231-LB: Prolactin Modulates the Differentiation of Pancreatic Ductal Cell to Islet-Like Cell

Abstract

Objective: Pancreatic ductal cells will be differentiated into islet-like cell with the activation of PRL-PRLR signaling pathway in vitro. Methods: Primary mouse and human pancreatic ductal cells were obtained by CK-19 labelling and MACS sorting of digested pancreatic tissue, and selected cells were cultured with recombinant mouse prolactin and recombinant human prolactin. Subsequently, the cell clusters formed by the pancreatic ductal cells were stained by anti-insulin, anti-GCK, anti-GLUT2, anti-PC1/3 Abs. In vitro, glucose stimulated insulin secretion (GSIS) was quantified. Prolactin related signaling pathway, glucose transportation and insulin secretion related proteins were assessed by immunoblotting. Lastly, the in vitro formed islet-like cells were transplanted under the kidney capsule of diabetic mice. To assess the role for blood glucose regulation of transplanted islet-like cells, blood glucose, c-peptide and intravenous glucose tolerance tests were performed and immunofluorescence staining for insulin and glucagon was performed on the grafts. Results: At day 7, the pancreatic ductal cells, also called progenitor cells, expressed CK-19; at day 14, the ductal cells were aggregated and formed sphere-shaped cell clusters. At day 21, the cell clusters stained positive for insulin and etc. By in vitro GSIS test, we measured a 2.58±0.32-fold increase of insulin secretion in response to a 16.7 mM glucose stimulation. Immunoblotting demonstrated that PDX-1, MafA and other genes were expressed. At day 21, the cell clusters expressed PDX-1, insulin and glucagon. In vivo studies showed that diabetic mice transplanted with islet-like cells, maintained blood glucose levels below 300 mg/dL with c-peptide detected in the serum. Further tissue staining indicated that the transplanted islet-like cells contained both β-cells and α-cells. Conclusion: Out study shows that in vitro stimulation by prolactin might differentiate pancreatic ductal cells into islet-like cells. Disclosure Y. Wang: None. Funding National Natural Science Foundation of China (81802504); Sichuan Science and Technology Bureau (2022YFH0005, 2023YFH0010)