title: Enrichment of human embryonic stem cell-derived NKX6.1-expressing pancreatic progenitor cells accelerates the maturation of insulin-secreting cells in vivo.
abstract: Human embryonic stem cells (hESCs) are considered a potential alternative to cadaveric islets as a source of transplantable cells for treating patients with diabetes. We previously described a differentiation protocol to generate pancreatic progenitor cells from hESCs, composed of mainly pancreatic endoderm (PDX1/NKX6.1-positive), endocrine precursors (NKX2.2/synaptophysin-positive, hormone/NKX6.1-negative), and polyhormonal cells (insulin/glucagon-positive, NKX6.1-negative). However, the relative contributions of NKX6.1-negative versus NKX6.1-positive cell fractions to the maturation of functional β-cells remained unclear. To address this question, we generated two distinct pancreatic progenitor cell populations using modified differentiation protocols. Prior to transplant, both populations contained a high proportion of PDX1-expressing cells (~85%-90%) but were distinguished by their relatively high (~80%) or low (~25%) expression of NKX6.1. NKX6.1-high and NKX6.1-low progenitor populations were transplanted subcutaneously within macroencapsulation devices into diabetic mice. Mice transplanted with NKX6.1-low cells remained hyperglycemic throughout the 5-month post-transplant period whereas diabetes was reversed in NKX6.1-high recipients within 3 months. Fasting human C-peptide levels were similar between groups throughout the study, but only NKX6.1-high grafts displayed robust meal-, glucose- and arginine-responsive insulin secretion as early as 3 months post-transplant. NKX6.1-low recipients displayed elevated fasting glucagon levels. Theracyte devices from both groups contained almost exclusively pancreatic endocrine tissue, but NKX6.1-high grafts contained a greater proportion of insulin-positive and somatostatin-positive cells, whereas NKX6.1-low grafts contained mainly glucagon-expressing cells. Insulin-positive cells in NKX6.1-high, but not NKX6.1-low grafts expressed nuclear MAFA. Collectively, this study demonstrates that a pancreatic endoderm-enriched population can mature into highly functional β-cells with only a minor contribution from the endocrine subpopulation.
authors.affiliation: BetaLogics Venture, Janssen R & D LLC, Raritan, New Jersey, USA.
authors.forename: Alireza
authors.initials: A
authors.lastname: Rezania
authors.forename: Jennifer E
authors.initials: JE
authors.lastname: Bruin
authors.forename: Jean
authors.initials: J
authors.lastname: Xu
authors.forename: Kavitha
authors.initials: K
authors.lastname: Narayan
authors.forename: Jessica K
authors.initials: JK
authors.lastname: Fox
authors.forename: John J
authors.initials: JJ
authors.lastname: O'Neil
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Kieffer
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000502:physiology; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism* / Q000637:transplantation; D004707:Endoderm / Q000166:cytology / Q000378:metabolism; D000090322:Homeobox Protein Nkx-2.2; D018398:Homeodomain Proteins / Q000096:biosynthesis* / Q000235:genetics; D006801:Humans; D050417:Insulin-Secreting Cells / Q000166:cytology* / Q000378:metabolism; D008297:Male; D051379:Mice; D016513:Mice, SCID; D009687:Nuclear Proteins; D010179:Pancreas / Q000166:cytology* / Q000378:metabolism; D014157:Transcription Factors
chemical_list: D000090322:Homeobox Protein Nkx-2.2; D018398:Homeodomain Proteins; C000715631:NKX2-2 protein, human; C105201:NKX6-1 protein, human; C000715627:Nkx2-2 protein, mouse; D009687:Nuclear Proteins; D014157:Transcription Factors
keywords: Beta cells; Cell therapy; Diabetes; Human embryonic stem cells; Transplantation