Osteoarthritis (OA) is a chronic musculoskeletal disease characterised by the destruction of articular cartilage, synovial inflammation and bone remodelling. Disease aetiology is complex and highly ...heritable, with genetic variation estimated to contribute to 50% of OA occurrence. Epigenetic alterations, including DNA methylation changes, have also been implicated in OA pathophysiology. This review examines what genetic and DNA methylation studies have taught us about the genes and pathways involved in OA pathology. The influence of DNA methylation on the molecular mechanisms underlying OA genetic risk and the consequence of this interaction on disease susceptibility and penetrance are also discussed.
GDF5
encodes an extracellular signalling molecule that is essential for normal skeletal development. The rs144383 C to T SNP located in the 5ʹUTR of this gene is functional and has a pleiotropic ...effect on the musculoskeletal system, being a risk factor for knee-osteoarthritis (OA), congenital hip dysplasia, lumbar disc degeneration and Achilles tendon pathology. rs143383 exerts a joint-wide effect on
GDF5
expression, with expression of the OA-associated T allele being significantly reduced relative to the C allele, termed allelic expression imbalance. We have previously reported that the
GDF5
locus is subject to DNA methylation and that allelic imbalance of rs143383 is mediated by SP1, SP3 and DEAF1 transcriptional repressors. In this study, we have assayed
GDF5
methylation in normal and osteoarthritic cartilage, and investigated the effect of methylation on the allelic imbalance of rs143383. We observed demethylation of the
GDF5
5ʹUTR in OA knee cartilage relative to both OA (
p
= 0.009) and non-OA (
p
= 0.001) hip cartilage, with the most significant demethylation observed at the highly conserved +37 CpG site located 4 bp upstream of rs143383. Methylation modulates the level and direction of allelic imbalance of rs143383, with methylation of the +37 CpG dinucleotide within the SP1/SP3 binding site having an allele-specific effect on SP1 and SP3 binding. Furthermore, methylation attenuated the repressive effects of SP1, SP3 and DEAF1 on
GDF5
promoter activity. This data suggest that the differential methylation of the +37 CpG site between osteoarthritic hip and knee cartilage may be responsible for the knee-specific effect of rs143383 on OA susceptibility.
Objective
The aim of this study was to characterize the genome‐wide DNA methylation profile of chondrocytes from knee and hip cartilage obtained from patients with osteoarthritis (OA) and hip ...cartilage obtained from patients with femoral neck fracture, providing the first comparison of DNA methylation between OA and non‐OA hip cartilage, and between OA hip and OA knee cartilage.
Methods
The study was performed using the Illumina Infinium HumanMethylation450 BeadChip array, which allows the annotation of ∼480,000 CpG sites. Genome‐wide methylation was assessed in chondrocyte DNA extracted from 23 hip OA patients, 73 knee OA patients, and 21 healthy hip control patients with femoral neck fracture.
Results
Analysis revealed that chondrocytes from the hip cartilage of OA patients and healthy controls have unique methylation profiles, with 5,322 differentially methylated loci (DMLs) identified between the 2 groups. In addition, a comparison between hip and knee OA chondrocytes revealed 5,547 DMLs between the 2 groups, including DMLs in several genes known to be involved in the pathogenesis of OA. Hip OA samples were found to cluster into 2 groups. A total of 15,239 DMLs were identified between the 2 clusters, with an enrichment of genes involved in inflammation and immunity. Similarly, we confirmed a previous report of knee OA samples that also clustered into 2 groups.
Conclusion
We demonstrated that global DNA methylation using a high‐density array can be a powerful tool in the characterization of OA at the molecular level. Identification of pathways enriched in DMLs between OA and OA‐free cartilage highlight potential etiologic mechanisms that are involved in the initiation and/or progression of the disease and that could be therapeutically targeted.
Osteoarthritis (OA) is a common disease of older individuals that impacts detrimentally on the quality and the length of life. It is characterised by the painful loss of articular cartilage and is ...polygenic and multifactorial. Genome-wide association scans have highlighted over 90 osteoarthritis genetic signals, some of which reside within or close to highly plausible candidate genes. An example is an association to polymorphisms within and adjacent to the matrix Gla protein gene MGP. We set out to undertake a functional study of this gene.
