A 300 μm Organotypic Bone Slice Culture Model for Temporal Investigation of Endochondral Osteogenesis

Sriveena Srinivasaiah, Giuseppe Musumeci, Tamilselvan Mohan, Paola Castrogiovanni, Markus Absenger-Novak, Ulrike Zefferer, Sepideh Mostofi, Ehsan Bonyadi Rad, Nicole Gabriele Grün, Annelie Martina Weinberg, Ute Schafer*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review


Translational studies to elucidate the response of immature bone to biologic and physical stimuli have been held back by the lack of a viable long-term functional bone explant model. This study attempts to bridge this gap between cell culture and animal model studies. In this study, we describe a methodology to derive a 300 μm organotypic femur slice comprising physiological zones (epiphysis and meta-diaphysis) essential for endochondral bone development. The unique capability of slice culture model incorporating enhanced nutrient access to distinct bone tissue components associated with linear bone growth facilitates the investigation of the orchestrated cellular transition of chondrogenic and osteogenic cells involved in endochondral bone development in an ex vivo setup. Bone slices of 300 μm were prepared from 4-day-old postnatal rats and were viable in culture up to 21 days. On days 7 and 15, an increase in chondrogenic and osteogenic modulations was confirmed in epiphysis, metaphysis, and diaphysis. An increase in osteocytes, osteoblasts, and hypertrophic cells were found at these time points, as well as a noticeable increased expression of chondrogenic and osteogenic markers (collagen II, Runx2, and osteocalcin) confirmed endochondral progression. Osteoclast-mediated bone resorption was demonstrated on day 15 by tartrate-resistant acid phosphatase staining. Attenuated total reflection infrared spectroscopic analyses, furthermore, confirmed a time-dependent increase in phosphate levels, bone minerals, and hydroxyapatite for 15 days. Our establishment of a bone slice culture model closely mimicking the in vivo cellular transitions and endochondral microenvironment of a mineralizing bone provides a vital new tool for the elucidation of cellular and endochondral mechanisms of bone development, maturation, and growth plate modulations. The presented model has the potential to be utilized in implementation of preclinical, toxicological, and therapeutic investigations.

Original languageEnglish
Pages (from-to)197-212
Number of pages16
JournalTissue Engineering / C
Issue number4
Publication statusPublished - 1 Apr 2019


  • bone
  • bone morphogenesis
  • bone slice model
  • cartilage
  • endochondral ossification
  • organotypic bone culture

ASJC Scopus subject areas

  • Bioengineering
  • Medicine (miscellaneous)
  • Biomedical Engineering


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