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The present study describes a simple and cost-effective islet isolation procedure. Using this method, allogeneic islets reverse diabetes in cynomolgus monkeys.Pancreatic tissue from 11 cynomolgus monkeys were digested, collected, and purified using a simplified method. Islet quantification, purity, viability, and glucose static incubation were conducted immediately after isolation. Five streptozotocin-induced monkeys with diabetes were transplanted intrahepatically, and liver biopsies from 3 of these monkeys were taken at different time points for histologic study.The mean (SD) of viability, purity, and static glucose incubation stimulation index were 94.4% (2.3%), 91.8% (3.4%), and 2.6 (1.7), respectively. Monkeys who received a mean (SD) dose of 19,968 (2273) islet equivalent per kilogram (n = 4) from 2 to 3 donors who achieved prolonged normoglycemia (57–232 days), whereas the single monkey who received an islet dose of 8000 islet equivalent per kilogram did not experience diabetes reversal. Immunohistochemical assessment of the liver biopsies taken from the monkeys with normoglycemia revealed an insulin- and glucagon-positive islet graft for up to 6 months with minimal peri-islet inflammatory infiltration.This study demonstrates that cynomolgus monkey islets can be successfully and efficiently harvested using a simple isolation method, and these islets can restore normoglycemia in monkeys with diabetes.