title: Colony-forming cells in the adult mouse pancreas are expandable in Matrigel and form endocrine/acinar colonies in laminin hydrogel.
abstract: The study of hematopoietic colony-forming units using semisolid culture media has greatly advanced the knowledge of hematopoiesis. Here we report that similar methods can be used to study pancreatic colony-forming units. We have developed two pancreatic colony assays that enable quantitative and functional analyses of progenitor-like cells isolated from dissociated adult (2-4 mo old) murine pancreas. We find that a methylcellulose-based semisolid medium containing Matrigel allows growth of duct-like "Ring/Dense" colonies from a rare (∼1%) population of total pancreatic single cells. With the addition of roof plate-specific spondin 1, a wingless-int agonist, Ring/Dense colony-forming cells can be expanded more than 100,000-fold when serially dissociated and replated in the presence of Matrigel. When cells grown in Matrigel are then transferred to a Matrigel-free semisolid medium with a unique laminin-based hydrogel, some cells grow and differentiate into another type of colony, which we name "Endocrine/Acinar." These Endocrine/Acinar colonies are comprised mostly of endocrine- and acinar-like cells, as ascertained by RNA expression analysis, immunohistochemistry, and electron microscopy. Most Endocrine/Acinar colonies contain beta-like cells that secrete insulin/C-peptide in response to D-glucose and theophylline. These results demonstrate robust self-renewal and differentiation of adult Ring/Dense colony-forming units in vitro and suggest an approach to producing beta-like cells for cell replacement of type 1 diabetes. The methods described, which include microfluidic expression analysis of single cells and colonies, should also advance study of pancreas development and pancreatic progenitor cells.
authors: Department of Diabetes and Metabolic Diseases Research, Beckman Research Institute of City of Hope, Duarte, CA 91010, USA.
authors: Liang
authors: L
authors: Jin
authors: Tao
authors: T
authors: Feng
authors: Hung Ping
authors: HP
authors: Shih
authors: Ricardo
authors: R
authors: Zerda
authors: Angela
authors: A
authors: Luo
authors: Jasper
authors: J
authors: Hsu
authors: Alborz
authors: A
authors: Mahdavi
authors: Maike
authors: M
authors: Sander
authors: David A
authors: DA
authors: Tirrell
authors: Arthur D
authors: AD
authors: Riggs
authors: Hsun Teresa
authors: HT
authors: Ku
mesh_terms: D061354:Acinar Cells / Q000166:cytology / Q000378:metabolism; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000378:metabolism; D002454:Cell Differentiation; D003094:Collagen; D003114:Colony-Forming Units Assay / Q000379:methods*; D003470:Culture Media; D004338:Drug Combinations; D020100:Hydrogels; D050417:Insulin-Secreting Cells / Q000166:cytology / Q000378:metabolism; D007515:Islets of Langerhans / Q000166:cytology / Q000378:metabolism; D007797:Laminin; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D046529:Microscopy, Electron, Transmission; D009419:Nerve Tissue Proteins / Q000378:metabolism; D010179:Pancreas / Q000166:cytology* / Q000254:growth & development / Q000378:metabolism; D011509:Proteoglycans; D060449:Wnt Signaling Pathway
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D003094:Collagen; D003470:Culture Media; D004338:Drug Combinations; D020100:Hydrogels; D007797:Laminin; D009419:Nerve Tissue Proteins; D011509:Proteoglycans; C407737:Neurog3 protein, mouse; C059485:matrigel