Hypertrophy and physiological death of equine chondrocytes in vitro.
Abstract: Equine osteochondrosis results from a failure of endochondral ossification during skeletal growth. Endochondral ossification involves chondrocyte proliferation, hypertrophy and death. Until recently no culture system was available to study these processes in equine chondrocytes. Objective: To optimise an in vitro model in which equine chondrocytes can be induced to undergo hypertrophy and physiological death as seen in vivo. Methods: Chondrocytes isolated from fetal or older (neonatal, growing and mature) horses were cultured as pellets in 10% fetal calf serum (FCS) or 10% horse serum (HS). The pellets were examined by light and electron microscopy. Total RNA was extracted from the pellets, and quantitative PCR carried out to investigate changes in expression of a number of genes regulating endochondral ossification. Results: Chondrocytes from fetal foals, grown as pellets, underwent hypertrophy and died by a process morphologically similar to that seen in vivo. Chondrocytes from horses age >5 months did not undergo hypertrophy in pellet culture. They formed intramembranous inclusion bodies and the cultures included cells of osteoblastic appearance. Pellets from neonatal foals cultured in FCS resembled pellets from older horses, however pellets grown in HS underwent hypertrophy but contained inclusion bodies. Chondrocytes from fetal foals formed a typical cartilage-like tissue grossly and histologically, and expressed the cartilage markers collagen type II and aggrecan mRNA. Expression of Sox9, collagen type II, Runx2, matrix metalloproteinase-13 and connective tissue growth factor mRNA increased at different times in culture. Expression of fibroblast growth factor receptor-3 and vascular endothelial growth factor mRNA decreased with time in culture. Conclusions: Freshly isolated cells from fetal growth cartilage cultured as pellets provide optimal conditions for studying hypertrophy and death of equine chondrocytes. Conclusions: This culture system should greatly assist laboratory studies aimed at elucidating the pathogenesis of osteochondrosis.
Publication Date: 2007-12-11 PubMed ID: 18065314DOI: 10.2746/042516407X223699Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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This research article presents an optimized model for studying hypertrophy and physiological death in equine chondrocytes, which can contribute to the understanding of osteochondrosis in horses.
Methods Used in the Research
- To study the processes involved in endochondral ossification, which is the failure of which results in equine osteochondrosis, the researchers isolated chondrocytes from horses of different ages, and cultured them as pellets in fetal calf serum (FCS) or horse serum (HS).
- The pellets were then examined under both light and electron microscopy.
- Further analysis was conducted by extracting total RNA from the pellets and using quantitative PCR to investigate changes in the expression of certain genes that regulate endochondral ossification.
Research Findings
- The research found that the chondrocytes taken from fetal foals underwent hypertrophy and died in a way that was morphologically similar to the process seen in vivo. However, chondrocytes taken from horses older than five months did not undergo hypertrophy in the pellet culture.
- The cultures that did not undergo hypertrophy formed intramembranous inclusion bodies and included cells that appeared osteoblastic.
- It was observed that pellets formed from neonatal foals and cultured in FCS appeared similar to pellets from older horses, while pellets grown in HS underwent hypertrophy but contained inclusion bodies.
- The chondrocytes from fetal foals formed a cartilage-like tissue both grossly and histologically, and expressed cartilage markers collagen Type II and aggrecan mRNA.
- The gene expressions of Sox9, collagen Type II, Runx2, matrix metalloproteinase-13, and connective tissue growth factor mRNA increased at different times in culture. Conversely, fibroblast growth factor receptor-3 and vascular endothelial growth factor mRNA expressions decreased with time in culture.
Conclusions and Implications of the Research
- The research concluded that freshly isolated cells from fetal growth cartilage provide optimal conditions for studying hypertrophy and death of equine chondrocytes when cultured as pellets.
- This method will aid laboratory studies focused on understanding the pathogenesis of osteochondrosis, providing new insight which could lead to improved preventative and treatment strategies for the condition.
Cite This Article
APA
Ahmed YA, Tatarczuch L, Pagel CN, Davies HM, Mirams M, Mackie EJ.
(2007).
Hypertrophy and physiological death of equine chondrocytes in vitro.
Equine Vet J, 39(6), 546-552.
https://doi.org/10.2746/042516407X223699 Publication
Researcher Affiliations
- School of Veterinary Science, University of Melbourne, Parkville, Victoria 3010, Australia.
MeSH Terms
- Age Factors
- Aging / physiology
- Analysis of Variance
- Animals
- Animals, Newborn
- Apoptosis
- Cell Death
- Cells, Cultured
- Chondrocytes / physiology
- Chondrocytes / ultrastructure
- Collagen Type II / metabolism
- Gene Expression Regulation
- Horse Diseases / pathology
- Horses
- Inclusion Bodies
- Matrix Metalloproteinase 13 / metabolism
- Microscopy, Electron / veterinary
- Osteochondritis / pathology
- Osteochondritis / veterinary
- Osteogenesis / physiology
- Polymerase Chain Reaction / veterinary
- RNA, Messenger / metabolism
Citations
This article has been cited 4 times.- Pilgrim CR, McCahill KA, Rops JG, Dufour JM, Russell KA, Koch TG. A Review of Fetal Bovine Serum in the Culture of Mesenchymal Stromal Cells and Potential Alternatives for Veterinary Medicine. Front Vet Sci 2022;9:859025.
- Rampin A, Skoufos I, Raghunath M, Tzora A, Diakakis N, Prassinos N, Zeugolis DI. Allogeneic Serum and Macromolecular Crowding Maintain Native Equine Tenocyte Function in Culture. Cells 2022 May 5;11(9).
- Ayodele BA, Mirams M, Pagel CN, Mackie EJ. The vacuolar H(+) ATPase V(0) subunit d(2) is associated with chondrocyte hypertrophy and supports chondrocyte differentiation. Bone Rep 2017 Dec;7:98-107.
- Franke J, Abs V, Zizzadoro C, Abraham G. Comparative study of the effects of fetal bovine serum versus horse serum on growth and differentiation of primary equine bronchial fibroblasts. BMC Vet Res 2014 May 26;10:119.
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