title: Bioreactor-Induced Chondrocyte Maturation Is Dependent on Cell Passage and Onset of Loading.
abstract: OBJECTIVE
  To explore the effect of shifting in vitro culture conditions regarding cellular passage and onset of loading within matrix-associated bovine articular chondrocytes cultured under free-swelling and/or dynamical loading conditions on general chondrocyte maturation.
  
  
  METHODS
  Primary or passage 3 bovine chondrocytes were seeded in fibrin-polyurethane scaffolds. Constructs were cultured either free-swelling for 2 or 4 weeks, under direct mechanical loading for 2 or 4 weeks, or free swelling for 2 weeks followed by 2 weeks of loading. Samples were collected for glycosaminoglycan (GAG) quantification, mRNA expression of chondrogenic genes, immunohistochemistry, and histology.
  
  
  RESULTS
  Mechanical loading generally stimulated GAG synthesis, up-regulated chondrogenic genes, and improved the accumulation of matrix in cell-laden constructs when compared with free-swelling controls. Primary chondrocytes underwent more effective cartilage maturation when compared with passaged chondrocytes. Constructs of primary chondrocytes that were initially free-swelling followed by 2 weeks of mechanical load (delayed) had overall highest GAG with strongest responsiveness to load regarding matrix synthesis. Constructs that experienced the delayed loading regime also demonstrated most favorable chondrogenic gene expression profiles in both primary and third passage cells. Furthermore, most intense matrix staining and immunostaining of collagen type II and aggrecan were visualized in these constructs.
  
  
  CONCLUSIONS
  Primary chondrocytes were more effective than passage 3 chondrocytes when chondrogenesis was concerned. The most efficient chondrogenesis resulted from primary articular chondrocytes, which were initially free-swelling followed by a standardized loading protocol.
authors.affiliation: Department of Orthopaedics, General Hospital of Chinese PLA, Beijing, People's Republic of China ; Department of Orthopaedic and Trauma Surgery, University Medical Center, Albert-Ludwigs University Freiburg, Freiburg, Germany ; AO Research Institute Davos, Davos, Switzerland.
authors.forename: Ning
authors.initials: N
authors.lastname: Wang
authors.affiliation: AO Research Institute Davos, Davos, Switzerland.
authors.forename: Sibylle
authors.initials: S
authors.lastname: Grad
authors.affiliation: AO Research Institute Davos, Davos, Switzerland.
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Stoddart
authors.affiliation: Department of Orthopaedic and Trauma Surgery, University Medical Center, Albert-Ludwigs University Freiburg, Freiburg, Germany.
authors.forename: Philipp
authors.initials: P
authors.lastname: Niemeyer
authors.affiliation: Department of Orthopaedic and Trauma Surgery, University Medical Center, Albert-Ludwigs University Freiburg, Freiburg, Germany.
authors.forename: Norbert P
authors.initials: NP
authors.lastname: Südkamp
authors.affiliation: Department of Orthopaedic and Trauma Surgery, University Medical Center, Albert-Ludwigs University Freiburg, Freiburg, Germany.
authors.forename: Jan
authors.initials: J
authors.lastname: Pestka
authors.affiliation: AO Research Institute Davos, Davos, Switzerland.
authors.forename: Mauro
authors.initials: M
authors.lastname: Alini
authors.affiliation: Department of Orthopaedics, General Hospital of Chinese PLA, Beijing, People's Republic of China.
authors.forename: Jiying
authors.initials: J
authors.lastname: Chen
authors.affiliation: Department of Orthopaedic and Trauma Surgery, University Medical Center, Albert-Ludwigs University Freiburg, Freiburg, Germany.
authors.forename: Gian M
authors.initials: GM
authors.lastname: Salzmann
keywords: bioreactor; cartilage repair; chondrocytes; polyurethane scaffold; tissue engineering