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Objective Cartilage epigenetic changes are strongly associated with human osteoarthritis (OA). However, the influence of individual environmental OA risk factors on these epigenetic patterns has not been determined; herein we characterize cartilage DNA methylation patterns associated with aging and OA in a mouse model. Methods Murine knee cartilage DNA was extracted from healthy young (16‐week, n = 6), old (82‐week, n = 6), and young 4‐week post–destabilization of the medial meniscus (DMM) OA (n = 6) C57BL6/J mice. Genome‐wide DNA methylation patterns were determined via Illumina BeadChip. Gene set enrichment analysis was performed by Ingenuity Pathway Analysis. The top seven most differentially methylated positions (DMPs) were confirmed by pyrosequencing in an independent animal set. Results were compared to previously published human OA methylation data. Results Aging was associated with 20,940 DMPs, whereas OA was associated with 761 DMPs. Merging these two conditions revealed 279 shared DMPs. All demonstrated similar directionality and magnitude of change (Δβ 1.0% ± 0.2%, mean methylation change ± SEM). Shared DMPs were enriched in OA‐associated pathways, including RhoA signaling (P = 1.57 × 10−4), protein kinase A signaling (P = 3.38 × 10−4), and NFAT signaling (P = 6.14 × 10−4). Upstream regulators, including TET3 (P = 6.15 × 10−4), immunoglobulin (P = 6.14 × 10−4), and TLR7 (P = 7.53 × 10−4), were also enriched. Pyrosequencing confirmed six of the seven top DMPs in an independent cohort. Conclusion Aging and early OA following DMM surgery induce similar DNA methylation changes within a murine OA model, suggesting that aging may induce pro‐OA epigenetic “poising” within articular cartilage. Future research should focus on confirming and expanding these findings to other environmental OA risk factors, including obesity, as well as determining late OA changes in mice.

Introduction: Osteoarthritis (OA) is a chronic musculoskeletal disease characterized by progressive loss of joint function. Historically, it has been characterized as a disease caused by mechanical trauma, so-called ‘wear and tear’. Over the past two decades, it has come to be understood as a complex systemic disorder involving gene-environmental interactions. Epigenetic changes have been increasingly implicated. Recent improvements in microarray and next-generation sequencing (NGS) technologies have allowed for ever more complex evaluations of epigenetic aberrations associated with the development and progression of OA. Methods: A systematic review was conducted in the Pubmed database. We curated studies that presented the results of DNA methylation and noncoding RNA research in human OA and OA animal models since 1985. Results: Herein, we discuss recent findings and methodological advancements in OA epigenetics, including a discussion of DNA methylation, including microarray and NGS studies, and noncoding RNAs. Beyond cartilage, we also highlight studies in subchondral bone and peripheral blood mononuclear cells, which highlight widespread and potentially clinically important alterations in epigenetic patterns seen in OA patients. Finally, we discuss epigenetic editing approaches in the context of OA. Conclusions: Although a substantial body of literature has already been published in OA, much is still unknown. Future OA epigenetics studies will no doubt continue to broaden our understanding of underlying pathophysiology and perhaps offer novel diagnostics and/or treatments for human OA.

ObjectiveAdult elastic cartilage has limited repair capacity. MRL/MpJ (MRL) mice, by contrast, are capable of spontaneously healing ear punctures. This study was undertaken to characterize microbiome differences between healer and non-healer mice and to evaluate whether this healing phenotype can be transferred via gut microbiome transplantation.MethodsWe orally transplanted C57BL/6J (B6) mice with MRL/MpJ cecal contents at weaning and as adults (n = 57) and measured ear hole closure 4 weeks after a 2.0mm punch and compared to vehicle-transplanted MRL and B6 (n = 25) and B6-transplanted MRL (n = 20) mice. Sex effects, timing of transplant relative to earpunch, and transgenerational heritability were evaluated. In a subset (n = 58), cecal microbiomes were profiled by 16S sequencing and compared to ear hole closure. Microbial metagenomes were imputed using PICRUSt.ResultsTransplantation of B6 mice with MRL microbiota, either in weanlings or adults, improved ear hole closure. B6-vehicle mice healed ear hole punches poorly (0.25±0.03mm, mm ear hole healing 4 weeks after a 2mm ear hole punch [2.0mm-final ear hole size], mean±SEM), whereas MRL-vehicle mice healed well (1.4±0.1mm). MRL-transplanted B6 mice healed roughly three times as well as B6-vehicle mice, and half as well as MRL-vehicle mice (0.74±0.05mm, P = 6.9E-10 vs. B6-vehicle, P = 5.2E-12 vs. MRL-vehicle). Transplantation of MRL mice with B6 cecal material did not reduce MRL healing (B6-transplanted MRL 1.3±0.1 vs. MRL-vehicle 1.4±0.1, p = 0.36). Transplantation prior to ear punch was associated with the greatest ear hole closure. Offspring of transplanted mice healed significantly better than non-transplanted control mice (offspring:0.63±0.03mm, mean±SEM vs. B6-vehicle control:0.25±0.03mm, n = 39 offspring, P = 4.6E-11). Several microbiome clades were correlated with healing, including Firmicutes (R = 0.84, P = 8.0E-7), Lactobacillales (R = 0.65, P = 1.1E-3), and Verrucomicrobia (R = -0.80, P = 9.2E-6). Females of all groups tended to heal better than males (B6-vehicle P = 0.059, MRL-transplanted B6 P = 0.096, offspring of MRL-transplanted B6 P = 0.0038, B6-transplanted MRL P = 1.6E-6, MRL-vehicle P = 0.0031). Many clades characteristic of female mouse cecal microbiota vs. males were the same as clades characteristic of MRL and MRL-transplanted B6 mice vs. B6 controls, including including increases in Clostridia and reductions in Verrucomicrobia in female mice.ConclusionIn this study, we found an association between the microbiome and tissue regeneration in MRL mice and demonstrate that this trait can be transferred to non-healer mice via microbiome transplantation. We identified several microbiome clades associated with healing.