Impact of the two-layer method on the quality of isolated pancreatic islets.
OBJECTIVE
We have already reported that the two-layer method (UW/PFC) reduces warm and cold ischemic injuries before islet isolation, and results in improvement of islet yield and viability. In this study, we try to evaluate the effect of the two-layer method on isolated islets.


METHODS
We used male Wister rats. Isolated islets were cultured or preserved in various conditions for 24 hours. In group 1, islets were not cultured (control). In group 2, islets were cultured in RPMI at 37 degrees C. In groups 3 and 4, islets were cultured with "modified" two-layer method (RPMI/PFC) at 37 degrees C and 4 degrees C, respectively. In groups 5 and 6, islets were preserved in UW and with the two-layer method (UW/PFC), respectively at 4 degrees C. Islets in each group were evaluated in terms of function and viability in vitro.


RESULTS
Stimulation Indices were 1.3, 2.6, 3.7, 1.2, 1.4, and 2.4 in groups 1, 2, 3, 4, 5 and 6, respectively. Islets in groups 2, 3 and 6 showed clear response to glucose stimulation. Among these 3 groups, the total viability of islets assessed by FDA/PI staining was 88%, 92%, and 76% in groups 2, 3, 6, respectively.


CONCLUSIONS
Although in vivo studies are mandatory, the present study is supportive that the "modified" two-layer method (RPMI/PFC), which uses oxygenated PFC and RPMI, may be superior to conventional culture method with RPMI. This method may achieve further improvement of islet viability before implantation.
Takahashi
Tsuyoshi
T
Division of Gastroenterological Surgery, Kobe University Graduate School of Medicine Kobe, Japan.
Tanioka
Yasuki
Y
Matsuda
Takeru
T
Toyama
Hirochika
H
Kakinoki
Keitaro
K
Li
Seijitu
S
Hiraoka
Kunihiko
K
Fijino
Yasuhiro
Y
Suzuki
Yasuyuki
Y
Kuroda
Yoshikazu
Y
D000241:Adenosine; D000493:Allopurinol; D000818:Animals; D003080:Cold Temperature; D011464:Epoprostenol; D005466:Fluorocarbons; D005973:Glutamine; D005978:Glutathione; D006895:Hydroxyethyl Starch Derivatives; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000502:physiology* / Q000601:surgery; D008278:Magnesium Sulfate; D008297:Male; D009536:Niacinamide; D019188:Organ Preservation Solutions; D011887:Raffinose; D051381:Rats; D017208:Rats, Wistar; D014021:Tissue Preservation / Q000379:methods*; D014022:Tissue Survival; D020858:Tissue and Organ Harvesting; D014199:Trehalose
D005466:Fluorocarbons; D006895:Hydroxyethyl Starch Derivatives; D007328:Insulin; D019188:Organ Preservation Solutions; C431558:PFC-Kyoto solution; C055206:University of Wisconsin-lactobionate solution; D005973:Glutamine; D009536:Niacinamide; D000493:Allopurinol; D008278:Magnesium Sulfate; D014199:Trehalose; D011464:Epoprostenol; D005978:Glutathione; D000241:Adenosine; D011887:Raffinose