title: A modified aggregate culture for chondrogenesis of human adipose-derived stem cells genetically modified with growth and differentiation factor 5.
abstract: Adipose-derived stem cells (ADSCs) are an attractive cell source for tissue engineering, and recently a modified aggregate culture of human ADSCs (hADSCs) was established based on preparation of three-dimensional (3D) cell aggregates in growth factor-enriched low serum medium using the hanging droplet method. Growth and differentiation factor 5 (GDF5) plays a critical role in chondrogenesis and cartilage development. In the present study, we examine (1) whether the modified aggregate culture is feasible for chondrogenic induction of hADSCs, (2) whether overexpressed GDF5 can promote chondrogenesis, and (3) the gene expression profile during chondrogenesis in this aggregate culture. hADSCs were infected with an adenovirus carrying the GDF5 gene (Ad-GDF5). Cells were cultured with chondrogenic media either in a modified aggregate culture or in an attached micromass culture that served as a control. The chondrogenic phenotype was assessed by morphology (n=8), biochemistry (n=3), and histology (n=2). Expression of 12 genes was determined by quantitative real-time polymerase chain reaction (n=3). We found that ADSCs cultured in the modified aggregates exhibited denser pellets and higher content of sulfated glycosaminoglycan (sGAG) compared with those cultured in the micromass. Infection of cells with Ad-GDF5 increased the aggregate size and sGAG content. It also up-regulated expression of GDF5, aggrecan, and leptin and down-regulated expression of COL I, while expression of COL II and COL 10 remained unchanged. We concluded that the modified aggregate culture is feasible for chondrogenic induction of human ADSCs. Infection with Ad-GDF5 appears to promote the chondrogenesis. These findings suggest that genetic modification of ADSCs with GDF5 in the modified aggregate culture could be useful for treating diseases with cartilage defects.
authors.affiliation: Orthopedic Research Laboratory, Department of Orthopedic Surgery, University of Virginia , Charlottesville, Virginia.
authors.forename: Xinlin
authors.initials: X
authors.lastname: Yang
authors.forename: Hulan
authors.initials: H
authors.lastname: Shang
authors.forename: Adam
authors.initials: A
authors.lastname: Katz
authors.forename: Xudong
authors.initials: X
authors.lastname: Li
keywords: GDF5; aggregate culture; cartilage regeneration; chondrogenesis; genetic modification; human adipose-derived stem cell
references.citation: Osteoarthritis Cartilage. 2002 May;10(5):394-401
references.pmid: 12027540
references.citation: Cytotherapy. 2007;9(7):637-46
references.pmid: 17917877
references.citation: Eur Cell Mater. 2011 Jun 20;21:533-47
references.pmid: 21710444
references.citation: Biomaterials. 2010 May;31(13):3564-71
references.pmid: 20153525
references.citation: Connect Tissue Res. 2011;52(4):353-64
references.pmid: 21250863
references.citation: Cytotechnology. 2006 May;51(1):1-5
references.pmid: 19002889
references.citation: Mol Ther. 2012 Jul;20(7):1298-304
references.pmid: 22525514
references.citation: J Tissue Eng Regen Med. 2013 Jan;7(1):73-84
references.pmid: 22095700
references.citation: Growth Factors. 2008 Jun;26(3):132-42
references.pmid: 18569021
references.citation: Biomed Mater. 2011 Apr;6(2):025011
references.pmid: 21436509
references.citation: J Cell Physiol. 2005 Mar;202(3):731-42
references.pmid: 15389573
references.citation: Tissue Eng Part A. 2010 May;16(5):1595-606
references.pmid: 20038211
references.citation: J Tissue Eng Regen Med. 2011 Apr;5(4):e36-51
references.pmid: 21413156
references.citation: Bone. 2005 Nov;37(5):607-21
references.pmid: 16039170
references.citation: Tissue Eng Part A. 2010 Sep;16(9):2941-51
references.pmid: 20575691
references.citation: Biochem J. 2012 May 1;443(3):603-18
references.pmid: 22507128
references.citation: Spine J. 2010 Jan;10(1):32-41
references.pmid: 19926342
references.citation: Cytokine. 2000 Nov;12(11):1630-8
references.pmid: 11052813
references.citation: Spine J. 2008 Mar-Apr;8(2):287-95
references.pmid: 17974491
references.citation: Adv Drug Deliv Rev. 2010 Sep 30;62(12):1167-74
references.pmid: 20920540
references.citation: J Orthop Trauma. 2005 Oct;19(9):629-34
references.pmid: 16247308
references.citation: Tissue Eng Part A. 2010 May;16(5):1633-43
references.pmid: 19995150
references.citation: Clin Orthop Relat Res. 2011 Oct;469(10):2706-15
references.pmid: 21403984
references.citation: J Knee Surg. 2009 Jan;22(1):60-71
references.pmid: 19216354
references.citation: Biotechnol Bioeng. 2003 Jul 20;83(2):173-80
references.pmid: 12768623
references.citation: Biomaterials. 2004 Jul;25(16):3211-22
references.pmid: 14980416
references.citation: Exp Cell Res. 2009 Jul 1;315(11):1937-52
references.pmid: 19331821
references.citation: Exp Mol Pathol. 2010 Feb;88(1):163-70
references.pmid: 19818765
references.citation: J Bone Joint Surg Am. 2010 Oct 20;92(14):2470-90
references.pmid: 20962200
references.citation: Methods. 2001 Dec;25(4):402-8
references.pmid: 11846609
references.citation: Mech Dev. 2003 Jul;120(7):823-36
references.pmid: 12915232
references.citation: Biochem Biophys Res Commun. 2004 Dec 10;325(2):453-60
references.pmid: 15530414
references.citation: Biochem Biophys Res Commun. 2010 Nov 12;402(2):329-34
references.pmid: 20937261
references.citation: Stem Cells. 2005 Mar;23(3):412-23
references.pmid: 15749936
references.citation: J Bone Joint Surg Am. 2010 Apr;92(4):994-1009
references.pmid: 20360528
references.citation: J Cell Biol. 1999 Jan 11;144(1):161-73
references.pmid: 9885252
references.citation: Plast Reconstr Surg. 2004 Feb;113(2):585-94
references.pmid: 14758221
references.citation: Annu Rev Physiol. 2000;62:413-37
references.pmid: 10845097
references.citation: Growth Factors. 2006 Dec;24(4):268-78
references.pmid: 17381068
references.citation: J Biosci Bioeng. 2011 Apr;111(4):489-92
references.pmid: 21185779
references.citation: Spine (Phila Pa 1976). 2004 Oct 15;29(20):2229-34
references.pmid: 15480133