The inflammatory speech of fibroblasts.

Activation of fibroblasts is a key event during normal tissue repair after injury and the dysregulated repair processes that result in organ fibrosis. To most researchers, fibroblasts are rather unremarkable spindle-shaped cells embedded in the fibrous collagen matrix of connective tissues and/or deemed useful to perform mechanistic studies with adherent cells in culture. For more than a century, fibroblasts escaped thorough classification due to the lack of specific markers and were treated as the leftovers after all other cells have been identified from a tissue sample. With novel cell lineage tracing and single cell transcriptomics tools, bona fide fibroblasts emerge as only one heterogeneous sub-population of a much larger group of partly overlapping cell types, including mesenchymal stromal cells, fibro-adipogenic progenitor cells, pericytes, and/or perivascular cells. All these cells are activated to contribute to tissue repair after injury and/or chronic inflammation. "Activation" can entail various functions, such as enhanced proliferation, migration, instruction of inflammatory cells, secretion of extracellular matrix proteins and organizing enzymes, and acquisition of a contractile myofibroblast phenotype. We provide our view on the fibroblastic cell types and activation states playing a role during physiological and pathological repair and their crosstalk with inflammatory macrophages. Inflammation and fibrosis of the articular synovium during rheumatoid arthritis and osteoarthritis are used as specific examples to discuss inflammatory fibroblast phenotypes. Ultimately, delineating the precursors and functional roles of activated fibroblastic cells will contribute to better and more specific intervention strategies to treat fibroproliferative and fibrocontractive disorders.

Cell and transcriptomic diversity of infrapatellar fat pad during knee osteoarthritis.

OBJECTIVES: In this study, we employ a multiomic approach to identify major cell types and subsets, and their transcriptomic profiles within the infrapatellar fat pad (IFP), and to determine differences in the IFP based on knee osteoarthritis (KOA), sex and obesity status. METHODS: Single-nucleus RNA sequencing of 82 924 nuclei from 21 IFPs (n=6 healthy control and n=15 KOA donors), spatial transcriptomics and bioinformatic analyses were used to identify contributions of the IFP to KOA. We mapped cell subclusters from other white adipose tissues using publicly available literature. The diversity of fibroblasts within the IFP was investigated by bioinformatic analyses, comparing by KOA, sex and obesity status. Metabolomics was used to further explore differences in fibroblasts by obesity status. RESULTS: We identified multiple subclusters of fibroblasts, macrophages, adipocytes and endothelial cells with unique transcriptomic profiles. Using spatial transcriptomics, we resolved distributions of cell types and their transcriptomic profiles and computationally identified putative cell-cell communication networks. Furthermore, we identified transcriptomic differences in fibroblasts from KOA versus healthy control donor IFPs, female versus male KOA-IFPs and obese versus normal body mass index (BMI) KOA-IFPs. Finally, using metabolomics, we defined differences in metabolite levels in supernatants of naïve, profibrotic stimuli-treated and proinflammatory stimuli-treated fibroblasts from obese compared to normal BMI KOA-IFPs. CONCLUSIONS: Overall, by employing a multiomic approach, this study provides the first comprehensive map of the cellular and transcriptomic diversity of human IFP and identifies IFP fibroblasts as key cells contributing to transcriptomic and metabolic differences related to KOA disease, sex or obesity.

Association of synovial fluid and urinary C2C-HUSA levels with surgical outcomes post-total knee arthroplasty.

