Abstract
A method for embedded-culture of islet-like cell clusters (ICCs) from neonatal pig pancreas is described. The procedure is based on a sequential treatment of the pancreas with a proteolytic enzyme and ethylenediaminetetraacetate (EDTA). During the exposure to EDTA-Dispase, small pieces of pancreas were gradually digested, with gentle stirring. Under- and over-digested pancreatic cell clusters were separated from the pancreatic fragments by filtration, and formed whole islet-like cell clusters in RPMI 1640 containing 11.0 mM d-glucose and 10% fetal bovine serum (FBS). The ICCs were embedded in agarose with or without a gel containing a random copolymer hydroxethylmethacrylatevinylbenzyl maltonamide (HEVM) of hydroxethylmethacrylate- (HEMA)-vinylbenzyl maltonamide (VMA) and nicotinamide. They remained morphologically intact and a physiological response to acute stimulation with glucose was obtained when they were embedded in the agarose containing HEVM and nicotinamide. These findings suggest that ICCs in agarose containing HEVM and nicotinamide could be a useful tool for morphological, biochemical and molecular biological studies, and also as a potential source of material for transplantation.
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