Primary osteoarthritis (OA) is a late-onset, degenerative condition of synovial joints, and is the major cause of pain and disability in older persons. OA represents a significant disease burden and focus of research, especially as no disease-modifying therapies exist to manage the condition. The genetic influence to OA is complex and polygenic. The arcOGEN study, the most powerful genome-wide association study yet to investigate OA in humans, identified the 9q33.1 locus to be significantly associated with hip OA in females. TRIM32 lies within the 9q33.1 susceptibility locus and may have strong biological relevance to OA; it encodes a protein with E3 ubiquitin ligase activity. Sanger sequencing of TRIM32 in the youngest 500 female patients with hip OA from the arcOGEN study was performed to identify rare variants in TRIM32 that are associated with OA of the hip in females. Polymorphisms were identified in the proximal promoter, and 3’untranslated regions (3’UTR) of TRIM32 that are disproportionately represented in female patients with hip OA, compared to the control population. In vitro studies identified expression of TRIM32 in human femoral head cartilage; reduced expression of TRIM32 was also demonstrated in femoral head primary articular chondrocytes from patients with hip OA compared to control patients. Trim32 knockout resulted in increased aggrecanolysis in murine femoral head explants. Murine chondrocytes deficient in Trim32 also exhibited increased expression of markers of a mature chondrocyte phenotype in response to anabolic cytokine stimulation, and increased expression of markers of a hypertrophic chondrocyte phenotype upon catabolic cytokine stimulation. In vivo studies of joint degeneration in Trim32 knockout mice demonstrated increased cartilage degradation and tibial epiphyseal bone changes after surgically induced knee joint instability, compared to wild-type mice. Increased cartilage degradation and medial knee subchondral bone changes were also identified upon ageing of Trim32 knockout mice. These results further implicate TRIM32 in the genetic predisposition to OA, and indicate a role for TRIM32 in the joint degeneration evident in OA. These results support the further study of TRIM32 in the pathophysiology of OA and development of novel therapeutic strategies to manage OA.

Abstract This work engaged Finnish females affected with osteoarthritis (OA) of the hand to define the role of common sequence variations within the genes of the important structural protein of cartilage, aggrecan (AGC1), and the genes of inflammatory mediators, the interleukin 1 gene cluster and interleukin 6 (IL6), as possible risk factors for the disease. Also, a genome-wide linkage analysis was performed in a sample consisting of Finnish families with multiple individuals affected with hip and knee OA in order to reveal new chromosomal areas that are likely to contain disease associated variations. OA is a chronic disease that leads to the degeneration of articular cartilage in synovial joints. The etiology of the disease is for the most part unknown. Joints of the hand, hip and knee are most commonly affected, and obesity, trauma and excess mechanical stress are known risk factors for the disease. OA also has a significant genetic component. AGC1 carries a variable number of tandem repeats (VNTR) polymorphism, which may be significant for the biomechanical properties of cartilage. It was shown that the most common allele with 27 tandem repeats is protective against hand OA (HOA) (odds ratio 0.46, 95% confidence interval 0.27–0.78). Also, carrying two copies of any of the shorter or longer alleles increased the risk of the disease. Inflammation seems to play a role in the etiology of OA and certain polymorphisms within the interleukin 1 gene cluster and IL6 have been previously shown to increase the transcription of these molecules and to associate with OA. In this study it was shown that the G alleles in three common IL6 promoter single nucleotide polymorphism (SNP) sites are associated with the risk of more severe forms of HOA (p =  0.001 for GGG haplotype). A SNP in IL1B associated with the bilateral form of the disease (p =  0.006) and two IL1B-IL1RN extended haplotype alleles were associated with the same phenotype. Genome-wide and fine mapping linkage analyses recognized chromosomal locus 2q21 with a multipoint LOD score of 3.96. Despite the association analyses of several candidate genes within the locus, no disease-associating sequence variants were identified.

Osteoarthritis (OA) is a common, multifactorial disease that is characterized by focal degeneration of the smooth articular cartilage in any of the synovial joints. Although the underlying molecular mechanisms for OA are still not fully understood, epidemiological studies have evidenced a significant genetic component to OA susceptibility. Genome-wide linkage scans and large-scale association studies have had success in unraveling the genetic architecture underlying OA with the identification of a number of susceptibility genes. In this work, functional analyses are reported of OA associated polymorphisms within two susceptibility genes: BMP5 and GDF5, both members of the TGF-β superfamily of secreted proteins. The extent of differential allelic expression (DAE) of BMP5 in human mesenchymal tissues was first examined with significant differences in BMP5 allelic output observed (allelic ratios exceeding 4:1 in the tissues of some donors). Significant variability in allelic expression within the different tissues of donors was also observed, suggesting that polymorphism in cis-regulation of BMP5 expression is common and that there is a considerable effect of tissue specific elements on BMP5 expression. DAE was then used as a phenotype to map tissue-specific cis-regulatory polymorphisms with the identification of a single nucleotide polymorphism (SNP) located downstream of BMP5 that was significantly associated with DAE as well as OA, suggesting that variability in cis-regulation of BMP5 is important in OA susceptibility. Moreover, the functional effect of a previously identified OA associated microsatellite within intron 1 of BMP5 was investigated using luciferase reporter assays and electrophoretic mobility shift assays (EMSAs) with significant differences observed both in the ability of various microsatellite alleles to modulate BMP5 promoter activity and to bind GATA-3 nuclear proteins, further suggesting a role for variability in BMP5 expression in OA susceptibility that may in part be due to altered GATA-3 binding. Finally, functional characterization of a previously reported OA associated SNP in the 5′ UTR of GDF5 is presented in which EMSAs show differential binding of nuclear factors between the two SNP alleles, strengthening the possible functional contribution of this SNP to OA susceptibility. Overall, this work demonstrates that polymorphism in cis-regulation is likely to play a role in OA susceptibility.