OBJECTIVES: After total knee arthroplasty (TKA), ∼30% of knee osteoarthritis (KOA) patients show little symptomatic improvement. Earlier studies have correlated urinary (u) type 2 collagen C terminal cleavage peptide assay (C2C-HUSA), which detects a fragment of cartilage collagen breakdown, with KOA progression. This study determines whether C2C levels in urine, synovial fluid, or their ratio, are associated with post-surgical outcomes. METHODS: From a large sample of 489 subjects, diagnosed with primary KOA undergoing TKA, Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) pain and function scores were collected at baseline (time of surgery) and one-year post-TKA. Baseline urine (u) and synovial fluid (sf) were analysed using the IBEX-C2C-HUSA assay, with higher values indicating higher amounts of cartilage degradation. For urine, results were normalised to creatinine. Furthermore, subjects' changes in WOMAC scores were categorised based on percent reduction in pain or improvement in function, compared to baseline, such that >66.7%, >33.3 to ≤66.7%, and ≤33.3% denoted "strong", "moderate" and "mild/worse" responses, respectively. Associations of individual biofluid C2C-HUSA levels, or their ratio, with change in WOMAC pain and function scores up to one-year post-TKA, or category of change, were analysed by linear, logistic, or cumulative odds models. RESULTS: Higher baseline uC2C-HUSA levels or a lower ratio of baseline sfC2C-HUSA to uC2C-HUSA were associated with improvements in WOMAC pain by linear multivariable modelling [odds ratio -0.40 (95% confidence interval -0.76, -0.05) p = 0.03; 0.36 (0.01, 0.71), p = 0.04, respectively], while sfC2C-HUSA alone was not. However, lower ratios of sfC2C-HUSA to uC2C-HUSA were associated with improvements in WOMAC function [1.37 (0.18, 2.55), p = 0.02], while sfC2C-HUSA and uC2C-HUSA alone were not. Lower ratios of sfC2C-HUSA to uC2C-HUSA were also associated with an increased likelihood of a subject being categorised in a group where TKA was beneficial in both univariable [pain, 0.81 (0.68, 0.96), p = 0.02; function, 0.92 (0.85, 0.99), p = 0.035] and multivariable [pain, 0.81 (0.68, 0.97) p = 0.02; function, 0.92 (0.85, 1.00), p = 0.043] ordinal modelling, while sfC2C-HUSA and uC2C-HUSA alone were not. CONCLUSIONS: Overall, ratios of baseline sfC2C-HUSA to uC2C-HUSA, and baseline uC2C-HUSA, may play an important role in studying post-TKA surgical outcomes.

Three decades of advancements in osteoarthritis research: insights from transcriptomic, proteomic, and metabolomic studies.

OBJECTIVE: Osteoarthritis (OA) is a complex disease involving contributions from both local joint tissues and systemic sources. Patient characteristics, encompassing sociodemographic and clinical variables, are intricately linked with OA rendering its understanding challenging. Technological advancements have allowed for a comprehensive analysis of transcripts, proteomes and metabolomes in OA tissues/fluids through omic analyses. The objective of this review is to highlight the advancements achieved by omic studies in enhancing our understanding of OA pathogenesis over the last three decades. DESIGN: We conducted an extensive literature search focusing on transcriptomics, proteomics and metabolomics within the context of OA. Specifically, we explore how these technologies have identified individual transcripts, proteins, and metabolites, as well as distinctive endotype signatures from various body tissues or fluids of OA patients, including insights at the single-cell level, to advance our understanding of this highly complex disease. RESULTS: Omic studies reveal the description of numerous individual molecules and molecular patterns within OA-associated tissues and fluids. This includes the identification of specific cell (sub)types and associated pathways that contribute to disease mechanisms. However, there remains a necessity to further advance these technologies to delineate the spatial organization of cellular subtypes and molecular patterns within OA-afflicted tissues. CONCLUSIONS: Leveraging a multi-omics approach that integrates datasets from diverse molecular detection technologies, combined with patients' clinical and sociodemographic features, and molecular and regulatory networks, holds promise for identifying unique patient endophenotypes. This holistic approach can illuminate the heterogeneity among OA patients and, in turn, facilitate the development of tailored therapeutic interventions.

Evolution and advancements in genomics and epigenomics in OA research: How far we have come.

OBJECTIVE: Osteoarthritis (OA) is the most prevalent musculoskeletal disease affecting articulating joint tissues, resulting in local and systemic changes that contribute to increased pain and reduced function. Diverse technological advancements have culminated in the advent of high throughput "omic" technologies, enabling identification of comprehensive changes in molecular mediators associated with the disease. Amongst these technologies, genomics and epigenomics - including methylomics and miRNomics, have emerged as important tools to aid our biological understanding of disease. DESIGN: In this narrative review, we selected articles discussing advancements and applications of these technologies to OA biology and pathology. We discuss how genomics, deoxyribonucleic acid (DNA) methylomics, and miRNomics have uncovered disease-related molecular markers in the local and systemic tissues or fluids of OA patients. RESULTS: Genomics investigations into the genetic links of OA, including using genome-wide association studies, have evolved to identify 100+ genetic susceptibility markers of OA. Epigenomic investigations of gene methylation status have identified the importance of methylation to OA-related catabolic gene expression. Furthermore, miRNomic studies have identified key microRNA signatures in various tissues and fluids related to OA disease. CONCLUSIONS: Sharing of standardized, well-annotated omic datasets in curated repositories will be key to enhancing statistical power to detect smaller and targetable changes in the biological signatures underlying OA pathogenesis. Additionally, continued technological developments and analysis methods, including using computational molecular and regulatory networks, are likely to facilitate improved detection of disease-relevant targets, in-turn, supporting precision medicine approaches and new treatment strategies for OA.

