Osteoarthritis Genetic Risk Research Articles

Genetic risk of osteoarthritis operates during human skeletogenesis.

Osteoarthritis (OA) is a polygenic disease of older people resulting in the breakdown of cartilage within articular joints. Although it is a leading cause of disability, there are no disease-modifying therapies. Evidence is emerging to support the origins of OA in skeletogenesis. Whereas methylation quantitative trait loci (mQTLs) co-localizing with OA genome-wide association study signals have been identified in aged human cartilage and used to identify effector genes and variants, such analyses have never been conducted during human development. Here, for the first time, we have investigated the developmental origins of OA genetic risk at seven well-characterized OA risk loci, comprising 39 OA-mQTL CpGs, in human fetal limb (FL) and cartilage (FC) tissues using a range of molecular genetic techniques. We identified significant OA-mQTLs at 14 and 29 CpGs in FL and FC tissues, respectively, and compared our results with aged cartilage samples (AC). Differential methylation was observed at 26 sites between FC and AC, with the majority becoming actively hypermethylated in old age. Notably, 6/9 OA effector genes showed allelic expression imbalances during fetal development. Finally, we conducted ATAC-sequencing in cartilage from the developing and aged hip and knee to identify accessible chromatin regions and found enrichment for transcription factor binding motifs including SOX9 and FOS/JUN. For the first time, we have demonstrated the activity of OA-mQTLs and expression imbalance of OA effector genes during human skeletogenesis. We show striking differences in the spatiotemporal function of these loci, contributing to our understanding of OA aetiology, with implications for the timing and strategy of pharmacological interventions.

EPIGENOMIC ANALYSIS OF OSTEOARTHRITIS GENETIC RISK DURING HUMAN FETAL DEVELOPMENT

Purpose: Over 100 single nucleotide variants (SNVs) contributing to osteoarthritis (OA) disease risk have now been reported. Risk-conferring variants typically operate by altering expression of a target gene and epigenetic mechanisms within chondrocytes can mediate this regulation. DNA methylation (DNAm) at CpG dinucleotides (CpGs) can correlate with SNV genotype, creating methylation quantitative trait loci (mQTLs). Previously, we have reported mQTLs associated with OA genetic risk loci in aged articular cartilage.

Chondrocyte protein co-synthesis network analysis links ECM mechanosensing to metabolic adaptation in osteoarthritis

Background Knee osteoarthritis (OA) is one of the most common structural OA disorders globally. Incomplete understanding of the fundamental biological aspects of osteoarthritis underlies the current lack of effective treatment or disease modifying drugs. Research Design and Methods We implemented a systems approach by making use of the statistical network concepts in Weighted Gene Co-expression Analysis to reconstruct the organisation of the core proteome network in chondrocytes obtained from OA patients and healthy individuals. Protein modules reflect groups of tightly co-ordinated changes in protein abundance across healthy and OA chondrocytes. Results The unbiased systems analysis identified extracellular matrix (ECM) mechanosensing and glycolysis as two modules that are most highly correlated with ΟΑ. The ECM module was enriched in the OA genetic risk factors tenascin-C (TNC) and collagen 11A1 (COL11A1), as well as in cartilage oligomeric matrix protein (COMP), a biomarker associated with cartilage integrity. Mapping proteins that are unique to OA or healthy chondrocytes onto the core interactome, which connects microenvironment sensing and regulation of glycolysis, identified differences in metabolic and anti-inflammatory adaptation. Conclusion The interconnection between cartilage ECM remodeling and metabolism is indicative of the dynamic chondrocyte states and their significance in osteoarthritis.

Multi-Tissue Epigenetic and Gene Expression Analysis Combined With Epigenome Modulation Identifies RWDD2B as a Target of Osteoarthritis Susceptibility.

Osteoarthritis (OA) is polygenic, with more than 90 risk loci currently mapped, including at the single-nucleotide polymorphism rs6516886. Previous analysis of OA cartilage DNA identified 6 CpG dinucleotides whose methylation levels correlated with the rs6516886 genotype, forming methylation quantitative trait loci (mQTLs). We undertook this study to investigate these mQTLs and to map expression quantitative trait loci (eQTLs) across joint tissues in order to prioritize a particular gene as a target of the rs6516886 association effect. Nucleic acids were extracted from the cartilage, fat pad, synovium, and peripheral blood from OA patients. Methylation of CpGs and allelic expression imbalance of potential target genes were assessed by pyrosequencing. A chondrocyte cell line expressing deactivated Cas9 (dCas9)-TET1 was used to directly alter CpG methylation levels, with effects on gene expression quantified by polymerase chain reaction. Multiple mQTLs were detected, with effects strongest in joint tissues and with methylation at CpG cg20220242 correlating most significantly with the rs6516886 genotype. CpG cg20220242 is located upstream of RWDD2B. Significant rs6516886 eQTLs were observed for this gene, with the OA risk-conferring allele of rs6516886 correlating with reduced expressionCpG methylation also correlated with allelic expression ofRWDD2B, forming methylation-expression QTLs (meQTLs). Deactivated Cas9-TET1 reduction in the methylation of cg20220242 increased expression of RWDD2B. The rs6516886 association signal is a multi-tissue meQTL involving cg20220242 and acting onRWDD2B. Modulating CpG methylation reverses the impact of the risk allele. RWDD2B codes for a protein about which little is currently known. Its further analysis as a target of OA genetic risk will provide novel insight into this complex disease.

