Differential glucocorticoid metabolism in patients with persistent versus resolving inflammatory arthritis

Introduction: Impairment in the ability of the inflamed synovium to generate cortisol has been proposed to be a factor in the persistence and severity of inflammatory arthritis. In the inflamed synovium, cortisol is generated from cortisone by the 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) enzyme. The objective of this study was to determine the role of endogenous glucocorticoid metabolism in the development of persistent inflammatory arthritis. Methods: Urine samples were collected from patients with early arthritis (symptoms ≤12 weeks duration) whose final diagnostic outcomes were established after clinical follow-up and from patients with established rheumatoid arthritis (RA). All patients were free of disease-modifying anti-rheumatic drugs at the time of sample collection. Systemic measures of glucocorticoid metabolism were assessed in the urine samples by gas chromatography/mass spectrometry. Clinical data including CRP and ESR were also collected at baseline. Results: Systemic measures of 11β-HSD1 activity were significantly higher in patients with early arthritis whose disease went on to persist, and also in the subgroup of patients with persistent disease who developed RA, when compared with patients whose synovitis resolved over time. We observed a significant positive correlation between systemic 11β-HSD1 activity and ESR/CRP in patients with established RA but not in any of the early arthritis patients group. Conclusions: The present study demonstrates that patients with a new onset of synovitis whose disease subsequently resolved had significantly lower levels of systemic 11β-HSD1 activity when compared with patients whose synovitis developed into RA or other forms of persistent arthritis. Low absolute levels of 11β-HSD1 activity do not therefore appear to be a major contributor to the development of RA and it is possible that a high total body 11β-HSD1 activity during early arthritis may reduce the probability of disease resolution

The role of extracellular vesicle miRNAs and tRNAs in synovial fibroblast senescence

Extracellular vesicles are mediators of intercellular communication with critical roles in cellular senescence and ageing. In arthritis, senescence is linked to the activation of a pro-inflammatory phenotype contributing to chronic arthritis pathogenesis. We hypothesised that senescent osteoarthritic synovial fibroblasts induce senescence and a pro-inflammatory phenotype in non-senescent osteoarthritic fibroblasts, mediated through extracellular vesicle cargo. Small RNA-sequencing and mass spectrometry proteomics were performed on extracellular vesicles isolated from the secretome of non-senescent and irradiation-induced senescent synovial fibroblasts. β-galactosidase staining confirmed senescence in SFs. RNA sequencing identified 17 differentially expressed miRNAs, 11 lncRNAs, 14 tRNAs and one snoRNA and, 21 differentially abundant proteins were identified by mass spectrometry. Bioinformatics analysis of miRNAs identified fibrosis, cell proliferation, autophagy, and cell cycle as significant pathways, tRNA analysis was enriched for signaling pathways including FGF, PI3K/AKT and MAPK, whilst protein analysis identified PAX3-FOXO1, MYC and TFGB1 as enriched upstream regulators involved in senescence and cell cycle arrest. Finally, treatment of non-senescent synovial fibroblasts with senescent extracellular vesicles confirmed the bystander effect, inducing senescence in non-senescent cells potentially through down regulation of NF-κβ and cAMP response element signaling pathways thus supporting our hypothesis. Understanding the exact composition of EV-derived small RNAs of senescent cells in this way will inform our understanding of their roles in inflammation, intercellular communication, and as active molecules in the senescence bystander effect

Polygenic basis and biomedical consequences of telomere length variation.

Funder: Health Data Research UK EU/EFPIA Innovative Medicines Initiative Joint Undertaking BigData@Heart (11607).Funder: Health Data Research UKTelomeres, the end fragments of chromosomes, play key roles in cellular proliferation and senescence. Here we characterize the genetic architecture of naturally occurring variation in leukocyte telomere length (LTL) and identify causal links between LTL and biomedical phenotypes in 472,174 well-characterized UK Biobank participants. We identified 197 independent sentinel variants associated with LTL at 138 genomic loci (108 new). Genetically determined differences in LTL were associated with multiple biological traits, ranging from height to bone marrow function, as well as several diseases spanning neoplastic, vascular and inflammatory pathologies. Finally, we estimated that, at the age of 40 years, people with an LTL >1 s.d. shorter than the population mean had a 2.5-year-lower life expectancy compared with the group with ≥1 s.d. longer LDL. Overall, we furnish new insights into the genetic regulation of LTL, reveal wide-ranging influences of LTL on physiological traits, diseases and longevity, and provide a powerful resource available to the global research community

Obesity defined molecular endotypes in the synovium of patients with osteoarthritis provides a rationale for therapeutic targeting of fibroblast subsets

