Long-term culture of free or encapsulated islets isolated from specific pathogen-free (SPF) pigs

As the risk of recipient contamination is a limiting factor for xenotransplantation, the use of specific pathogen-free (SPF) pigs is mandatory. This study investigated the long-term culture of SPF pig islets and evaluated their insulin production when encapsulated in AN69 hollow fibres. Insulin secretion was studied after 3 weeks (n = 10), 2 months (n = 8) and 3 months (n = 3) by 45-min incubation in the presence of 2.8, 5.5, 11 and 16.5mM glucose. Although a decrease in the amount of secreted insulin occurred (1385 +/- 421 and 4323 +/- 1068 microns U/ml at 3 weeks for 2.8 and 16.5 mM glucose respectively; 702 +/- 261 and 2397 +/- 1047 microU/ml at 2 months; 59 +/- 23 and 154 +/- 34 microU/ml at 3 months), glucose-dependent insulin secretion was observed in all cases, i.e. stimulation indices of 8.1 +/- 3.1 (p < 0.05 vs the presence of 5.5 mM glucose) at 3 weeks, 3.3 +/- 1.1 at 2 months and 3.0 +/- 0.7 at 3 months. The insulin secretion of encapsulated SPF pig islets, cultured for 1 or 3 weeks, was evaluated under perifusion conditions using a stimulus of 10mM glucose plus 5.5 mM theophylline. Glucose stimulation resulted in a significant two-fold increase in insulin secretion (p < 0.05), which was maintained over culture time. These results indicate that SPF-isolated islets remained functional when cultured for several weeks either as free or encapsulated islets, although the magnitude of insulin secretion decreased dramatically after three months of culture.