Nucleic acid was extracted from cartilage, infrapatellar fat pad, synovium, trabecular bone, trapezium and peripheral whole blood from OA patients and also from mesenchymal stem cells (MSCs) subjected to chondrogenesis. Expression of MGP was measured by quantitative PCR (qPCR), RNA-sequencing and allelic expression imbalance (AEI) analysis. Matrix Gla protein was depleted in chondrocytes by knocking down MGP expression using RNA interference (RNAi) and the effect on a range of genes assessed by qPCR.
MGP is expressed in joint tissues, blood and chondrocytes cultured from MSCs. There is a higher expression in diseased versus non-diseased cartilage. Polymorphisms that are associated with OA also correlate with the expression of MGP, with the OA risk-conferring allele showing significantly reduced expression in cartilage, fat pad and synovium but increased expression in blood. Depletion of Matrix Gla protein had a significant effect on the majority of genes tested, with an increased expression of catabolic genes that encode enzymes that degrade cartilage.
MGP expression is subject to cis-acting regulators that correlate with the OA association signal. These are active in a range of joint tissues but have effects which are particularly strong in cartilage. An opposite effect is observed in blood, highlighting the context-specific nature of the regulation of this gene's expression. Recapitulation of the genetic deficit in cartilage chondrocytes is pro-catabolic.
Bayesian networks can be used to identify possible causal relationships between variables based on their conditional dependencies and independencies, which can be particularly useful in complex ...biological scenarios with many measured variables. Here we propose two improvements to an existing method for Bayesian network analysis, designed to increase the power to detect potential causal relationships between variables (including potentially a mixture of both discrete and continuous variables). Our first improvement relates to the treatment of missing data. When there is missing data, the standard approach is to remove every individual with any missing data before performing analysis. This can be wasteful and undesirable when there are many individuals with missing data, perhaps with only one or a few variables missing. This motivates the use of imputation. We present a new imputation method that uses a version of nearest neighbour imputation, whereby missing data from one individual is replaced with data from another individual, their nearest neighbour. For each individual with missing data, the subsets of variables to be used to select the nearest neighbour are chosen by sampling without replacement the complete data and estimating a best fit Bayesian network. We show that this approach leads to marked improvements in the recall and precision of directed edges in the final network identified, and we illustrate the approach through application to data from a recent study investigating the causal relationship between methylation and gene expression in early inflammatory arthritis patients. We also describe a second improvement in the form of a pseudo-Bayesian approach for upweighting certain network edges, which can be useful when there is prior evidence concerning their directions.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Genetic studies have revealed that most loci associated with osteoarthritis (OA) show ethnic stratification, with limited overlap between Asian and European populations. Consequently, such studies ...have often focused on particular ethnic groups, with those performed in European cohorts yielding the most replicated associations. As for other common diseases, the OA susceptibility loci mapped to date account for only a fraction of disease heritability. Nevertheless, analysis of these loci could identify biological pathways related to OA pathogenesis. Such an approach is taken in this Review and provides valuable insights into OA aetiology. For example, several of the loci associated with OA contain genes encoding key regulators of skeletogenesis and endochondral ossification. Furthermore, direct and indirect regulation of gene transcription is highlighted as an important factor in this disease. Interestingly, genes encoding structural proteins of the cartilage extracellular matrix do not seem to be a repository for OA susceptibility. Therefore, susceptibility might operate at a regulatory rather than a structural level, which is a potentially promising finding, as the activities of regulators are amenable to therapeutic modulation. Greater clarity will emerge as more association signals are identified; nonetheless, patterns of aetiology are clearly discernible, from a molecular perspective, even with the relatively small number currently available.
The human syndrome of dendritic cell, monocyte, B and natural killer lymphoid deficiency presents as a sporadic or autosomal dominant trait causing susceptibility to mycobacterial and other ...infections, predisposition to myelodysplasia and leukemia, and, in some cases, pulmonary alveolar proteinosis. Seeking a genetic cause, we sequenced the exomes of 4 unrelated persons, 3 with sporadic disease, looking for novel, heterozygous, and probably deleterious variants. A number of genes harbored novel variants in person, but only one gene, GATA2, was mutated in all 4 persons. Each person harbored a different mutation, but all were predicted to be highly deleterious and to cause loss or mutation of the C-terminal zinc finger domain. Because GATA2 is the only common mutated gene in 4 unrelated persons, it is highly probable to be the cause of dendritic cell, monocyte, B, and natural killer lymphoid deficiency. This disorder therefore constitutes a new genetic form of heritable immunodeficiency and leukemic transformation.