Duration of symptoms prior to partial meniscectomy is not associated with the expression of osteoarthritis genes in the injured meniscus.

While there is emerging data on how duration of symptoms prior to surgery relates to outcomes of patients undergoing arthroscopic partial meniscectomy, little is known about how duration of symptoms relates to the biology of the knee in these patients. The purpose of this study was to test the hypothesis that duration of symptoms prior to arthroscopic partial meniscectomy is associated with expression of osteoarthritis (OA)-related genes in the meniscus. We collected resected meniscus from patients (N = 76) undergoing clinically indicated arthroscopic partial meniscectomy from knees without advanced degenerative changes. RNA from 64 patients was analyzed for 28 candidate OA transcripts by real-time polymerase chain reaction (PCR). RNA was also probed for identification of novel genes by RNA microarray in 12 patients followed by validation of selected candidates by real-time PCR. The association of gene expression with duration of symptoms prior to surgery was tested. Additional screening was performed with known OA genetic risk alleles assembled from published literature and with gene transcripts differentially expressed between non-OA and OA cartilage and menisci. Our data revealed that duration of symptoms did not predict expression of OA genes in the meniscus, other than limited association with CXCL3, BMP2, and HLA-DQA1. Microarray identified new genes and pathways with unknown role(s) in meniscus injury and OA and validation of a subset of genes by real-time PCR showed expression pattern highly concordant with the microarray data. While duration of symptoms prior to arthroscopic partial meniscectomy does not significantly alter the expression of OA related genes, the association with novel genes and pathways deserves further investigation.

Identification of a novel, methylation-dependent, RUNX2 regulatory region associated with osteoarthritis risk.

Osteoarthritis (OA) is a common, multifactorial and polygenic skeletal disease that, in its severest form, requires joint replacement surgery to restore mobility and to relieve chronic pain. Using tissues from the articulating joints of 260 patients with OA and a range of in vitro experiments, including CRISPR-Cas9, we have characterized an intergenic regulatory element. Here, genotype at an OA risk locus correlates with differential DNA methylation, with altered gene expression of both a transcriptional regulator (RUNX2), and a chromatin remodelling protein (SUPT3H). RUNX2 is a strong candidate for OA susceptibility, with its encoded protein being essential for skeletogenesis and healthy joint function. The OA risk locus includes single nucleotide polymorphisms (SNPs) located within and flanking the differentially methylated region (DMR). The OA association SNP, rs10948172, demonstrates particularly strong correlation with methylation, and two intergenic SNPs falling within the DMR (rs62435998 and rs62435999) demonstrate genetic and epigenetic effects on the regulatory activity of this region. We therefore posit that the OA signal mediates its effect by modulating the methylation of the regulatory element, which then impacts on gene expression, with RUNX2 being the principal target. Our study highlights the interplay between DNA methylation, OA genetic risk and the downstream regulation of genes critical to normal joint function.

Evaluation of shared genetic aetiology between osteoarthritis and bone mineral density identifies SMAD3 as a novel osteoarthritis risk locus.

Osteoarthritis (OA) is a common complex disease with high public health burden and no curative therapy. High bone mineral density (BMD) is associated with an increased risk of developing OA, suggesting a shared underlying biology. Here, we performed the first systematic overlap analysis of OA and BMD on a genome wide scale. We used summary statistics from the GEFOS consortium for lumbar spine (n = 31,800) and femoral neck (n = 32,961) BMD, and from the arcOGEN consortium for three OA phenotypes (hip, ncases=3,498; knee, ncases=3,266; hip and/or knee, ncases=7,410; ncontrols=11,009). Performing LD score regression we found a significant genetic correlation between the combined OA phenotype (hip and/or knee) and lumbar spine BMD (rg=0.18, P = 2.23 × 10−2), which may be driven by the presence of spinal osteophytes. We identified 143 variants with evidence for cross-phenotype association which we took forward for replication in independent large-scale OA datasets, and subsequent meta-analysis with arcOGEN for a total sample size of up to 23,425 cases and 236,814 controls. We found robustly replicating evidence for association with OA at rs12901071 (OR 1.08 95% CI 1.05–1.11, Pmeta=3.12 × 10−10), an intronic variant in the SMAD3 gene, which is known to play a role in bone remodeling and cartilage maintenance. We were able to confirm expression of SMAD3 in intact and degraded cartilage of the knee and hip. Our findings provide the first systematic evaluation of pleiotropy between OA and BMD, highlight genes with biological relevance to both traits, and establish a robust new OA genetic risk locus at SMAD3.

