Background/Purpose: Osteoarthritis (OA) is a complex disease involving pathological processes in joint tissues such as the cartilage and synovium. Cartilage is degraded by matrix metalloproteinases (MMPs) secreted by chondrocytes in response to inflammatory stimuli. Monocytes/macrophages also secrete MMPs and pro-inflammatory signals and are the most abundant immune cell type in the OA-inflamed synovium. Previous literature has shown that Nr4a1-expressing monocytes/macrophages are major secretors of MMPs. Data from the Accelerating Medicines Partnership (AMP) RA Phase I found that as the degree of synovial inflammation increases, multiple cell populations in the OA synovium, including monocytes/macrophages, upregulate the expression of the transcription factor Nr4a1. However, the role of Nr4a1 in cells within different joint tissues during OA is not yet fully understood.

Methods: We used flow cytometry to study the cellular heterogeneity in the synovium of a murine model of OA. To understand the role of Nr4a1 in monocytes/macrophages and other cells within the knee joint during OA, we performed MLI on Nr4a1 wild-type (WT), Nr4a1 Heterozygous (Het) and Nr4a1 Knockout (KO) littermate mice (WT n=3, Het n=4, KO n=6). Knee joints were harvested 4 weeks later for histology. In vitro studies using RT-qPCR were performed to assess the effects of loss of Nr4a1 in both ATDC5 chondrogenic cells transfected with Nr4a1 silencing RNA (siRNA) and primary Nr4a1 KO chondrocytes. Statistics were performed using unpaired Student’s t-test and One-way or Two-way ANOVA followed by Tukey’s post hoc test. Statistical significance was defined as p< 0.05.  

Results: We found a higher percentage of monocytes/macrophages in cells disaggregated from the synovium of C57BL6/J mouse knee joints subjected to meniscal/ligamentous injury (MLI) compared to sham joints (n=5, p< 0.05). Moreover, Nr4a1-GFP reporter mouse knee joints subjected to MLI showed a greater percentage of monocytes/macrophages expressing Nr4a1 than sham joints (n=5, p< 0.05). Histomorphometry showed a significant total tibial cartilage area loss in Nr4a1 WT compared to Het and KO knee joints (p< 0.01 and p< 0.05, respectively). Loss of Nr4a1 in both ATDC5 chondrogenic cells transfected with Nr4a1 silencing RNA (siRNA) and primary Nr4a1 KO chondrocytes resulted in a significant decrease in the gene expression of MMPs after IL-1ß stimulation. Interestingly, the expression of various pro-inflammatory genes increased significantly after IL-1ß stimulation in ATDC5 chondrogenic cells transfected with Nr4a1 siRNA.

Conclusion: Our findings indicate that monocytes/macrophages and chondrocytes express Nr4a1 during OA-related inflammation, and its downregulation reduces MMP gene expression in chondrocytes. This suggests that dampening MMP secretion, even in the presence of pro-inflammatory signals, could be protective for cartilage integrity in OA. Targeting Nr4a1 could be a potential strategy to explore this avenue.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/loss-of-nr4a1-expression-protects-cartilage-during-post-traumatic-osteoarthritis/