Early microRNA and metabolite changes after anterior cruciate ligament reconstruction surgery.

OBJECTIVES: Anterior cruciate ligament (ACL) reconstruction after injury does not prevent post-traumatic osteoarthritis (PTOA). Circulating microRNA (miRNA) and metabolite changes emerging shortly after ACL injury and reconstruction remain insufficiently defined, potentially harbouring early cues contributing to PTOA evolution. Moreover, their differential expression between females and males also may influence PTOA's natural trajectory. This study aims to determine alterations in plasma miRNA and metabolite levels in the early stages following ACL reconstruction and between females and males. METHODS: A cohort of 43 ACL reconstruction patients was examined. Plasma was obtained at baseline, 2 weeks, and 6 weeks post-surgery (129 biospecimens in total). High-throughput miRNA sequencing and metabolomics were conducted. Differentially expressed miRNAs and metabolites were identified using negative binomial and linear regression models, respectively. Associations between miRNAs and metabolites were explored using time and sex as co-variants, (pre-surgery versus 2 and 6 weeks post-surgery). Using computational biology, miRNA-metabolite-gene interaction and pathway analyses were performed. RESULTS: Levels of 46 miRNAs were increased at 2 weeks post-surgery compared to pre-surgery (baseline) using miRNA sequencing. Levels of 13 metabolites were significantly increased while levels of 6 metabolites were significantly decreased at 2 weeks compared to baseline using metabolomics. Hsa-miR-145-5p levels were increased in female subjects at both 2 weeks (log2-fold-change 0.71, 95%CI 0.22,1.20) and 6 weeks (log2-fold-change 0.75, 95%CI 0.07,1.43) post-surgery compared to males. In addition, hsa-miR-497-5p showed increased levels in females at 2 weeks (log2-fold-change 0.77, 95%CI 0.06,1.48) and hsa-miR-143-5p at 6 weeks (log2-fold-change 0.83, 95%CI 0.07,1.59). Five metabolites were decreased at 2 weeks post-surgery in females compared to males: L-leucine (-1.44, 95%CI -1.75,-1.13), g-guanidinobutyrate (-1.27, 95%CI 1.54,-0.99), creatinine (-1.17, 95%CI -1.44,-0.90), 2-methylbutyrylcarnitine (-1.76, 95%CI -2.17,-1.35), and leu-pro (-1.13, 95%CI -1.44,-0.83). MiRNA-metabolite-gene interaction analysis revealed key signalling pathways based on post-surgical time-point and in females versus males. CONCLUSION: MiRNA and metabolite profiles were modified by time and by sex early after ACL reconstruction surgery, which could influence surgical response and ultimately risk of developing PTOA.

Individual participant data meta-analysis of metabolomics on sustained knee pain in primary osteoarthritis patients.

OBJECTIVES: Knee pain is the major driver for OA patients to seek healthcare, but after pursuing both conservative and surgical pain interventions, ∼20% of patients continue to report long-term pain following total knee arthroplasty (TKA). This study aimed to identify a metabolomic signature for sustained knee pain after TKA to elucidate possible underlying mechanisms. METHODS: Two independent cohorts from St John's, NL, Canada (n = 430), and Toronto, ON, Canada (n = 495) were included in the study. Sustained knee pain was assessed using the WOMAC pain subscale (five questions) at least 1 year after TKA for primary OA. Those reporting any pain on all five questions were considered to have sustained knee pain. Metabolomic profiling was performed on fasted pre-operative plasma samples using the Biocrates Absolute IDQ p180 kit. Associations between metabolites and pair-wise metabolite ratios with sustained knee pain in each individual cohort were assessed using logistic regression with adjustment for age, sex and BMI. Random-effects meta-analysis using inverse variance as weights was performed on summary statistics from both cohorts. RESULTS: One metabolite, phosphatidylcholine (PC) diacyl (aa) C28:1 (odds ratio = 0.66, P = 0.00026), and three metabolite ratios, PC aa C32:0 to PC aa C28:1, PC aa C28:1 to PC aa C32:0, and tetradecadienylcarnitine (C14:2) to sphingomyelin C20:2 (odds ratios = 1.59, 0.60 and 1.59, respectively; all P < 2 × 10-5), were significantly associated with sustained knee pain. CONCLUSIONS: Though further investigations are needed, our results provide potential predictive biomarkers and drug targets that could serve as a marker for poor response and be modified pre-operatively to improve knee pain and surgical response to TKA.

