Functional dissection of genetic associations to osteoarthritis

Abstract

Osteoarthritis is a common, multifactorial disease, characterised by the progressive loss of articular cartilage from synovial joints. It has a major genetic component that is polygenic in nature. The aim of my thesis was to explore functional effects of several osteoarthritis susceptibility loci. I have carried out functional studies on five genes in total, covering three osteoarthritis susceptibility loci, including GDF5, MICAL3 and CHST11. I used protein, gene and allelic expression analyses. Firstly, I assessed whether human chondrocytes would respond in a predictable manner to GDF5. Reduced expression of GDF5 (chromosome 20q11.22) correlates with an increased risk for osteoarthritis and I assessed what effect providing exogenous GDF5 to chondrocytes had on expression of target genes. I observed that chondrocytes responded to exogenous GDF5 in a highly discordant manner. Secondly, I carried out gene and allelic expression analyses on MICAL3 (chromosome 22q11.21), to investigate if the association to osteoarthritis at this locus is mediated by an influence on gene expression. I observed allelic expression imbalance (AEI) at this locus, however, this did not correlate with genotype at the associated polymorphism in the human joint tissues investigated. I then focused on the susceptibility locus at CHST11 (chromosome 12q23.3). AEI was common at this gene, although a correlation with genotype at the associated polymorphism was not observed in the human joint tissues investigated. I carried out sequence analysis of CHST11 in osteoarthritic patients to identify rare amino acid coding variants, and found no evidence for any accounting for disease susceptibility. I explored the role of CHST11 in mesenchymal stem cells during chondrogenesis and observed that CHST11 knockdown leads to gene expression changes in chondrocyte marker genes and a reduction in cartilage extracellular matrix synthesis. My studies highlight the complexity in performing functional studies to identify and characterise the causal polymorphisms influencing osteoarthritis susceptibility.