FETAL PIG PANCREAS PREPARATION AND ASSESSMENT OF TISSUE FOR TRANSPLANTATION, AND ITS IN VIVO DEVELOPMENT AND FUNCTION IN ATHYMIC (NUDE) MICE

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Abstract
The possibility of using xenogeneic islets for transplantation in insulin-dependent diabetes mellitus (IDDM) necessitates characterization of their potential for growth and functional differentiation. Fetal pig pancreas (FPP) of various gestational ages was examined with respect to morphology, ability to produce insulin before and during culture, and development and function in nude mice. Insulin-containing beta cells were present, but distinct islets were not apparent in FPP even in late gestation, and did not develop during culture. FPP remained viable and produced insulin for up to 30 days in vitro. Mitotic figures were seen in cultured tissue. Culture on a gelfoam raft resulted in more viable tissue than free-floating culture. Culture in a high concentration of O2 (90%O2/10% CO2) was detrimental compared with culture in 10% CO2 in air. Response to static incubation in secretagogues showed that IMBX, theophylline, and tolbutamide all stimulated insulin secretion, but high glucose concentration (5 g/L)j, arginine, and leucine did not. The potential of this tissue for growth and its ability to regulate blood glucose levels appropriately were tested in athymic (nu/nu) mice. Pancreatic tissue from fetuses as young as 4 weeks gestation showed growth after transplantation into athymic mice, with representation of the major pancreatic endocrine cells demonstrated by selective immunochemical staining. The increase in the size of the grafts showed an impressive proliferative capacity, and histology confirmed mitotic activity and islet structure in the graft. The amount of endocrine tissue in grafts reflected the condition of the explants at the time of grafting, and prolonged culture times were detrimental to eventual graft size. Functional capability of the grafted FPP to release insulin in response to hyperglycemia was tested by transplantation into mice made diabetic with streptozotocin. Blood glucose levels, animal weights and survival, and the histological appearance of the tissue after graft nephrectomy indicated that either fresh tissue or tissue cultured for up to 8 days (Gelfoam; 10% CO2 in air) had better eventual graft function then FPP grown in 90% O2 or transplanted as a secondary graft following an interim period to allow gestational maturation in a nondiabetic nu/nu host. Return to euglycemia took 3-4 months after transplantation of FPP. The in vitro characteristics of FPP are similar to those reported for human fetal tissue, and since FPP is capable of growth and proliferation in vivo and has the ability to normalize hyperglycemia, further investigation of FPP to establish its suitability as a source of xenogeneic insulin-secreting tissue for human transplantation is warranted.