Anti-fibrotic mechanisms of exogenously-expanded mesenchymal stromal cells for fibrotic diseases.

Most chronic diseases involving inflammation have a fibrotic component that involves remodeling and excess accumulation of extracellular matrix components. Left unchecked, fibrosis leads to organ failure and death. Mesenchymal stromal cells (MSCs) are emerging as a potent cell-based therapy for a wide spectrum of fibrotic conditions due to their immunomodulatory, anti-inflammatory and anti-fibrotic properties. This review provides an overview of known mechanisms by which MSCs mediate their anti-fibrotic actions and in relation to animal models of pulmonary, liver, renal and cardiac fibrosis. Recent MSC clinical trials results in liver, lung, skin, kidney and hearts are discussed and next steps for future MSC-based therapies including pre-activated or genetically-modified cells, or extracellular vesicles are also considered.

Understanding osteoarthritis pathogenesis: a multiomics system-based approach.

PURPOSE OF REVIEW: Osteoarthritis is a heterogeneous, multifactorial condition regulated by complex biological interactions at multiple levels. Comprehensive understanding of these regulatory interactions is required to develop feasible advances to improve patient outcomes. Improvements in technology have made extensive genomic, transcriptomic, epigenomic, proteomic, and metabolomic profiling possible. This review summarizes findings over the past 20 months related to omics technologies in osteoarthritis and examines how using a multiomics approach is necessary for advancing our understanding of osteoarthritis as a disease to improve precision osteoarthritis treatments. RECENT FINDINGS: Using the search terms 'genomics' or 'transcriptomics' or 'epigenomics' or 'proteomics' or 'metabolomics' and 'osteoarthritis' from January 1, 2018 to August 31, 2019, we identified advances in omics approaches applied to osteoarthritis. Trends include untargeted whole genome, transcriptome, proteome, and metabolome analyses leading to identification of novel molecular signatures, cell subpopulations and multiomics validation approaches. SUMMARY: To address the complexity of osteoarthritis, integration of multitissue analyses by multiomics approaches with the inclusion of longitudinal clinical data is necessary for a comprehensive understanding of the disease process, and for appropriate development of efficacious diagnostics, prognostics, and biotherapeutics.

microRNA-181a-5p antisense oligonucleotides attenuate osteoarthritis in facet and knee joints.

OBJECTIVES: We recently identified microRNA-181a-5p (miR-181a-5p) as a critical mediator involved in the destruction of lumbar facet joint (FJ) cartilage. In this study, we tested if locked nucleic acid (LNA) miR-181a-5p antisense oligonucleotides (ASO) could be used as a therapeutic to limit articular cartilage degeneration. METHODS: We used a variety of experimental models consisting of both human samples and animal models of FJ and knee osteoarthritis (OA) to test the effects of LNA-miR-181a-5p ASO on articular cartilage degeneration. Histopathological analysis including immunohistochemistry and in situ hybridisation were used to detect key OA catabolic markers and microRNA, respectively. Apoptotic/cell death markers were evaluated by flow cytometry. qPCR and immunoblotting were applied to quantify gene and protein expression. RESULTS: miR-181a-5p expression was increased in human FJ OA and knee OA cartilage as well as injury-induced FJ OA (rat) and trauma-induced knee OA (mouse) cartilage compared with control cartilage, correlating with classical OA catabolic markers in human, rat and mouse cartilage. We demonstrated that LNA-miR-181a-5p ASO in rat and mouse chondrocytes reduced the expression of cartilage catabolic and chondrocyte apoptotic/cell death markers in vitro. Treatment of OA-induced rat FJ or mouse knee joints with intra-articular injections of in vivo grade LNA-miR-181a-5p ASO attenuated cartilage destruction, and the expression of catabolic, hypertrophic, apoptotic/cell death and type II collagen breakdown markers. Finally, treatment of LNA-miR-181a-5p ASO in cultures of human knee OA chondrocytes (in vitro) and cartilage explants (ex vivo) further demonstrated its cartilage protective effects. CONCLUSIONS: Our data demonstrate, for the first time, that LNA-miR-181a-5p ASO exhibit cartilage-protective effects in FJ and knee OA.

