Calcified Tissue International

, Volume 88, Issue 5, pp 388–401 | Cite as

A Scaffold-Free Multicellular Three-Dimensional In Vitro Model of Osteogenesis

  • Umut A. Gurkan
  • Vipuil Kishore
  • Keith W. Condon
  • Teresita M. Bellido
  • Ozan AkkusEmail author
Original Research


In vitro models of osteogenesis are essential for investigating bone biology and the effects of pharmaceutical, chemical, and physical cues on bone formation. Osteogenesis takes place in a complex three-dimensional (3D) environment with cells from both mesenchymal and hematopoietic origins. Existing in vitro models of osteogenesis include two-dimensional (2D) single type cell monolayers and 3D cultures. However, an in vitro scaffold-free multicellular 3D model of osteogenesis is missing. We hypothesized that the self-inductive ossification capacity of bone marrow tissue can be harnessed in vitro and employed as a scaffold-free multicellular 3D model of osteogenesis. Therefore, rat bone marrow tissue was cultured for 28 days in three settings: 2D monolayer, 3D homogenized pellet, and 3D organotypic explant. The ossification potential of marrow in each condition was quantified by micro-computed tomography. The 3D organotypic marrow explant culture resulted in the greatest level of ossification with plate-like bone formations (up to 5 mm in diameter and 0.24 mm in thickness). To evaluate the mimicry of the organotypic marrow explants to newly forming native bone tissue, detailed compositional and morphological analyses were performed, including characterization of the ossified matrix by histochemistry, immunohistochemistry, Raman microspectroscopy, energy dispersive X-ray spectroscopy, backscattered electron microscopy, and micromechanical tests. The results indicated that the 3D organotypic marrow explant culture model mimics newly forming native bone tissue in terms of the characteristics studied. Therefore, this platform holds significant potential to be used as a model of osteogenesis, offering an alternative to in vitro monolayer cultures and in vivo animal models.


Scaffold-free In vitro model Marrow explant culture Osteogenesis Bone tissue engineering 



This study was funded by a grant from the Musculoskeletal Transplant Foundation. We thank David VanSickle, PhD, DVM, and Paul Snyder, PhD, DVM, for their insights on the histological results. We also thank Purdue Life Science Microscopy Facility director Debby Sherman for her support and help with the EDS and BSEM systems.


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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Umut A. Gurkan
    • 1
  • Vipuil Kishore
    • 2
  • Keith W. Condon
    • 3
  • Teresita M. Bellido
    • 3
  • Ozan Akkus
    • 2
    Email author
  1. 1.Center for Biomedical Engineering at Brigham and Women’s Hospital, Harvard Medical SchoolHarvard-MIT Division of Health Sciences and TechnologyBostonUSA
  2. 2.Weldon School of Biomedical EngineeringPurdue UniversityWest LafayetteUSA
  3. 3.Department of Anatomy and Cell BiologyIndiana University School of MedicineIndianapolisUSA

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