Enrichment of human embryonic stem cell‐derived NKX6.1‐expressing pancreatic progenitor cells accelerates the maturation of insulin‐secreting cells in vivo

• Published in: Stem cells (Dayton, Ohio) Vol. 31; no. 11; pp. 2432 - 2442
• Main Authors: Rezania, Alireza, Bruin, Jennifer E., Xu, Jean, Narayan, Kavitha, Fox, Jessica K., O'Neil, John J., Kieffer, Timothy J.
• Format: Journal Article
• Language: English
• Published: United States Oxford University Press 01.11.2013
• Subjects: Allografts; Animals; Beta cells; Cell Differentiation - physiology; Cell therapy; Diabetes; Embryonic Stem Cells - cytology; Embryonic Stem Cells - metabolism; Embryonic Stem Cells - transplantation; Endoderm - cytology; Endoderm - metabolism; Homeodomain Proteins - biosynthesis; Homeodomain Proteins - genetics; Human embryonic stem cells; Humans; Insulin; Insulin-Secreting Cells - cytology; Insulin-Secreting Cells - metabolism; Male; Medical research; Mice; Mice, SCID; Pancreas - cytology; Pancreas - metabolism; Stem cells; Transplantation; Beta cells; Human embryonic stem cells; Transplantation; Cell therapy; Diabetes
• ISSN: 1066-5099; 1549-4918
• DOI: 10.1002/stem.1489

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. Stem Cells 2013;31:2432–2442