Tissue Culture of Human Fetal Pancreas: Effects of Human Serum on Development and Endocrine Function of Isletlike Cell Clusters

The human fetal pancreas represents a source of insulin-producing β-cells with a potential for transplantation to diabetic patients. It has previously been shown that such cells can be viably maintained in tissue culture media containing fetal calf serum (FCS) and that these explants continue to synthesize and release insulin. In this study the effects of human serum (HS) on the growth and function of human fetal pancreatic explants have been compared with those of FCS. For this purpose, pancreatic glands, obtained after prostaglandin-induced abortions, were briefly exposed to collagenase, and the digest was cultured in RPMI-1640 medium plus 10% pooled HS or FCS. The outgrowth of isletlike cell clusters (ICCs) was monitored. In 31 of 58 consecutively explanted glands, development of ICCs was observed. In the presence of FCS the outgrowth of ICC took place on top of a fibroblast monocellular cell layer; HS effected less growth of fibroblasts and increased the formation of ICCs about sevenfold compared with explants from the same glands maintained in FCS. However, in the explant cultures with HS, the cell number per ICC, expressed as DNA content, was reduced by 50%. In both FCS and HS the insulin content of the medium showed great variability and progressively declined from day 2 to day 5. The medium glucagon concentration also decreased but not to the same extent as that of insulin. Immunocytochemical-stained ICCs showed insulin- and glucagon-positive cells scattered among most nonstained, presumably nonendocrine cells. The ultrastructure of the cultured cells was well preserved, with frequent granulated cells, some of which were identified as β-cells. Measurements of [³H]thymidine incorporation into ICCs indicated a high rate of DNA replication. ICCs maintained in HS showed significant rates of insulin biosynthesis and release. We conclude that in comparison with explants cultured with FCS, the yield of ICCs is markedly increased by HS. The size and geometry of these ICCs may make them particularly suited for transplantation.