title: Hematopoietic Stem and Progenitor Cell Expansion in Contact with Mesenchymal Stromal Cells in a Hanging Drop Model Uncovers Disadvantages of 3D Culture.
abstract: Efficient ex vivo expansion of hematopoietic stem cells with a concomitant preservation of stemness and self-renewal potential is still an unresolved ambition. Increased numbers of methods approaching this issue using three-dimensional (3D) cultures were reported. Here, we describe a simplified 3D hanging drop model for the coculture of cord blood-derived CD34(+) hematopoietic stem and progenitor cells (HSPCs) with bone marrow-derived mesenchymal stromal cells (MSCs). When seeded as a mixed cell suspension, MSCs segregated into tight spheroids. Despite the high expression of niche-specific extracellular matrix components by spheroid-forming MSCs, HSPCs did not migrate into the spheroids in the initial phase of coculture, indicating strong homotypic interactions of MSCs. After one week, however, HSPC attachment increased considerably, leading to spheroid collapse as demonstrated by electron microscopy and immunofluorescence staining. In terms of HSPC proliferation, the conventional 2D coculture system was superior to the hanging drop model. Furthermore, expansion of primitive hematopoietic progenitors was more favored in 2D than in 3D, as analyzed in colony-forming assays. Conclusively, our data demonstrate that MSCs, when arranged with a spread (monolayer) shape, exhibit better HSPC supportive qualities than spheroid-forming MSCs. Therefore, 3D systems are not necessarily superior to traditional 2D culture in this regard.
authors.affiliation: Center for Medical Research, Department of Medicine II, University of Tübingen, 72072 Tübingen, Germany.
authors.forename: Olga
authors.initials: O
authors.lastname: Schmal
authors.affiliation: Institute of Applied Physics, University of Tübingen, 72076 Tübingen, Germany.
authors.forename: Jan
authors.initials: J
authors.lastname: Seifert
authors.affiliation: Institute of Applied Physics, University of Tübingen, 72076 Tübingen, Germany.
authors.forename: Tilman E
authors.initials: TE
authors.lastname: Schäffer
authors.affiliation: Department of Obstetrics and Gynecology, University of Tübingen, 72076 Tübingen, Germany.
authors.forename: Christina B
authors.initials: CB
authors.lastname: Walter
authors.affiliation: Department of Urology, University of Tübingen, 72076 Tübingen, Germany.
authors.forename: Wilhelm K
authors.initials: WK
authors.lastname: Aicher
authors.affiliation: Center for Medical Research, Department of Medicine II, University of Tübingen, 72072 Tübingen, Germany.
authors.forename: Gerd
authors.initials: G
authors.lastname: Klein
references.citation: Tissue Eng Part C Methods. 2011 Dec;17(12):1223-32
references.pmid: 21895494
references.citation: J Cell Physiol. 2009 Oct;221(1):18-25
references.pmid: 19492404
references.citation: Macromol Biosci. 2011 Jun 14;11(6):739-47
references.pmid: 21341371
references.citation: Adv Funct Mater. 2014 Apr 23;24(16):2396-2403
references.pmid: 25530746
references.citation: Nature. 2012 Jan 25;481(7382):457-62
references.pmid: 22281595
references.citation: Stem Cells. 2004;22(3):253-8
references.pmid: 15153602
references.citation: Br J Haematol. 2002 Oct;119(1):212-20
references.pmid: 12358928
references.citation: Blood. 2014 Jul 17;124(3):334-43
references.pmid: 24914138
references.citation: Nature. 2006 Feb 2;439(7076):599-603
references.pmid: 16382241
references.citation: Nat Biotechnol. 2010 Oct;28(10):1123-8
references.pmid: 20890282
references.citation: Clin Exp Ophthalmol. 2011 Nov;39(8):808-19
references.pmid: 22050567
references.citation: J Exp Med. 2011 Mar 14;208(3):421-8
references.pmid: 21402747
references.citation: PLoS One. 2010 Feb 08;5(2):e9093
