CX3CR1+ Synovial Macrophages for Barrier Restore in Rheumatoid Arthritis: A New Target for Mesenchymal Stem Cell-Based Therapy?

Objective: In this study, we explored the interaction between Mesenchymal stem cells (MSCs) and synovial CX3CR1+ macrophages with rheumatoid arthritis (RA) model mice during the process of barrier reconstruction, deeply analyzed the therapeutic mechanism of MSCs and the underlying mechanisms of the dynamic state changes of the macrophage barrier. Design: ScRNA-seq analysis was conducted 4 days after MSCs/PBS injection in serum transfer-induced arthritis model mice. The role of mitochondria transfer and transplantation of MSCs were also explored. Bulk RNA-seq analysis was performed after the co-culture of MSCs and synovial CX3CR1+ macrophages in vitro. MSCs were retrieved 0/2/4/7 days after injection for Bulk RNA-seq analysis to explore their fate in vivo. Results: Intra-articular injection of MSCs could help restore CX3CR1+ macrophage barrier and attenuate the pathological process of RA mice. ScRNA-seq analysis revealed that an Atf3high Ccl3high subset of CX3CR1+ macrophages with poor oxidative phosphorylation increased in abundance during RA progression. MSCs reduced this subset's abundance through mitochondria transfer. Mitochondria transplantation by MSCs was also effective in treating RA. Bulk RNA-seq and scRNA- seq analyses revealed multiple interaction mechanisms between MSCs and CX3CR1+ macrophages such as Cd74/Mif axis and GAS6/MERTK axis to restore CX3CR1+ macrophage barrier. Bulk RNA-seq analysis showed that MSCs primarily contribute to tissue repair and immune regulation subsequently. Conclusions: Our results suggest that MSCs enhanced the oxidative phosphorylation capacity of CX3CR1+ lining macrophages through mitochondria transfer and promote barrier restoration. In addition, we elucidated the fate of MSCs and the therapeutic potential of mitochondria in RA treatment. Overall design: To investigate the potential interaction between ADSCs and Macrophages, in vito separately-cultured and co-cultured ADSCs and Macrophages were collected for RNA-seq. To investigate the heterogeneity changes of CX3CR1+ synovial lining macrophages in the barrier disruption and restoration process, CX3CR1+ macrophages of RA model mice with or without ADSCs treatment were collected for single cell RNA-seq. To investigate the in vivo fate of ADSCs during RA treatment, injected ADSCs were collected from mice knee joints at different time point after injection for RNA-seq.