Pancreas preservation by the 2-layer cold storage method before islet isolation protects isolated islets against apoptosis through the mitochondrial pathway.
BACKGROUND
Apoptosis in isolated islets has been implicated in primary nonfunction or early graft failure after islet transplantation. Recently, pancreas preservation by the 2-layer method (TLM) before islet isolation has been proved to improve the islet yield, quality, and transplant results not only in experimental models, but also in clinical settings. We examined the influence of TLM on apoptosis of isolated islets.


METHODS
Rat islets freshly isolated and after pancreas preservation by TLM or conventional cold storage in University of Wisconsin solution (UW) were examined and compared. Islet apoptosis was assessed by TUNEL and annexin V assays. The apoptosis pathways involved were investigated by measurement of caspase 3, 8, and 9 activities and by immunoblotting for total and phosphorylated c-Jun NH2-terminal kinase (JNK) and p38.


RESULTS
Islet apoptosis in the UW group was significantly increased compared with the fresh and TLM groups. Both caspase 3 and 9 activities in the UW group were higher than in the fresh and TLM groups with an approximate increase of 2- to 3-fold. On the other hand, there was no significant difference in caspase 8 activity among these 3 groups. JNKs were strongly activated both in the TLM and UW groups; although they were not activated in the fresh group, p38 was activated to almost the same levels in these 3 groups.


CONCLUSIONS
Pancreas preservation by TLM before islet isolation protects isolated islets against apoptosis mainly through the mitochondrial pathway. Pancreas storage before islet isolation even with TLM triggers activation of JNKs in isolated islets.
Matsuda
Takeru
T
Department of Gastroenterological Surgery, Graduate School of Medical Sciences, Kobe University, Kobe, Japan.
Suzuki
Yasuyuki
Y
Tanioka
Yasuki
Y
Toyama
Hirochika
H
Kakinoki
Keitaro
K
Hiraoka
Kunihiko
K
Fujino
Yasuhiro
Y
Kuroda
Yoshikazu
Y
D000818:Animals; D017304:Annexin A5 / Q000032:analysis; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D003080:Cold Temperature; D004789:Enzyme Activation; D020287:In Situ Nick-End Labeling; D007515:Islets of Langerhans / Q000473:pathology*; D016381:Islets of Langerhans Transplantation / Q000379:methods*; D048031:JNK Mitogen-Activated Protein Kinases; D008297:Male; D008928:Mitochondria / Q000502:physiology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009926:Organ Preservation / Q000379:methods*; D051381:Rats; D017208:Rats, Wistar; D048051:p38 Mitogen-Activated Protein Kinases
D017304:Annexin A5; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D020169:Caspases