Long-Term Culture of Self-renewing Pancreatic Progenitors Derived from Human Pluripotent Stem Cells

Abstract

SummaryPluripotent stem cells have been proposed as an unlimited source of pancreatic β cells for studying and treating diabetes. However, the long, multi-step differentiation protocols used to generate functional β cells inevitably exhibit considerable variability, particularly when applied to pluripotent cells from diverse genetic backgrounds. We have developed culture conditions that support long-term self-renewal of human multipotent pancreatic progenitors, which are developmentally more proximal to the specialized cells of the adult pancreas. These cultured pancreatic progenitor (cPP) cells express key pancreatic transcription factors, including PDX1 and SOX9, and exhibit transcriptomes closely related to their in vivo counterparts. Upon exposure to differentiation cues, cPP cells give rise to pancreatic endocrine, acinar, and ductal lineages, indicating multilineage potency. Furthermore, cPP cells generate insulin+ β-like cells in vitro and in vivo, suggesting that they offer a convenient alternative to pluripotent cells as a source of adult cell types for modeling pancreatic development and diabetes.

Reconstructing human pancreatic differentiation by mapping specific cell populations during development

“…However, our attempts so far to culture so few cells sorted from human fetal pancreas have failed. Discovering new and efficient culture conditions such as co-culture on feeder layers could alleviate this issue ( Trott et al, 2017 ).…”

Current Advances and Limitations in Modeling ALS/FTD in a Dish Using Induced Pluripotent Stem Cells

“…Another concern which was raised from research with mouse iPSCs is that the selection of reprogramming factors can influence the differentiation potential of iPSCs (Buganim et al, 2014 ). The use of the standardized integration free “Sendai Reprogramming Kit” with well-characterized reprogramming factors and protocols may help to avoid these unwanted effects (Trott et al, 2017 ). In addition, a novel approach was proposed recently by using microspheres conjugated with reprogramming factors for delivery into human fibroblasts (Unciti-Broceta et al, 2015 ).…”

CENP-A overexpression drives distinct cell fates depending on p53 status

“…Multicolor fluorescence in situ hybridization (mFISH) karyotyping Cells were grown with or without 10 ng/ml Dox (as in the growth curve assay) for 15 days, then treated with colcemid (100 ng/ml, Roche) for 1.5 hours and prepared as previously described for mFISH (MetaSystems) staining (Trott et al, 2017). Briefly, mitotic cells were collected by mitotic shake-off after a short trypsin treatment and centrifuged at 1000 rpm for 10 min.…”