6.4±2.2 ng/ml), SOD (84.3±2.2 vs 92.2±13.0 % inhibition) or TUNEL positivity in pancreas (0.03±0.01 vs 0.03±0.02) or islets (0.14±0.04 vs 0.06±0.01), but 'old' pancreases had significantly higher lactate levels (1.7±0.2 vs 1.1±0.1 nmol; p<0.05). 

Conclusion
In this study, donor age had minimal impact on pancreas quality and subsequent islet yields.  However, islets from older donors demonstrated greater cytotoxicity during culture.  Higher levels of lactate found in pancreases from this donor group may be an indication of mitrochondrial dysfunction.

 


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