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Effects of hypoxia on anabolic and catabolic gene expression and DNA methylation in OA chondrocytes

BACKGROUND: Cartilage is an avascular and aneural tissue. Chondrocytes thrive in this restricted environment of low oxygen tension and poor nutrient availability which has led to suggestions that hypoxia may be a protective mechanism against the development of osteoarthritis (OA). There is also a gr...

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Autores principales: Alvarez, Karl, de Andrés, María C, Takahashi, Atsushi, Oreffo, Richard O C
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2014
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4301830/
https://www.ncbi.nlm.nih.gov/pubmed/25510649
http://dx.doi.org/10.1186/1471-2474-15-431
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author Alvarez, Karl
de Andrés, María C
Takahashi, Atsushi
Oreffo, Richard O C
author_facet Alvarez, Karl
de Andrés, María C
Takahashi, Atsushi
Oreffo, Richard O C
author_sort Alvarez, Karl
collection PubMed
description BACKGROUND: Cartilage is an avascular and aneural tissue. Chondrocytes thrive in this restricted environment of low oxygen tension and poor nutrient availability which has led to suggestions that hypoxia may be a protective mechanism against the development of osteoarthritis (OA). There is also a growing body of evidence to support the role of epigenetic factors in the pathogenesis of OA. However, few studies have investigated the epigenetic-OA process within a hypoxic environment. The current study has investigated the effects of hypoxia on gene expression and DNA methylation of anabolic and catabolic genes involved in the pathogenesis of OA. METHODS: Chondrocytes extracted from OA femoral heads were incubated in normoxia and hypoxia (20% and 2% oxygen concentrations respectively). Interleukin 1-beta (IL-1β) plus oncostatin M (OSM), 5-azadeoxycytidine (5-aza-dC) or media alone (control) were added twice weekly to the incubated samples. After 5 weeks, levels of Collagen type IX (COL9A1), IL1B, and matrix metalloproteinase-13 (MMP13) gene expression were measured using SYBR Green-based qRT-PCR and were correlated with methylation status analysed by pyrosequencing methodology. RESULTS: Hypoxia resulted in a >50-fold and >10-fold increase in relative expression of COL9A1 and IL1B respectively. This was inversely correlated to the DNA methylation status of these genes. Expression of MMP13 was reduced at 2% oxygen tension in control cells. Relative expression of MMP13 increased in cells stimulated with IL-1β and 5-aza-dC in normoxic conditions, and this effect was eliminated at low oxygen tension although no correlation with methylation status was observed. CONCLUSIONS: These findings demonstrate a role for hypoxia in the regulation of anabolic and catabolic gene expression and the influence of changes in DNA methylation. These results further support the role of epigenetics in OA and, critically, highlight the complex relationship between the physiological environment of cartilaginous cells and the osteoarthritic process with implications for therapeutic intervention and our understanding of OA pathophysiology. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/1471-2474-15-431) contains supplementary material, which is available to authorized users.
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spelling pubmed-43018302015-01-22 Effects of hypoxia on anabolic and catabolic gene expression and DNA methylation in OA chondrocytes Alvarez, Karl de Andrés, María C Takahashi, Atsushi Oreffo, Richard O C BMC Musculoskelet Disord Research Article BACKGROUND: Cartilage is an avascular and aneural tissue. Chondrocytes thrive in this restricted environment of low oxygen tension and poor nutrient availability which has led to suggestions that hypoxia may be a protective mechanism against the development of osteoarthritis (OA). There is also a growing body of evidence to support the role of epigenetic factors in the pathogenesis of OA. However, few studies have investigated the epigenetic-OA process within a hypoxic environment. The current study has investigated the effects of hypoxia on gene expression and DNA methylation of anabolic and catabolic genes involved in the pathogenesis of OA. METHODS: Chondrocytes extracted from OA femoral heads were incubated in normoxia and hypoxia (20% and 2% oxygen concentrations respectively). Interleukin 1-beta (IL-1β) plus oncostatin M (OSM), 5-azadeoxycytidine (5-aza-dC) or media alone (control) were added twice weekly to the incubated samples. After 5 weeks, levels of Collagen type IX (COL9A1), IL1B, and matrix metalloproteinase-13 (MMP13) gene expression were measured using SYBR Green-based qRT-PCR and were correlated with methylation status analysed by pyrosequencing methodology. RESULTS: Hypoxia resulted in a >50-fold and >10-fold increase in relative expression of COL9A1 and IL1B respectively. This was inversely correlated to the DNA methylation status of these genes. Expression of MMP13 was reduced at 2% oxygen tension in control cells. Relative expression of MMP13 increased in cells stimulated with IL-1β and 5-aza-dC in normoxic conditions, and this effect was eliminated at low oxygen tension although no correlation with methylation status was observed. CONCLUSIONS: These findings demonstrate a role for hypoxia in the regulation of anabolic and catabolic gene expression and the influence of changes in DNA methylation. These results further support the role of epigenetics in OA and, critically, highlight the complex relationship between the physiological environment of cartilaginous cells and the osteoarthritic process with implications for therapeutic intervention and our understanding of OA pathophysiology. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/1471-2474-15-431) contains supplementary material, which is available to authorized users. BioMed Central 2014-12-15 /pmc/articles/PMC4301830/ /pubmed/25510649 http://dx.doi.org/10.1186/1471-2474-15-431 Text en © Alvarez et al.; licensee BioMed Central Ltd. 2014 This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Research Article
Alvarez, Karl
de Andrés, María C
Takahashi, Atsushi
Oreffo, Richard O C
Effects of hypoxia on anabolic and catabolic gene expression and DNA methylation in OA chondrocytes
title Effects of hypoxia on anabolic and catabolic gene expression and DNA methylation in OA chondrocytes
title_full Effects of hypoxia on anabolic and catabolic gene expression and DNA methylation in OA chondrocytes
title_fullStr Effects of hypoxia on anabolic and catabolic gene expression and DNA methylation in OA chondrocytes
title_full_unstemmed Effects of hypoxia on anabolic and catabolic gene expression and DNA methylation in OA chondrocytes
title_short Effects of hypoxia on anabolic and catabolic gene expression and DNA methylation in OA chondrocytes
title_sort effects of hypoxia on anabolic and catabolic gene expression and dna methylation in oa chondrocytes
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4301830/
https://www.ncbi.nlm.nih.gov/pubmed/25510649
http://dx.doi.org/10.1186/1471-2474-15-431
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