Genetic risk factors for knee OA display good predictive power in a high risk group of overweight and obese females

Purpose: Over the last few years, several genetic markers have been identified to be robustly associated with knee osteoarthritis (OA). These genetic markers have been identified by gathering all cases and controls available. In general, small effect sizes of these risk factors were found, making OA genetic risk prediction difficult. However, it is known that the association of the genetic marker with the phenotype can be context specific. It could be that in persons with or without major environmental risk factors, different sets of genes are important. We therefore set out to test predictive value of DNA variants previously identified as being associated to knee OA, in a high risk group of overweight and obese, middle-aged women. Methods: Study Sample: This study was performed within the PRevention of knee Osteoarthritis in Overweight Females (PROOF) study. A total of 407 women met the inclusion criteria (BMI ≥ 27, no clinical knee OA (ACR criteria), no contraindications to MRI, no rheumatic diseases), were invited for baseline measurements. Pre-specified primary outcome was incidence of knee OA, defined by incidence of either K&L ≥ 2, joint space narrowing of ≥ 1.0 mm or incident clinical knee OA (ACR criteria). After 2.5 years of follow-up, forty-three women (11%) were lost to follow-up and for a further 72 women no DNA was available. DNA-variants: We studied DNA variants that have been reported as being associated with knee OA at genome-wide significance level (P<5x10-8) in women. These included variants in or near: GDF5 (rs143383), GNL3 (rs11177) and DUS4L (also referred to as the Chr 7 locus, rs4730250) and MCF2L (rs11842874). The assay for the MCF2L variant failed due to technical problems. We subsequently computed a genetic risk score for the 3 variants that were genotyped successfully, where the number of carried risk alleles where added per person. Association between the genetic markers, the genetic risk score and incident knee OA was tested by multivariate logistic regression analysis, adjusting for age and BMI. Results: In total 92 (31%) women developed OA during the 2.5 year follow-up time. When the risk for the separate DNA variants were tested, for all three variants we found increased risk for incident knee OA with the previously identified risk allele (OR(95% CI): GDF5: 1.30 (0.91-1.85); GNL3: 1.23(0.86-1.78); Chr7: 1.75(1.10-2.77)). All these risks were at least twice as large as compared to those previously reported in large meta-analysis. However, only the odds ratio for the chr 7 locus reached nominally significance (p=0.018). The genetic risk score, which was composed of the three SNPs under study, was significantly associated with incident knee OA (p=0.007). With each additional risk allele, the women had 38% more chance for incident knee OA. When we stratified the women into 3 groups according to their genetic risk score, we observed that women with 4 or 5 genetic risk alleles had almost 3.5 times higher risk for incident knee OA compared to women that had 0 or 1 risk allele (OR 3.45 (95%CI 1.41-8.47), p=0.007). Conclusions: The previously identified genetic risk markers for knee OA, were significantly associated with incident knee OA in this high risk group of overweight and obese women. The effect sizes of the studies DNA variants were larger than those reported previously. These results indicate that genetic prediction of OA can be dependent on the presence of other strong clinical risk factors

Genetics of osteoarthritis

Epidemiological studies have demonstrated a major genetic component to osteoarthritis (OA), with heritability estimates of over 50% for most joint sites. These studies have also highlighted differences in the degree of OA heritability between joint sites and between the sexes, implying a high level of heterogeneity. We published a genome-wide scan in 1999. We had focused on families containing siblings with severe large-joint OA ascertained by joint-replacement surgery. Our data showed that OA genetic susceptibility did exhibit joint specificity and that this susceptibility had a greater role in female disease. During the past 5 years we have been investigating our linkage regions and we have so far identified FRZB (chromosome 2q32.1), COL9A1 (6q12-q13), BMP5 (6p12.1) and IL4R (16p12.1-p11.2) as encoding for OA susceptibility. Common variants at these genes affect either the structural properties of the protein (FRZB and IL4R) or the transcription of the gene (BMP5 and COL9A1). The variants are particularly relevant to the development of hip OA in females. What is particularly interesting about our recent discoveries is that the proteins encoded for by IL4R, BMP5 and FRZB are involved in chondrocyte cell signalling and signal transduction pathways. It appears probable, therefore, that OA genetic risk for the hip is principally accounted for by aberrant cell signalling. This was not anticipated. In this presentation I will focus on our latest genetic findings. I will also discuss the results from other studies. A number of OA genome-wide scans have been performed, some on large joints and others investigating hand disease. Many loci appear unique to one particular study, with only some loci being positive in multiple studies. The reasons for this will be discussed. A concern often expressed at orthopaedic and rheumatology meetings is that genetic linkages and associations are not consistently reproduced. These are reasonable criticisms but it needs to be remembered that the genetic component of a complex trait such as OA will not be mediated by fully penetrant risk alleles. Instead, any one allele will contribute only a fraction of the overall risk and this allele will, by chance, have varying frequencies in different cohorts. It is unreasonable, therefore, to expect a linkage or an association in one cohort to be replicated in all cohorts.