Research article

Normal proliferation and differentiation of Hoxc-8 transgenic chondrocytes in vitro

Stephania A Cormier¹ , Maria Alice Mello^2,3 and Claudia Kappen^1,2,4

¹  Samuel C. Johnson Medical Research Center, Mayo Clinic Scottsdale, Scottdale, AZ 85259, USA

²  Center for Human Molecular Genetics, Munroe-Meyer Institute, University of Nebraska Medical Center, Omaha, NE 68198, USA

³  Current address: NIAMS/NIH, Cartilage Biology and Orthopedics Branch, Bethesda, MD 20892-8022, USA

⁴  Department of Genetics, Cell Biology and Anatomy, University of Nebraska Medical School, Omaha, NE 68198, USA

author email corresponding author email

BMC Developmental Biology 2003, 3:4doi:10.1186/1471-213X-3-4

Published:	24 April 2003

Abstract

Background

Hox genes encode transcription factors that are involved in pattern formation in the skeleton, and recent evidence suggests that they also play a role in the regulation of endochondral ossification. To analyze the role of Hoxc-8 in this process in more detail, we applied in vitro culture systems, using high density cultures of primary chondrocytes from neonatal mouse ribs.

Results

Cultured cells were characterized on the basis of morphology (light microscopy) and production of cartilage-specific extracellular matrix (sulfated proteoglycans and type II Collagen). Hypertrophy was demonstrated by increase in cell size, alkaline phosphatase activity and type X Collagen immunohistochemistry. Proliferation was assessed by BrdU uptake and flow cytometry. Unexpectedly, chondrocytes from Hoxc-8 transgenic mice, which exhibit delayed cartilage maturation in vivo [1], were able to proliferate and differentiate normally in our culture systems. This was the case even though freshly isolated Hoxc-8 transgenic chondrocytes exhibited significant molecular differences as measured by real-time quantitative PCR.

Conclusions

The results demonstrate that primary rib chondrocytes behave similar to published reports for chondrocytes from other sources, validating in vitro approaches for studies of Hox genes in the regulation of endochondral ossification. Our analysis of cartilage-producing cells from Hoxc-8 transgenic mice provides evidence that the cellular phenotype induced by Hoxc-8 overexpression in vivo is reversible in vitro.