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Abstract
We previously reported that dye was effective to prevent the leakage of enzyme solutions from pancreatic glands during an islet isolation procedure. However, the dye used for islet isolation has not yet been optimized. In this study, we focused on pyoktanin blue (PB), diagnogreen (DG), and indigo carmine (IC) as potential candidates among clinically established dyes. A serial dilution assay was performed to determine minimal effective concentrations of each dye for detecting damaged pancreatic tissues. According to the outcome of serial dilution assays, double minimum effective concentrations of each dye were used for in vitro toxicity assays on islets and used in the isolation procedure to investigate whether they adversely affect islet isolation efficiency. The evaluations included islet yield, ADP/ATP, ATP/DNA, glucose stimulation test, and insulin/DNA assays. Islet viability cultured with PB contained medium was significantly lower than the other dyes. DG and IC appeared to be non-toxic to the islets. In isolation experiments, the islet yield in the DG group was considerably lower than that in the Control group, suggesting that DG might inhibit enzyme activity. The present study demonstrates that IC could be a promising candidate for an effective dye to detect damaged pancreatic tissues without affecting the enzyme activity and islet quality.
Highlights
Pancreatic islet transplantation is an attractive and promising treatment for type 1 diabetic patients
A serial dilution assay revealed that the minimum effective concentrations of each dye for detecting damaged pancreatic tissues were 2.1 μg/ml (PB), 27.8 μg/ml (DG), and 6.7 μg/ml (IC), respectively (Fig 1)
The adenosine diphosphate (ADP)/adenosine triphosphate (ATP) ratio was significantly higher in the pyoktanin blue (PB) group than the other groups (Control: 0.00, PB: 0.59±0.04, DG: 0.00, and indigo carmine (IC): 0.00, n = 5, p
Summary
Pancreatic islet transplantation is an attractive and promising treatment for type 1 diabetic patients. A low yield of isolated islets is caused by various factors, inadequate delivery of dissociation enzymes to the pancreatic parenchyma due to tissue damages during organ harvesting procedures could be a primary issue [3, 4]. We previously reported that both dye and glue were effective to prevent the leakage of enzyme solutions from pancreatic glands [3]. These methods are especially useful for the cases of islet autotransplantation associated with severe inflammation caused by pancreatic arteriovenous malformation [5, 6]. Methylthionine chloride was proposed as a potential dye solution to detect the damaged area of the pancreas [3].
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