Ephrins and Eph Receptor Signaling in Tissue Repair and Fibrosis.

PURPOSE OF REVIEW: Fibrosis is a pathological feature of many human diseases that affect multiple organs. The development of anti-fibrotic therapies has been a difficult endeavor due to the complexity of signaling pathways associated with fibrogenic processes, complicating the identification and modulation of specific targets. Evidence suggests that ephrin ligands and Eph receptors are crucial signaling molecules that contribute to physiological wound repair and the development of tissue fibrosis. Here, we discuss recent advances in the understanding of ephrin and Eph signaling in tissue repair and fibrosis. RECENT FINDINGS: Ephrin-B2 is implicated in fibrosis of multiple organs. Intercepting its signaling may help counteract fibrosis. Ephrins and Eph receptors are candidate mediators of fibrosis. Ephrin-B2, in particular, promotes fibrogenic processes in multiple organs. Thus, therapeutic strategies targeting Ephrin-B2 signaling could yield new ways to treat organ fibrosis.

Specification of chondrocytes and cartilage tissues from embryonic stem cells.

Osteoarthritis primarily affects the articular cartilage of synovial joints. Cell and/or cartilage replacement is a promising therapy, provided there is access to appropriate tissue and sufficient numbers of articular chondrocytes. Embryonic stem cells (ESCs) represent a potentially unlimited source of chondrocytes and tissues as they can generate a broad spectrum of cell types under appropriate conditions in vitro. Here, we demonstrate that mouse ESC-derived chondrogenic mesoderm arises from a Flk-1(-)/Pdgfrα(+) (F(-)P(+)) population that emerges in a defined temporal pattern following the development of an early cardiogenic F(-)P(+) population. Specification of the late-arising F(-)P(+) population with BMP4 generated a highly enriched population of chondrocytes expressing genes associated with growth plate hypertrophic chondrocytes. By contrast, specification with Gdf5, together with inhibition of hedgehog and BMP signaling pathways, generated a population of non-hypertrophic chondrocytes that displayed properties of articular chondrocytes. The two chondrocyte populations retained their hypertrophic and non-hypertrophic properties when induced to generate spatially organized proteoglycan-rich cartilage-like tissue in vitro. Transplantation of either type of chondrocyte, or tissue generated from them, into immunodeficient recipients resulted in the development of cartilage tissue and bone within an 8-week period. Significant ossification was not observed when the tissue was transplanted into osteoblast-depleted mice or into diffusion chambers that prevent vascularization. Thus, through stage-specific manipulation of appropriate signaling pathways it is possible to efficiently and reproducibly derive hypertrophic and non-hypertrophic chondrocyte populations from mouse ESCs that are able to generate distinct cartilage-like tissue in vitro and maintain a cartilage tissue phenotype within an avascular and/or osteoblast-free niche in vivo.

Transcriptomics and metabolomics: Challenges of studying obesity in osteoarthritis.

Objective: Obesity is a leading risk factor for both the incidence and progression of osteoarthritis (OA). Omic technologies, including transcriptomics and metabolomics are capable of identifying RNA and metabolite profiles in tissues and biofluids of OA patients. The objective of this review is to highlight studies using transcriptomics and metabolomics that contribute to our understanding of OA pathology in relation to obesity. Design: We conducted a targeted search of PUBMED for articles, and GEO for datasets, published up to February 13, 2024, screening for those using high-throughput transcriptomic and metabolomic techniques to study human or pre-clinical animal model tissues or biofluids related to obesity-associated OA. We describe relevant studies and discuss challenges studying obesity as a disease-related factor in OA. Results: Of the 107 publications identified by our search criteria, only 15 specifically used transcriptomics or metabolomics to study joint tissues or biofluids in obesity-related OA. Specific transcriptomic and metabolomic signatures associated with obesity-related OA have been defined in select local joint tissues, biofluids and other biological material. However, considerable challenges exist in understanding contributions of obesity-associated modifications of transcriptomes and metabolomes related to OA, including sociodemographic, anthropometric, dietary and molecular redundancy-related factors. Conclusions: A number of additional transcriptomic and metabolomic studies are needed to comprehensively understand how obesity affects OA incidence, progression and outcomes. Integration of transcriptome and metabolome signatures from multiple tissues and biofluids, using network-based approaches will likely help to better define putative therapeutic targets that could enable precision medicine approaches to obese OA patients.

