Searchable abstracts of presentations at key conferences in endocrinology
Endocrine Abstracts (2011) 25 P17

SFEBES2011 Poster Presentations Bone (26 abstracts)

A 5′-untranslated region mutation of the growth and differentiation factor 5 (Gdf5) gene increases expression and is associated with decreased urinary excretion of the cartilage degradation product, CTX-II: relevance to osteoarthritis

M Andrew Nesbit 1 , Chris Esapa 1 , Rosie Head 1 , Katie Gaynor 1 , Roger Cox 2 , Steve Brown 2 & Rajesh Thakker 1

1University of Oxford, Oxford, UK; 2MRC Harwell, Oxfordshire, UK.

Osteoarthritis (OA) may be associated with endocrine disorders such as hypothyroidism, obesity, primary hyperparathyroidism or acromegaly, although often its cause remains undefined. To facilitate investigations of the underlying molecular mechanisms of OA we have investigated N-ethyl-N-nitrosourea (ENU) mutant mice using a genotype-driven approach in which candidate genes are examined for mutations. One such investigated gene is growth and differentiation factor 5 (GDF5), as in man a single nucleotide polymorphism (SNP) in its 5′ untranslated region, which reduces GDF5 expression in joints, has been reported to be associated with susceptibility to knee and hip OA. Our analysis of 10 000 DNA samples from ENU mutagenised mice identified an A/G SNP in a conserved nucleotide 47 bp downstream of the human OA susceptibility SNP and 225 bp upstream of the translation start site of the Gdf5 gene. The in vitro and in vivo effects of this polymorphism on GDF5 expression were investigated. Luciferase reporter assays of Gdf5 promoter polymorphisms in MG63 (osteoblast cells) and CH8 (cartilage cells) demonstrated that the ENU mutant (−225G) increases Gdf5 expression (1.5- and 2-fold respectively, P<0.005) in contrast to the human OA-associated polymorphism (−272T) which decreases expression (0.75- and 0.7-fold respectively, P<0.05). For in vivo studies, ENU mutant and wild-type mice were kept in accordance with national welfare guidelines and project license restrictions, aged for 14 weeks and investigated for the urinary excretion of the cartilage degradation product, CTX-II, which may be elevated in OA patients. This revealed that female homozygous mutant mice, when compared to wild-type littermates, had significantly reduced 24-h urinary CTX-II excretion (mutant 2.90±0.01 ng/ml; wild-type 4.87±0.036 ng/ml, P<0.05) thereby suggesting that the mutant confers a protective effect against OA. Thus, we have established a mouse model with a functional alteration in Gdf5 expression that will facilitate investigation of the molecular mechanisms of OA.

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