title: Expanding donor muscle-derived cells for transplantation.
abstract: Studies in mice showed tremendous promise for the eventual clinical utility of myoblast transplantation to treat human muscular dystrophies. Initial attempts to translate the murine studies to humans, however, were not successful, due in part to limited engraftability of expanded donor myoblasts. Conventionally, muscle cells have been cultured on collagen-coated tissue culture-treated polystyrene. However, this promotes lineage progression and differentiation of cells, which limits engraftment potential. This unit describes the isolation of canine muscle-derived cells, ex vivo expansion of cells on plates coated with a modified Notch ligand, and the xenotransplant method used to evaluate engraftment potential. Activation of Notch signaling in freshly isolated canine muscle-derived cells with Delta-1(ext)-IgG inhibits myogenic differentiation, and maintains cells earlier in myogenic lineage progression. Delta-1(ext)-IgG-expanded cells engraft into the regenerating muscle of NOD/SCID mice more effectively than control cells expanded on human IgG, as evidenced by a significant increase in the number of muscle fibers expressing canine dystrophin in recipient murine muscle. Therefore, this protocol provides the basis for further developing culture conditions for ex vivo expansion of donor muscle cells for transplant.
authors: Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, Washington, USA.
authors: Maura H
authors: MH
authors: Parker
authors: Stephen J
authors: SJ
authors: Tapscott
mesh_terms: D000818:Animals; D002452:Cell Count; D018929:Cell Culture Techniques / Q000379:methods*; D049109:Cell Proliferation; D002469:Cell Separation; D002478:Cells, Cultured; D016616:Cryoultramicrotomy; D004285:Dogs; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism; D007267:Injections; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000637:transplantation; D051379:Mice; D016513:Mice, SCID; D032342:Muscle Cells / Q000166:cytology* / Q000637:transplantation*
chemical_list: D007074:Immunoglobulin G
references: J Cell Biol. 1998 Sep 7;142(5):1257-67
references: 9732286
references: Exp Cell Res. 1959 Mar;16(3):477-505
references: 13653019
references: J Neurol Sci. 1977 Sep;33(3):425-35
references: 915527
references: J Cell Biol. 1990 Dec;111(6 Pt 1):2437-49
references: 2277066
references: Cell Tissue Res. 1999 Nov;298(2):371-5
references: 10571126
references: Exp Cell Res. 1960 Nov;21:414-20
references: 13752953
references: Nature. 1978 May 25;273(5660):306-8
references: 652035
references: Blood. 2003 Mar 1;101(5):1784-9
references: 12411302
references: J Cell Sci. 2000 Dec;113 Pt 23:4313-8
references: 11069775
references: J Morphol. 1968 Aug;125(4):447-72
references: 5687378
references: Dev Cell. 2002 Sep;3(3):397-409
references: 12361602
references: Nat Med. 2010 Feb;16(2):232-6
references: 20081862
references: Cell. 2005 Jul 29;122(2):289-301
references: 16051152
references: Dev Biol. 1986 May;115(1):129-39
references: 3516758
references: J Biophys Biochem Cytol. 1960 Oct;8:333-43
references: 13752951
references: Nature. 1989 Jan 12;337(6203):176-9
references: 2643055
references: Skelet Muscle. 2012 Feb 16;2(1):4
references: 22340947
references: Methods Enzymol. 1979;58:511-27
references: 423788
references: Stem Cells. 2012 Oct;30(10):2212-20
references: 22865615