Fusobacterium nucleatum drives CD40-mediated dendritic cell activation and Th17/Treg imbalance to exacerbate intestinal inflammation in Crohn's disease.

Fusobacterium nucleatum was significantly enriched in Crohn's disease patients and positively associated with severity of inflammation.

• Fecal and colonic mucosal samples from CD patients and healthy controls were analyzed by 16S rRNA sequencing.
• Fn abundance was positively correlated with markers of inflammation severity in CD patients.
• Fn was identified as a pathobiont enriched specifically in the CD patient population compared to healthy controls.

Fn aggravated colitis in mice and heightened immune dysregulation, especially dendritic cell activation.

• A TNBS-induced colitis mouse model was used to evaluate the impact of Fn on intestinal inflammation.
• Mice exposed to Fn showed heightened immune dysregulation compared to controls.
• DC activation was identified as a particularly prominent feature of Fn-driven immune dysregulation in the mouse model.

Adoptive transfer of Fn-primed dendritic cells aggravated TNBS-induced intestinal inflammation in recipient mice.

• Bone marrow-derived DCs (BMDCs) were primed with Fn prior to adoptive transfer into recipient mice.
• Recipient mice receiving Fn-primed DCs showed aggravated TNBS-induced colitis compared to controls.
• The aggravated inflammation was accompanied by Th17/Treg imbalance in recipient mice.
• The adoptive DC transfer model directly linked Fn-conditioned DCs to downstream immune dysregulation.

Transcriptomic analysis identified robust CD40 upregulation in Fn-exposed dendritic cells, which was corroborated in inflamed colons from CD patients.

• RNA sequencing was performed on Fn-exposed bone marrow-derived DCs (BMDCs) to identify key immune mediators.
• CD40 was identified as robustly upregulated in Fn-exposed DCs by transcriptomic analysis.
• Elevated CD40 expression was confirmed in inflamed colonic tissue from CD patients, providing clinical translational relevance.
• CD40 upregulation was identified as a key mechanistic link between Fn exposure and DC activation.

CD40 blockade with TRAF-STOP suppressed DC activation, restored Th17/Treg balance, and ameliorated intestinal inflammation.

• TRAF-STOP was used as a CD40 signaling inhibitor both in vivo and in vitro.
• CD40 inhibition with TRAF-STOP suppressed Fn-driven DC activation.
• TRAF-STOP treatment restored Th17/Treg balance that had been disrupted by Fn-primed DCs.
• Intestinal inflammation was ameliorated following CD40 blockade, supporting CD40 as a functional and therapeutically relevant target.

The study establishes a mechanistic link between Fusobacterium nucleatum-driven microbial dysbiosis and immune dysregulation via the CD40 signaling pathway in dendritic cells.

• The proposed pathway involves Fn enrichment leading to CD40-mediated DC activation, which drives downstream Th17/Treg imbalance and exacerbates mucosal inflammation.
• This mechanistic framework connects specific microbial species to defined immune dysfunction in Crohn's disease.
• CD40 is identified as a promising target for microbiota-directed immunotherapy in CD.