Effect on ligament marker expression by direct-contact co-culture of mesenchymal stem cells and anterior cruciate ligament cells

Jose A. Canseco, Koji Kojima, Ashley R. Penvose, Jason D. Ross, Haruko Obokata, Andreas H. Gomoll, Charles A. Vacanti*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

33 Scopus citations


Ligament and tendon repair is an important topic in orthopedic tissue engineering; however, the cell source for tissue regeneration has been a controversial issue. Until now, scientists have been split between the use of primary ligament fibroblasts or marrow-derived mesenchymal stem cells (MSCs). The objective of this study was to show that a co-culture of anterior cruciate ligament (ACL) cells and MSCs has a beneficial effect on ligament regeneration that is not observed when utilizing either cell source independently. Autologous ACL cells (ACLcs) and MSCs were isolated from Yorkshire pigs, expanded in vitro, and cultured in multiwell plates in varying %ACLcs/%MSCs ratios (100/0, 75/25, 50/50, 25/75, and 0/100) for 2 and 4 weeks. Quantitative mRNA expression analysis and immunofluorescent staining for ligament markers Collagen type I (Collagen-I), Collagen type III (Collagen-III), and Tenascin-C were performed. We show that Collagen-I and Tenascin-C expression is significantly enhanced over time in 50/50 co-cultures of ACLcs and MSCs (p≤0.03), but not in other groups. In addition, Collagen-III expression was significantly greater in MSC-only cultures (p≤0.03), but the Collagen-I-to-Collagen-III ratio in 50% co-culture was closest to native ligament levels. Finally, Tenascin-C expression at 4 weeks was significantly higher (p≤0.02) in ACLcs and 50% co-culture groups compared to all others. Immunofluorescent staining results support our mRNA expression data. Overall, 50/50 co-cultures had the highest Collagen-I and Tenascin-C expression, and the highest Collagen-I-to-Collagen-III ratio. Thus, we conclude that using a 50% co-culture of ACLcs and MSCs, instead of either cell population alone, may better maintain or even enhance ligament marker expression and improve healing.

Original languageEnglish (US)
Pages (from-to)2549-2558
Number of pages10
JournalTissue Engineering - Part A
Issue number23-24
StatePublished - Dec 1 2012

ASJC Scopus subject areas

  • Bioengineering
  • Biochemistry
  • Biomaterials
  • Biomedical Engineering


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