Insulin-Expressing Colonies Developed From Murine Embryonic Stem Cell–Derived Progenitors (original) (raw)

Previous studies describe a unique culture method for the commitment of murine embryonic stem cells to early endocrine pancreata. In this report, early pancreatic-like β-cell progenitors were enriched and a colony assay devised to allow these progenitors to differentiate into insulin-expressing colonies in vitro. An embryonic stem cell line with enhanced green fluorescent protein (EGFP) inserted into one allele of neurogenin 3 (Ngn3), a marker for pancreatic endocrine progenitors, was differentiated. During the late stage of culture, 20–30% of cells were Ngn3-EGFP+. Gene expression profiling using the PancChip microarray platform demonstrated that Ngn3-EGFP+ cells differentially express endocrine-related genes. A novel semisolid culture method was developed to support the formation of individual insulin/C-peptide–expressing colonies from dissociated single cells. Approximately 0.1–0.6% of Ngn3-EGFP+ cells gave rise to insulin-expressing colonies, a three- to fivefold enrichment of β...