HIF-1α and MIF enhance neutrophil-driven type 3 immunity and chondrogenesis in a murine spondyloarthritis model.

The hallmarks of spondyloarthritis (SpA) are type 3 immunity-driven inflammation and new bone formation (NBF). Macrophage migration inhibitory factor (MIF) was found to be a key driver of the pathogenesis of SpA by amplifying type 3 immunity, yet MIF-interacting molecules and networks remain elusive. Herein, we identified hypoxia-inducible factor-1 alpha (HIF1A) as an interacting partner molecule of MIF that drives SpA pathologies, including inflammation and NBF. HIF1A expression was increased in the joint tissues and synovial fluid of SpA patients and curdlan-injected SKG (curdlan-SKG) mice compared to the respective controls. Under hypoxic conditions in which HIF1A was stabilized, human and mouse neutrophils exhibited substantially increased expression of MIF and IL-23, an upstream type 3 immunity-related cytokine. Similar to MIF, systemic overexpression of IL-23 induced SpA pathology in SKG mice, while the injection of a HIF1A-selective inhibitor (PX-478) into curdlan-SKG mice prevented or attenuated SpA pathology, as indicated by a marked reduction in the expression of MIF and IL-23. Furthermore, genetic deletion of MIF or HIF1A inhibition with PX-478 in IL-23-overexpressing SKG mice did not induce evident arthritis or NBF, despite the presence of psoriasis-like dermatitis and blepharitis. We also found that MIF- and IL-23-expressing neutrophils infiltrated areas of the NBF in curdlan-SKG mice. These neutrophils potentially increased chondrogenesis and cell proliferation via the upregulation of STAT3 in periosteal cells and ligamental cells during endochondral ossification. Together, these results provide supporting evidence for an MIF/HIF1A regulatory network, and inhibition of HIF1A may be a novel therapeutic approach for SpA by suppressing type 3 immunity-mediated inflammation and NBF.

Emerging molecular biomarkers in osteoarthritis pathology.

Osteoarthritis (OA) is the most common form of arthritis resulting in joint discomfort and disability, culminating in decline in life quality. Attention has been drawn in recent years to disease-associated molecular biomarkers found in readily accessible biofluids due to low invasiveness of acquisition and their potential to detect early pathological molecular changes not observed with traditional imaging methodology. These biochemical markers of OA have been found in synovial fluid, blood, and urine. They include emerging molecular classes, such as metabolites and noncoding RNAs, as well as classical biomarkers, like inflammatory mediators and by-products of degradative processes involving articular cartilage. Although blood-based biomarkers tend to be most studied, the use of synovial fluid, a more isolated biofluid in the synovial joint, and urine as an excreted fluid containing OA biomarkers can offer valuable information on local and overall disease activity, respectively. Furthermore, larger clinical studies are required to determine relationships between biomarkers in different biofluids, and their impacts on patient measures of OA. This narrative review provides a concise overview of recent studies of OA using these four classes of biomarkers as potential biomarker for measuring disease incidence, staging, prognosis, and therapeutic intervention efficacy.

Identification of a differential metabolite-based signature in patients with late-stage knee osteoarthritis.

Objective: Multiple disease phenotypes have been identified in knee osteoarthritis (OA) patients based on anthropometric, sociodemographic and clinical factors; however, differential systemic metabolite-based signatures in OA patients are not well understood. We sought to identify differential plasma metabolome signatures in a cross-sectional sample of late-stage knee OA patients. Methods: Plasma from 214 (56.5% female; mean age â€‹= â€‹67.58 years) non-diabetic, non-obese (BMI <30 â€‹kg/m2, mean â€‹= â€‹26.25 â€‹kg/m2), radiographic KL 3/4 primary knee OA patients was analyzed by metabolomics. Patients with post-traumatic OA and rheumatoid arthritis were excluded. Hierarchical clustering was used to identify patient clusters based on metabolite levels. A refined metabolite signature differentiating patient clusters was determined based on ≥ 10% difference, significance by FDR-adjusted t-test (q-value < 0.05), and random forests importance score ≥1, and analyzed by AUROC. Bioinformatics analysis was used to identify genes linked to ≥2 annotated metabolites. Associated enriched pathways (q â€‹< â€‹0.05) were determined. Results: Two patient clusters were determined based on the levels of 151 metabolites identified. Metabolite signature refinement found 24 metabolites could accurately predict cluster classification within the sample (AUC â€‹= â€‹0.921). Fifty-six genes were linked to at least 2 â€‹KEGG annotated metabolites. Pathway analysis found 26/56 genes were linked to enriched pathways including tRNA acylation and B-vitamin metabolism. Conclusion: This study demonstrates systemic metabolites can classify a cross-sectional cohort of OA patients into distinct clusters. Links between metabolites, genes and pathways can help determine biological differences between OA patients, potentially improving precision medicine and decision-making.

