Rapamycin Suppresses the Expansion and Differentiation of Porcine Neonatal Pancreas Cell Clusters

Abstract

The role of rapamycin in pancreas stem cells remains to be clearly elucidated. Herein, we evaluated the effects of rapamycin on porcine neonatal pancreas cell clusters (NPCCs), which primarily comprised pancreatic precursors, and attempted to find an intracellular mechanism about the harmful effects of rapamycin. Porcine NPCCs were treated with rapamycin in a monolayer, and the apoptosis and proliferation were determined via caspase-3 assay and H-thymidine uptake analysis. The expression of transcription factors was assessed via reverse-transcriptase polymerase chain reaction and Western blotting. For the in vivo study, the porcine NPCCs were transplanted into the kidney subcapsules of normal nude mice and treated with rapamycin. Rapamycin treatment significantly reduced the number of β cells, glucose-stimulated insulin secretion, and the insulin contents in the monolayer-cultured porcine NPCCs. Furthermore, rapamycin treatment increased the apoptosis and inhibited the proliferation of β cells in the culture dishes. The expressions of the insulin, pancreatic and duodenal homeobox-1, and NeuroD/Beta2 genes were down-regulated via rapamycin treatment. The expression of insulin-like growth factor-II was significantly down-regulated, but the expression of Foxo1 was simultaneously inversely increased, and the translocation of Foxo1 from the cytoplasm to the nucleus was induced by rapamycin treatment. Moreover, rapamycin treatment induced a marked reduction in the relative volume and absolute mass of β cells in the porcine NPCCs grafts at 8 weeks after transplantation in the normal nude mice. Here, we demonstrate that rapamycin treatment suppresses the expansion and differentiation of porcine NPCCs, and the alteration of Foxo1 and insulin-like growth factor-II gene expression might be the crucial factors.