Widespread epigenomic, transcriptomic and proteomic differences between hip osteophytic and articular chondrocytes in osteoarthritis

Abstract

ObjectivesTo identify molecular differences between chondrocytes from osteophytic and articular cartilage tissue from OA patients.MethodsWe investigated genes and pathways by combining genome-wide DNA methylation, RNA sequencing and quantitative proteomics in isolated primary chondrocytes from the cartilaginous layer of osteophytes and matched areas of low- and high-grade articular cartilage across nine patients with OA undergoing hip replacement surgery.ResultsChondrocytes from osteophytic cartilage showed widespread differences to low-grade articular cartilage chondrocytes. These differences were similar to, but more pronounced than, differences between chondrocytes from osteophytic and high-grade articular cartilage, and more pronounced than differences between high- and low-grade articular cartilage. We identified 56 genes with significant differences between osteophytic chondrocytes and low-grade articular cartilage chondrocytes on all three omics levels. Several of these genes have known roles in OA, including ALDH1A2 and cartilage oligomeric matrix protein, which have functional genetic variants associated with OA from genome-wide association studies. An integrative gene ontology enrichment analysis showed that differences between osteophytic and low-grade articular cartilage chondrocytes are associated with extracellular matrix organization, skeletal system development, platelet aggregation and regulation of ERK1 and ERK2 cascade.ConclusionWe present a first comprehensive view of the molecular landscape of chondrocytes from osteophytic cartilage as compared with articular cartilage chondrocytes from the same joints in OA. We found robust changes at genes relevant to chondrocyte function, providing insight into biological processes involved in osteophyte development and thus OA progression.

Accelerating functional gene discovery in osteoarthritis

“…We isolated chondrocytes from each cartilage sample as described 113 and detailed in full in Supplementary Methods (see Supplementary Information). Assays and analyses were performed as described in Supplementary Methods (see Supplementary Information), and RNA sequencing was performed on the Illumina HiSeq2000 or Hiseq4000 (75 bp paired-ends), with quality control including FastQC 0.11.5 ( http://www.bioinformatics.babraham.ac.uk/projects/fastqc ).…”

A molecular quantitative trait locus map for osteoarthritis

“…For cohort 3, the extraction of chondrocytes has previously been described 35 in the majority of these samples, with the remaining samples following the same protocol. The protocol was based on that for cohorts 1, 2, 4 and highly similar as described in the following.…”

Decoding the genomic basis of osteoarthritis

“…Full details for confirmation of joint replacement for osteoarthritis and cartilage tissue scoring are listed in Supplementary Methods. We followed previously established protocols to isolate chondrocytes 37,38 , and synoviocytes 39 DNA, RNA and protein extraction DNA, RNA, and protein extraction was carried out using Qiagen AllPrep DNA/RNA/Protein Mini Kit following manufacturer's instructions, with small variations for cohort 3 as previously described 38 . Samples were frozen at -80C (cohorts 1, 2, 4) or -70C (cohort 3) prior to assays.…”