title: Simulated microgravity affects chondrogenesis and hypertrophy of human mesenchymal stem cells.
abstract: OBJECTIVE
  During in vitro chondrogenesis of human mesenchymal stem cells (hMSCs) hypertrophy is an inadvertent event associated with cell differentiation toward the osteogenic lineage. Up to now, there is no stringent experimental control mechanism to prevent hypertrophy of MSCs. Microgravity is known to have an impact on osteogenesis. In this study, the influence of simulated microgravity (SMG) on both chondrogenesis and hypertrophy of hMSCs was evaluated.
  
  
  METHODS
  A bioreactor using a rotating wall vessel was constructed to simulate microgravity. Pellet cultures formed from hMSCs (P5) were supplemented with human transforming growth factor-β3 (TGF-β3). The hMSC pellet cultures treated with TGF-β3 were either kept in SMG or in a control system. After three weeks of culture, the chondrogenic differentiation status and level of hypertrophy were examined by safranin-O staining, immunohistochemistry and quantitative real-time PCR.
  
  
  RESULTS
  SMG reduced the staining for safranin-O and collagen type II. The expression of collagen type X α1 chain (COL10A1) and collagen type II α1 chain (COL2A1) were both significantly reduced. There was a higher decrease in COL2A1 than in COL10A1 expression, resulting in a low COL2A1/COL10A1 ratio.
  
  
  CONCLUSIONS
  SMG reduced hypertrophy of hMSCs during chondrogenic differentiation. However, the expression of COL2A1 was likewise reduced. Even more, the COL2A1/COL10A1 ratio decreased under SMG conditions. We therefore assume that SMG has a significant impact on the chondrogenic differentiation of hMSCs. However, due to the high COL2A1 suppression under SMG, this culture system does not yet seem to be suitable for a potential application in cartilage repair.
authors.affiliation: Department of Orthopaedic Surgery, Ludwig-Maximilians-University, Campus Großhadern, Marchioninistr. 15, 81377, Munich, Germany, susanne.mayer@med.uni-muenchen.de.
authors.forename: Susanne
authors.initials: S
authors.lastname: Mayer-Wagner
authors.forename: Florian
authors.initials: F
authors.lastname: Hammerschmid
authors.forename: Julia I
authors.initials: JI
authors.lastname: Redeker
authors.forename: Bärbel
authors.initials: B
authors.lastname: Schmitt
authors.forename: Boris Michael
authors.initials: BM
authors.lastname: Holzapfel
authors.forename: Volkmar
authors.initials: V
authors.lastname: Jansson
authors.forename: Oliver B
authors.initials: OB
authors.lastname: Betz
authors.forename: Peter E
authors.initials: PE
authors.lastname: Müller
mesh_terms: D053674:Aggrecans / Q000378:metabolism; D002454:Cell Differentiation; D020219:Chondrogenesis / Q000502:physiology*; D024043:Collagen Type II / Q000378:metabolism; D024121:Collagen Type X / Q000378:metabolism; D006801:Humans; D006984:Hypertrophy; D007150:Immunohistochemistry; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000473:pathology*; D010012:Osteogenesis; D060888:Real-Time Polymerase Chain Reaction; D018474:Weightlessness Simulation
chemical_list: D053674:Aggrecans; C499177:COL2A1 protein, human; D024043:Collagen Type II; D024121:Collagen Type X