Optimized protocol for generating functional pancreatic insulin-secreting cells from human pluripotent stem cells

• Main Authors: Cherkaoui, Ines, Du, Qian, Egli, Dieter, Misra, Shivani, Rutter, Guy A.
• Other Authors: Lee Kong Chian School of Medicine (LKCMedicine)
• Format: Article
• Language: English
• Published: 2024
• Subjects: Medicine, Health and Life Sciences; Endocrine pancreas; Human embryonic stem cell
• Online Access: https://hdl.handle.net/10356/178514
• Institution: Nanyang Technological University

Human pluripotent stem cells (hPSCs) can differentiate into any kind of cell, making them an excellent alternative source of human pancreatic β-cells. hPSCs can either be embryonic stem cells (hESCs) derived from the blastocyst or induced pluripotent cells (hiPSCs) generated directly from somatic cells using a reprogramming process. Here a video-based protocol is presented to outline the optimal culture and passage conditions for hPSCs, prior to their differentiation and subsequent generation of insulin-producing pancreatic cells. This methodology follows the six-stage process for β-cell directed differentiation, wherein hPSCs differentiate into definitive endoderm (DE), primitive gut tube, posterior foregut fate, pancreatic progenitors, pancreatic endocrine progenitors, and ultimately pancreatic β-cells. It is noteworthy that this differentiation methodology takes a period of 27 days to generate human pancreatic β-cells. The potential of insulin secretion was evaluated through two experiments, which included immunostaining and glucose-stimulated insulin secretion.