Efficient Generation of NKX6-1+ Pancreatic Progenitors from Multiple Human Pluripotent Stem Cell Lines

Abstract

SummaryHuman pluripotent stem cells (hPSCs) represent a renewable source of pancreatic beta cells for both basic research and therapeutic applications. Given this outstanding potential, significant efforts have been made to identify the signaling pathways that regulate pancreatic development in hPSC differentiation cultures. In this study, we demonstrate that the combination of epidermal growth factor (EGF) and nicotinamide signaling induces the generation of NKX6-1+ progenitors from all hPSC lines tested. Furthermore, we show that the size of the NKX6-1+ population is regulated by the duration of treatment with retinoic acid, fibroblast growth factor 10 (FGF10), and inhibitors of bone morphogenetic protein (BMP) and hedgehog signaling pathways. When transplanted into NOD scid gamma (NSG) recipients, these progenitors differentiate to give rise to exocrine and endocrine cells, including monohormonal insulin+ cells. Together, these findings provide an efficient and reproducible strategy for generating highly enriched populations of hPSC-derived beta cell progenitors for studies aimed at further characterizing their developmental potential in vivo and deciphering the pathways that regulate their maturation in vitro.

A Novel Pre-Vascularized Subcutaneous Site Safely Accommodates Stem Cell Derived Therapies for Treating Diabetes

“…Consistent with previous studies, the PE cells we tested in our experiments require a long (more than five months) in vivo maturation period [10,17,20,21].…”

“…We speculate that longer in vivo maturation or a higher number of hESC-derived PE cells at the time of transplantation may be required to normalize glucose control in diabetic mice, although a more prominent insulin staining has been previously reported when transplanting these cells in the kidney subcapsular space and mammary fat pad [10]. Further experimentation will definitely be required to fully understand the maturation process in this new transplant site.…”

“…Nostro and G. Keller at the McEwen Centre for Regenerative Medicine in Toronto. Their differentiation protocol uses a combination of cytokines and small molecules to simulate pancreatic development and produces multipotent pancreatic progenitor cells with the potential to differentiate into all pancreatic lineages [10,11]. At the time of transplant, cells were harvested and shipped overnight to Edmonton for immediate implantation.…”

Ductal pancreatic cancer modeling and drug screening using human pluripotent stem cell– and patient-derived tumor organoids

“…Human embryonic stem cell (hESC)-derived pancreatic progenitors were generated in a monolayer format using a modification of our previously described staged differentiation protocol 30,31 . Stem cells were regularly tested to be mycoplasma free using MycoAlert (Lonza).…”

Minireview: Directed Differentiation and Encapsulation of Islet β-Cells—Recent Advances and Future Considerations

“…These studies demonstrated that the PH cells require additional transcriptional modulation to more closely resemble adult ␤-cells at both the functional and gene expression level. Additionally, Nostro et al observed that induction press.endocrine.org/journal/mend of NKX6.1 transcription factor expression may be a mediator of successful progenitor maturation in vivo, as opposed to an immature PH population lacking NKX6.1 (68). Success in generating mature ␤-cells in vitro will likely shorten the time required to prevent or reverse diabetes posttransplantation.…”