Abstract Osteoarthritis (OA) is a common age-related disease that affects the tissues of the synovial joint, leading to loss of function and pain. It impacts on both patient morbidity and mortality. ...It is a complex, polygenic disease that lacks any large-effect susceptibility loci. Instead, OA susceptibility alleles individually contribute only modestly to the overall disease risk, making their identification challenging. Despite this, breakthroughs have occurred with compelling associations so far reported to polymorphisms within the genes GDF5 and MCF2L and to the genomic region 7q22. The latter two have emerged from genome-wide association scans, which are likely to yield more hits in the near future. As for many complex diseases, it is now apparent that epigenetic effects are also important mediators of disease biology, with DNA methylation, histone modifications and non-coding RNAs all having a role. At present, much of the epigenetic focus has been on cartilage, the tissue at the center of the OA disease process. If we are to get close to a qualitative and quantitative understanding of the impact of epigenetics on OA, then in future the other tissues of the joint will also need to be investigated. One of the more exciting insights to have emerged recently is the fact that epigenetic effects can impact on OA genetic effects and this may be a particularly fruitful avenue for integrating both as we move toward a clearer understanding of the pathophysiology of this intriguing disease.
Objective
To identify the functional single‐nucleotide polymorphisms (SNPs) and mechanisms conferring increased risk of hand osteoarthritis (OA) at the ALDH1A2 locus, which is a retinoic acid ...regulatory gene.
Methods
Tissue samples from 247 patients with knee, hip, or hand OA who had undergone joint surgery were included. RNA‐sequencing analysis was used to investigate differential expression of ALDH1A2 and other retinoic acid signaling pathway genes in cartilage. Expression of ALDH1A2 in joint tissues obtained from multiple sites was quantified using quantitative reverse transcription–polymerase chain reaction. Allelic expression imbalance (AEI) was measured by pyrosequencing. The consequences of ALDH1A2 depletion by RNA interference were assessed in primary human chondrocytes. In silico and in vitro analyses were used to pinpoint which, among 62 highly correlated SNPs, could account for the association at the locus.
Results
ALDH1A2 expression was observed across multiple joint tissue samples, including osteochondral tissue from the hand. The expression of ALDH1A2 and of several retinoic acid signaling genes was different in diseased cartilage compared to non‐diseased cartilage, with ALDH1A2 showing lower levels in OA cartilage. Experimental depletion of ALDH1A2 resulted in changes in the expression levels of a number of chondrogenic markers, including SOX9. In addition, reduced expression of the OA risk–conferring allele was witnessed in a number of joint tissues, with the strongest effect in cartilage. The intronic SNP rs12915901 recapitulated the AEI observed in patient tissues, while the Ets transcription factors were identified as potential mediators of this effect.
Conclusion
The ALDH1A2 locus seems to increase the risk of hand OA through decreased expression of ALDH1A2 in joint tissues, with the effect dependent on rs12915901. These findings indicate a mechanism that may now be targeted to modulate OA risk.
Recent genome-wide association scans (GWASs) along with several adequately powered candidate gene studies have yielded a number of risk alleles for osteoarthritis (OA). This number is now ...sufficiently large to allow conclusions to be drawn regarding the nature of genetic susceptibility, including the fact that the risk alleles have variable effects depending on sex, ethnicity and on the skeletal site of the disease. Several of the alleles that have emerged from the GWASs are within or close to highly plausible candidate genes, including RUNX2 and CHST11. However, the majority of risk alleles do not map to genes previously reported to play a role in musculoskeletal biology, indicating that the GWAS datasets are telling us something new about the OA disease process. Functional studies have so far revealed that effects on gene expression are likely to be one of the main mechanisms through which OA susceptibility is acting. Epigenetic mechanisms such as DNA methylation also influence OA risk, and integration of genetic, transcriptomic and epigenetic data will allow us to use the genetic discoveries for informed development of new OA biological treatments.
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