In vitro Screening of Putative Compounds Inducing Fetal Porcine Pancreatic P-cell Differentiation: Implications for Cell Transplantation in Insulin-dependent Diabetes Mellitus

Abstract

Successful transplantation of fetal pancreatic p-cells to diabetic recipients requires that differentiation of the immature p-cells is achieved. Animal experiments have shown that this can occur in vivo, but it would be desirable to induce p-cell maturation in vitro prior to transplantation. For that purpose the effect of several putative inducers of p-cell differentiation and/or replication in explant cultures of fetal porcine pancreatic islet-like cell clusters (ICC) were investigated. Initial screening experiments indicated that dexamethasone (DEX; 200 ng/ml) and sodium butyrate (BUT; 2 mM) might promote p-cell differentiation as evidenced by increased insulin/DNA contents in the ICC. In subsequent experiments these two substances, and also nicotinamide (NIC; 10 mM) which previously has been found to promote fetal p-cell differentiation, were added alone or in combinations to the basal control medium consisting of RPMl 1640 + 1 % human serum. All three test agents alone or in combinations increased the insulin content/DNA of the ICC compared with that of the control group. The combination of NIC + DEX increased the insulin mRNA levels of the ICC. No significant stimulation of insulin release was observed in any test group after shortterm incubation with high glucose alone. Addition of 5 mM theophylline to high glucose stimulation, however, increased the insulin secretion in most groups of ICC. Finally, ICC in groups of about 600, which had developed in the presence of NIC or NIC + DEX, were transplanted under the kidney capsule of alloxan-diabetic nude mice. However, neither the time for reversal of diabetes (4 weeks) nor the amount of insulin secretion during perfusion from the grafted ICC were further affected by adding DEX to the NIC supplemented medium. The marked increase of the insulin content of the ICC cultured with DEX supplementation, appeared transient and was not manifested after transplantation. In conclusion, the present study demonstrated that some compounds can stimulate porcine fetal p-cells in an in vitro system, but in order to attain terminal differentiation of the p-cells including glucose-sensitive insulin secretion, longer observation periods might be required than used herein. Alternatively, an in vivo environment like that after transplantation is mandatory for this process.