references.pmid: 20161704
references.citation: Analyst. 2011 Feb 7;136(3):473-8
references.pmid: 20967331
references.citation: Blood. 1999 Aug 1;94(3):1143-5
references.pmid: 10454804
references.citation: Nature. 2003 Oct 23;425(6960):841-6
references.pmid: 14574413
references.citation: Curr Mol Med. 2012 Jan;12(1):34-49
references.pmid: 22082480
references.citation: Blood. 1998 Jan 1;91(1):111-7
references.pmid: 9414274
references.citation: J Cell Sci. 2012 Aug 15;125(Pt 16):3765-75
references.pmid: 22553208
references.citation: Nature. 2010 Aug 12;466(7308):829-34
references.pmid: 20703299
references.citation: Biomaterials. 2009 May;30(14):2705-15
references.pmid: 19215979
references.citation: J Exp Med. 2013 Jul 1;210(7):1351-67
references.pmid: 23776077
references.citation: Nat Med. 2006 Jun;12(6):657-64
references.pmid: 16715089
references.citation: PLoS One. 2014 Mar 24;9(3):e92511
references.pmid: 24663399
references.citation: Nature. 2014 Jan 16;505(7483):327-34
references.pmid: 24429631
references.citation: Stem Cell Res. 2010 Mar;4(2):129-39
references.pmid: 20116358
references.citation: Proc Natl Acad Sci U S A. 2010 Aug 3;107(31):13724-9
references.pmid: 20643923
references.citation: Proteomics. 2010 May;10(9):1886-90
references.pmid: 20162561
references.citation: Exp Hematol Oncol. 2012 May 14;1(1):12
references.pmid: 23210618
references.citation: Haematologica. 2009 Nov;94(11):1493-501
references.pmid: 19608669
references.citation: Nature. 2003 Oct 23;425(6960):836-41
references.pmid: 14574412
references.citation: N Engl J Med. 2006 Apr 27;354(17):1813-26
references.pmid: 16641398
references.citation: Nat Rev Clin Oncol. 2015 Mar;12(3):163-74
references.pmid: 25511187
references.citation: Matrix Biol. 1998 Apr;17(1):47-63
references.pmid: 9628252
references.citation: Cytotherapy. 2006;8(4):315-7
references.pmid: 16923606
references.citation: Stem Cells Dev. 2013 Dec 1;22(23):3052-61
references.pmid: 23879861
references.citation: Nat Rev Mol Cell Biol. 2011 Sep 02;12(10):643-55
references.pmid: 21886187
references.citation: J Cell Physiol. 1977 Jun;91(3):335-44
references.pmid: 301143
references.citation: Blood. 1998 Oct 15;92(8):2609-12
references.pmid: 9763542
references.citation: Matrix Biol. 1995 Feb;14(6):457-65
references.pmid: 7795884
references.citation: Nat Biotechnol. 2005 Jan;23(1):69-74
references.pmid: 15619619
references.citation: Proc Natl Acad Sci U S A. 1980 Aug;77(8):4756-9
references.pmid: 6933522
references.citation: Haematologica. 2012 May;97(5):651-60
references.pmid: 22058199
references.citation: Biomaterials. 2012 Oct;33(29):6987-97
references.pmid: 22800538
references.citation: Tissue Eng Part C Methods. 2012 May;18(5):319-28
references.pmid: 22082070
references.citation: Cell. 2008 Feb 22;132(4):631-44
references.pmid: 18295580
references.citation: Small. 2009 May;5(9):1008-13
references.pmid: 19334013
references.citation: Biotechnol Bioeng. 2012 May;109(5):1293-304
references.pmid: 22161651
references.citation: Cytotherapy. 2014 Nov;16(11):1486-500
references.pmid: 25231893
references.citation: Blood. 2003 Feb 1;101(3):877-85
references.pmid: 12393739
references.citation: Biomaterials. 2014 Jan;35(3):929-40
references.pmid: 24176196
references.citation: Biomaterials. 2011 Jan;32(2):321-9
references.pmid: 20952054
references.citation: Cell. 2007 Oct 19;131(2):324-36
references.pmid: 17956733
references.citation: Cell. 2006 Aug 25;126(4):677-89
references.pmid: 16923388
references.citation: Cell Rep. 2013 May 30;3(5):1714-24
references.pmid: 23623496
references.citation: Blood. 2000 Dec 15;96(13):4194-203
references.pmid: 11110691
references.citation: Blood. 1995 Sep 1;86(5):1740-8
references.pmid: 7655006
references.citation: Biomaterials. 2012 Feb;33(6):1736-47
references.pmid: 22136713