Abstract Background Osteoarthritis (OA), a multifaceted condition, poses a significant challenge for the successful clinical development of therapeutics due to heterogeneity. However, classifying molecular endotypes of OA pathogenesis could provide invaluable phenotype‐directed routes for stratifying subgroups of patients for targeted therapeutics, leading to greater chances of success in trials. This study establishes endotypes in OA soft joint tissue driven by obesity in both load‐bearing and non‐load bearing joints. Methods Hand, hip, knee and foot joint synovial tissue was obtained from OA patients (n = 32) classified as obese (BMI > 30) or normal weight (BMI 18.5–24.9). Isolated fibroblasts (OA SF) were assayed by Olink proteomic panel, seahorse metabolic flux assay, Illumina's NextSeq 500 bulk and Chromium 10X single cell RNA‐sequencing, validated by Luminex and immunofluorescence. Results Targeted proteomic, metabolic and transcriptomic analysis found the inflammatory landscape of OA SFs are independently impacted by obesity, joint loading and anatomical site with significant heterogeneity between obese and normal weight patients, confirmed by bulk RNAseq. Further investigation by single cell RNAseq identified four functional molecular endotypes including obesity specific subsets defined by an inflammatory endotype related to immune cell regulation, fibroblast activation and inflammatory signaling, with up‐regulated CXCL12, CFD and CHI3L1 expression. Luminex confirmed elevated chitase3‐like‐1(229.5 vs. 49.5 ng/ml, p < .05) and inhibin (20.6 vs. 63.8 pg/ml, p < .05) in obese and normal weight OA SFs, respectively. Lastly, we find SF subsets in obese patients spatially localise in sublining and lining layers of OA synovium and can be distinguished by differential expression of the transcriptional regulators MYC and FOS. Conclusion These findings demonstrate the significance of obesity in changing the inflammatory landscape of synovial fibroblasts in both load bearing and non‐load bearing joints. Describing multiple heterogeneous OA SF populations characterised by specific molecular endotypes, which drive heterogeneity in OA disease pathogenesis. These molecular endotypes may provide a route for the stratification of patients in clinical trials, providing a rational for the therapeutic targeting of specific SF subsets in specific patient populations with arthritic conditions

The role of extracellular vesicles in synovial fibroblast senescence

Extracellular vesicles are mediators of intercellular communication with critical roles in cellular senescence and ageing. In arthritis, senescence is linked to the activation of a pro-inflammatory phenotype contributing to chronic arthritis pathogenesis. We hypothesised that senescent osteoarthritic synovial fibroblasts induce senescence and a pro-inflammatory phenotype in non-senescent osteoarthritic fibroblasts, mediated through extracellular vesicle cargo. Small RNA-sequencing and mass spectrometry proteomics were performed on extracellular vesicles isolated from the secretome of non-senescent and irradiation-induced senescent synovial fibroblasts. β-galactosidase staining confirmed senescence in SFs. RNA sequencing identified 17 differentially expressed miRNAs, 11 lncRNAs, 14 tRNAs and one snoRNA and, 21 differentially abundant proteins were identified by mass spectrometry. Bioinformatics analysis of miRNAs identified fibrosis, cell proliferation, autophagy, and cell cycle as significant pathways, tRNA analysis was enriched for signaling pathways including FGF, PI3K/AKT and MAPK, whilst protein analysis identified PAX3-FOXO1, MYC and TFGB1 as enriched upstream regulators involved in senescence and cell cycle arrest. Finally, treatment of non-senescent synovial fibroblasts with senescent extracellular vesicles confirmed the bystander effect, inducing senescence in non-senescent cells potentially through down regulation of NF-κβ and cAMP response element signaling pathways thus supporting our hypothesis. Understanding the exact composition of EV-derived small RNAs of senescent cells in this way will inform our understanding of their roles in inflammation, intercellular communication, and as active molecules in the senescence bystander effect

DataSheet1_The role of extracellular vesicle miRNAs and tRNAs in synovial fibroblast senescence.docx

Extracellular vesicles are mediators of intercellular communication with critical roles in cellular senescence and ageing. In arthritis, senescence is linked to the activation of a pro-inflammatory phenotype contributing to chronic arthritis pathogenesis. We hypothesised that senescent osteoarthritic synovial fibroblasts induce senescence and a pro-inflammatory phenotype in non-senescent osteoarthritic fibroblasts, mediated through extracellular vesicle cargo. Small RNA-sequencing and mass spectrometry proteomics were performed on extracellular vesicles isolated from the secretome of non-senescent and irradiation-induced senescent synovial fibroblasts. β-galactosidase staining confirmed senescence in SFs. RNA sequencing identified 17 differentially expressed miRNAs, 11 lncRNAs, 14 tRNAs and one snoRNA and, 21 differentially abundant proteins were identified by mass spectrometry. Bioinformatics analysis of miRNAs identified fibrosis, cell proliferation, autophagy, and cell cycle as significant pathways, tRNA analysis was enriched for signaling pathways including FGF, PI3K/AKT and MAPK, whilst protein analysis identified PAX3-FOXO1, MYC and TFGB1 as enriched upstream regulators involved in senescence and cell cycle arrest. Finally, treatment of non-senescent synovial fibroblasts with senescent extracellular vesicles confirmed the bystander effect, inducing senescence in non-senescent cells potentially through down regulation of NF-κβ and cAMP response element signaling pathways thus supporting our hypothesis. Understanding the exact composition of EV-derived small RNAs of senescent cells in this way will inform our understanding of their roles in inflammation, intercellular communication, and as active molecules in the senescence bystander effect.</p