title: Hypertrophy in mesenchymal stem cell chondrogenesis: effect of TGF-beta isoforms and chondrogenic conditioning.
abstract: Induction of chondrogenesis in mesenchymal stem cells (MSCs) with TGF-beta leads to a hypertrophic phenotype. The hypertrophic maturation of the chondrocytes is dependent on the timed removal of TGF-beta and sensitive to hypertrophy-promoting agents in vitro. In this study, we have investigated whether TGF-beta3, which has been shown to be more prochondrogenic compared to TGF-beta1, similarly enhances terminal differentiation in an in vitro hypertrophy model of chondrogenically differentiating MSCs. In addition, we tested the impact of the time of chondrogenic conditioning on the enhancement of hypertrophy. MSCs were chondrogenically differentiated in pellet culture in medium containing TGF-beta1 or TGF-beta3. After 2 or 4 weeks, chondrogenic medium was switched to hypertrophy-inducing medium for 2 weeks. Aggregates were analyzed histologically and biochemically on days 14, 28 and 42. The switch to hypertrophy medium after 14 days induced hypertrophic cell morphology and significant increase in alkaline phosphatase activity compared to the chondrogenesis only control using both TGF-beta1 and TGF-beta3. After 28 days predifferentiation, differences between hypertrophic and control groups diminished compared to 14 days predifferentiation. In conclusion, chondrogenic conditioning with both TGF-beta isoforms similarly induced hypertrophy in our experiment and allowed the enhancement of the hypertrophic chondrocyte phenotype by hypertrophic medium. Enhancement of hypertrophy was seen more clearly after the shorter chondrogenic conditioning. Therefore, to utilize this experimental model as a tool to study hypertrophy in MSC chondrogenesis, a predifferentiation period of 14 days is recommended.
authors.affiliation: Department for Trauma Surgery, Regensburg University Medical Center, Regensburg, Germany. michaelbmueller@web.de
authors.forename: Michael B
authors.initials: MB
authors.lastname: Mueller
authors.forename: Maria
authors.initials: M
authors.lastname: Fischer
authors.forename: Johannes
authors.initials: J
authors.lastname: Zellner
authors.forename: Arne
authors.initials: A
authors.lastname: Berner
authors.forename: Thomas
authors.initials: T
authors.lastname: Dienstknecht
authors.forename: Lukas
authors.initials: L
authors.lastname: Prantl
authors.forename: Richard
authors.initials: R
authors.lastname: Kujat
authors.forename: Michael
authors.initials: M
authors.lastname: Nerlich
authors.forename: Rocky S
authors.initials: RS
authors.lastname: Tuan
authors.forename: Peter
authors.initials: P
authors.lastname: Angele
mesh_terms: D000328:Adult; D000469:Alkaline Phosphatase / Q000378:metabolism; D002113:Calcification, Physiologic / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology*; D049108:Cell Enlargement / Q000187:drug effects*; D019902:Chondrocytes / Q000166:cytology* / Q000378:metabolism; D020219:Chondrogenesis / Q000187:drug effects / Q000502:physiology*; D024042:Collagen Type I / Q000378:metabolism; D024043:Collagen Type II / Q000378:metabolism; D024121:Collagen Type X / Q000378:metabolism; D017077:Culture Media, Conditioned / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism; D005260:Female; D005994:Glycerophosphates / Q000494:pharmacology; D006025:Glycosaminoglycans / Q000378:metabolism; D006801:Humans; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D020033:Protein Isoforms / Q000494:pharmacology; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology; D053782:Transforming Growth Factor beta3 / Q000494:pharmacology; D055815:Young Adult
chemical_list: D024042:Collagen Type I; D024043:Collagen Type II; D024121:Collagen Type X; D017077:Culture Media, Conditioned; D005994:Glycerophosphates; D006025:Glycosaminoglycans; D020033:Protein Isoforms; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D053782:Transforming Growth Factor beta3; C013786:A73025; D004247:DNA; D000469:Alkaline Phosphatase; C031463:beta-glycerophosphoric acid
references.citation: Exp Cell Res. 1998 Jan 10;238(1):265-72
references.pmid: 9457080
references.citation: Biomaterials. 2005 Feb;26(6):599-609
references.pmid: 15282138
references.citation: Biomaterials. 2005 Oct;26(28):5677-86
references.pmid: 15878373
references.citation: Histochem Cell Biol. 1999 Aug;112(2):115-23
references.pmid: 10460464
references.citation: Birth Defects Res C Embryo Today. 2003 May;69(2):123-43
references.pmid: 12955857
references.citation: Arthritis Rheum. 2006 Oct;54(10):3254-66
references.pmid: 17009260
references.citation: Proc Natl Acad Sci U S A. 2002 Apr 2;99(7):4397-402
references.pmid: 11917104
references.citation: Dev Growth Differ. 2007 Aug;49(6):449-54
references.pmid: 17661739
references.citation: In Vitro Cell Dev Biol Anim. 1995 Apr;31(4):288-94
references.pmid: 7540918
references.citation: Endocrinology. 1997 Nov;138(11):4966-76
references.pmid: 9348228
references.citation: Arthritis Rheum. 2008 May;58(5):1377-88
references.pmid: 18438858
references.citation: Tissue Eng. 2006 Sep;12(9):2639-47
references.pmid: 16995797
references.citation: Exp Cell Res. 2001 Aug 15;268(2):189-200
references.pmid: 11478845
references.citation: J Orthop Res. 2006 Nov;24(11):2095-105
references.pmid: 16955422
references.citation: J Cell Biochem. 1997 Sep 1;66(3):394-403
references.pmid: 9257195
references.citation: Dev Biol. 1993 Aug;158(2):414-29
references.pmid: 8344460
references.citation: Cytotherapy. 2004;6(6):596-601
references.pmid: 15773023
references.citation: J Biol Chem. 2003 Oct 17;278(42):41227-36
references.pmid: 12893825
references.citation: Cell Tissue Res. 2005 Nov;322(2):217-26
references.pmid: 16091918
references.citation: J Bone Joint Surg Am. 1998 Dec;80(12):1745-57
references.pmid: 9875932
references.citation: Clin Orthop Relat Res. 1999 Oct;(367 Suppl):S156-62
references.pmid: 10546644
references.citation: J Cell Biochem. 2006 Jan 1;97(1):33-44
references.pmid: 16215986
references.citation: Biochem Biophys Res Commun. 2003 Jun 20;306(1):186-90
references.pmid: 12788086
references.citation: J Orthop Res. 2006 Aug;24(8):1791-8
references.pmid: 16779832
references.citation: J Bone Miner Res. 2001 Feb;16(2):309-18
references.pmid: 11204431
references.citation: Matrix Biol. 2001 Jan;19(8):707-15
references.pmid: 11223330
references.citation: Tissue Eng. 2002 Feb;8(1):131-44
references.pmid: 11886661
references.citation: Tissue Eng. 1998 Winter;4(4):415-28
references.pmid: 9916173
references.citation: J Orthop Res. 1995 Nov;13(6):838-45
references.pmid: 8544019
references.citation: Anat Rec. 1989 Jun;224(2):167-79
references.pmid: 2672883