Outer membrane vesicles from Fusobacterium nucleatum aggravate rheumatoid arthritis

Abstract

Researchers have identified a new interaction mode between gut microbiota and immune homeostasis of the synovium in modulating autoimmune responses in rheumatoid arthritis (RA), according to a new study published in Cell Host & Microbe.1 The study findings provide valuable information and insights into the causes of inflammation of RA and suggest Fusobacterium nucleatum (F. nucleatum) and its outer membrane vesicles (OMVs) as promising therapeutic targets for clinically ameliorating RA. The gut microbiota is gaining increasing attention as it plays essential but complex roles in maintaining host immune homeostasis of RA. However, due to physical and chemical restrictions, the gut microbiota can hardly translocate into distant organs intercellularly but primarily by secreting metabolites, proteins, and OMVs. “Accumulating evidence shows that microbiota-derived OMVs induce immune responses by carrying molecules including peptidoglycans, lipids, proteins, and nucleic acids targeting immune cells, especially macrophages and neutrophils. Moreover, the inherent surface of the lipid phase and physical structure of OMVs protect these effectors from exposure to the clearance mechanisms, suggesting OMV is one of the key players of interkingdom crosstalk between the host and microbes in pathogenic contexts. Therefore, we hypothesize that the gut microbiota associated OMVs may play a role in mediating the contribution of gut microbiota to the development of RA.” remarks Juan Li, one of the responding authors of the study. Through a combination of 16S rRNA-sequencing, clinical studies, and in vivo models, the researchers demonstrate that F. nucleatum is enriched in RA patients and positively correlated with disease activity. Importantly, transplanting F. nucleatum could aggravate inflammation in RA, while depleting F. nucleatum by Metronidazole could alleviate arthritis. “Previous studies mainly suggested F. nucleatum participates in various oral and gastrointestinal diseases, especially colorectal cancer. In this study, we found that F. nucleatum could affects distant joints and plays a role in mediating the aggravation of arthritis.” remarks Li. “OMV secretion is one of the most critical pathways of gram-negative bacteria interacting with the host. As one of common gram-negative bacteria, previous studies suggested that F. nucleatum derived OMV (F.n OMVs) could induce intestinal inflammation. All these studies inspired us to explore whether the effects of F. nucleatum on arthritis depend on OMVs.” explains Mukeng Hong, one of the authors of the study. To test whether F.n OMVs could translocate from the gut to the joints, authors labeled OMV with lipophilicity carbocyanine dyes DIL and observed that F.n OMVs translocated throughout the body and finally accumulated in the limbs. Also, the accumulated F.n OMVs in the joints aggravate arthritis, indicating that F. nucleatum promotes arthritis by secreting OMVs. Subsequently, considering that macrophages and neutrophils are major phagocytic cells that identify and internalize bacterial effectors, the authors administered F.n OMVs for the treatment of macrophage- or neutrophil-depleted arthritis and found that macrophage depletion, and not neutrophils, blocked the promotion effects of F.n OMVs on arthritic symptoms. The authors respectively depleted each contents of F.nOMVs (including nucleic acids, proteins and lipopolysaccharide). “FadA and Fap2 are the crucial proteins of F. nucleatum reported to mediate the systemic dissemination and pro-inflammatory cascades of F. nucleatum. Proteomic analysis of F.n OMVs identified the presence of FadA but not Fap2 in the F.n OMVs. Therefore, we subsequently focused on FadA.” remarks Li. By depleting FadA of F. nucleatum through homologous recombination, researchers found that F.n OMVs lacking FadA could not aggravate inflammation in artist mice, suggesting that FadA is critical for OMVs to exert its effects on arthritis locally. To elucidate the downstream molecular mechanisms underlying the F.n OMVs-aggravated RA, the authors performed transcriptomics and bioinformatics, revealing that GTPase Rab5a is responsive to F.n OMV-FadA stimulation. Also, on silencing Rab5a in vitro and vivo, F.n OMVs did not promote inflammation in arthritis, implying that Rab5a is the critical target by which FadA-containing F.n OMVs mediate aggravation effects on arthritis. “GTPase are involved in vesicle trafficking and inflammatory pathways modulation. We could still observe that F.n OMVs be internalized by the Rab5a depleting macrophages. Therefore, we hypothesized that Rab5a might interact with certain regulators of inflammatory pathways and activate the related inflammatory responses.” By pulling down proteins targeting Rab5a through coimmunoprecipitation and performing proteomics analysis, the authors noticed that Y-Box Binding Protein 1 (YB-1) is the most abundant transcription factor interconnected with Rab5a. Mechanistically, administration of Fn OMVs enhance the interaction between Rab5a and YB-1, which promotes phosphorylation and nuclear translocation of YB-1. Also, by performing luciferase reporter assays, the authors proved that YB-1 could directly bind the promoter region of interleukin-6 and tumor necrosis factor-α and modulate the promoter activity. To further verify our findings in clinical practice that F. nucleatum aggravates arthritis through the FadA-Rab5a-YB-1 axis, the authors collected joint fluid and synovial samples from RA patients. Importantly, all these critical factors underlying F. nucleatum-aggravated arthritis, that is, FadA, Rab5a, and YB-1, were elevated in RA patients compared to the controls. “These findings provide strong evidence to support the occurrence of the translocation of FadA-containing F.n OMVs from gut to joint and suggest its causal role in aggravating RA by targeting Rab5a-YB-1 axis.” remarks Li. The author declares no conflict of interest.