Can CD200R1 Agonists Slow the Progression of Osteoarthritis Secondary to Injury?

Post-traumatic knee osteoarthritis is characterized by cartilage degeneration, subchondral bone remodeling, osteophyte formation, and synovial changes. Therapeutic targeting of inflammatory activity in the knee immediately post injury may alter the course of osteoarthritis development. This study aimed to determine whether CD200R1 agonists, namely the protein therapeutic CD200Fc or the synthetic DNA aptamer CCS13, both known to act as anti-inflammatory agents, are able to delay the pathogenesis of injury-associated knee osteoarthritis in a murine model. Ten week old male C57BL/6 mice were randomized and surgical destabilization of the medial meniscus (DMM) to induce knee arthritis or sham surgery as a control were performed. CCS13 was evaluated as a therapeutic treatment along with CD200Fc and a phosphate-buffered saline vehicle control. Oligonucleotides were injected intra-articularly beginning one week after surgery, with a total of six injections administered prior to sacrifice at 12 weeks post-surgery. Histopathological assessment was used as the primary outcome measure to assess cartilage and synovial changes, while µCT imaging was used to compare the changes to the subchondral bone between untreated and treated arthritic groups. We did not find any attenuation of cartilage degeneration or synovitis in DMM mice with CD200Fc or CCS13 at 12 weeks post-surgery, nor stereological differences in the properties of subchondral bone. The use of CD200R1 agonists to blunt the inflammatory response in the knee are insufficient to prevent disease progression in the mouse DMM model of OA without anatomical restoration of the normal joint biomechanics.

Contribution of MicroRNA-27b-3p to Synovial Fibrotic Responses in Knee Osteoarthritis.

OBJECTIVE: Synovial fibrosis contributes to osteoarthritis (OA) pathology, but the underlying mechanisms remain unknown. We have observed increased microRNA-27b-3p (miR-27b-3p) levels in synovial fluid of patients with late-stage radiographic knee OA. Here, we investigated the contribution of miR-27b-3p to synovial fibrosis in patients with severe knee OA and in a mouse model of knee OA. METHODS: We stained synovium sections obtained from patients with radiographic knee OA scored according to the Kellgren/Lawrence scale and mice that underwent destabilization of the medial meniscus (DMM) for miR-27b-3p using in situ hybridization. We examined the effects of intraarticular injection of miR-27b-3p mimic into naive mouse knee joints and intraarticular injection of a miR-27b-3p inhibitor into mouse knee joints after DMM. We performed transfection with miR-27b-3p mimic and miR-27b-3p inhibitor in human OA fibroblast-like synoviocytes (FLS) using reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) array, RNA sequencing, RT-qPCR, Western blotting, immunofluorescence, and migration assays. RESULTS: We observed increased miR-27b-3p expression in the synovium from patients with knee OA and in mice with DMM-induced arthritis. Injection of the miR-27b-3p mimic in mouse knee joints induced a synovial fibrosis-like phenotype, increased synovitis scores, and increased COL1A1 and α-smooth muscle actin (α-SMA) expression. In the mouse model of DMM-induced arthritis, injection of the miR-27b-3p inhibitor decreased α-SMA but did not change COL1A1 expression levels or synovitis scores. Transfection with the miR-27b-3p mimic in human OA FLS induced profibrotic responses, including increased migration and expression of key extracellular matrix (ECM) genes, but transfection with the miR-27b-3p inhibitor had the opposite effects. RNA sequencing identified a PPARG/ADAMTS8 signaling axis regulated by miR-27b-3p in OA FLS. Human OA FLS transfected with miR-27b-3p mimic and then treated with the PPARG agonist rosiglitazone or with ADAMTS8 small interfering RNA exhibited altered expression of select ECM genes. CONCLUSION: Our findings demonstrate that miR-27b-3p has a key role in ECM regulation associated with synovial fibrosis during OA.