The exosomal miR-26b-3p derived from Crohn's Disease-associated mesenteric adipose tissue induces M1 macrophage polarization and exacerbates ileocolonic anastomosis inflammation via the p38-MAPK signaling pathway

Enhao Wu^1,2Wenwei Qian^3,4Xi Zhang⁵Lili Gu⁶Zhen Guo⁷Zeqian Yu⁶Yi Li⁶Weiming Zhu^1,6*

• ¹Nanjing University, Nanjing, China
• ²Nanjing Jinling Hospital,Affiliated Hospital of Medical School,Nanjing University, NanJing, China
• ³Soochow University, Taipei City, Taiwan
• ⁴The Fourth Affiliated Hospital of Soochow University, Suzhou, China
• ⁵Southeast University, Nanjing, China
• ⁶Nanjing Jinling Hospital,Affiliated Hospital of Medical School,Nanjing University, nanjing, China
• ⁷Nanjing Medical University, Nanjing, China

Purpose: Crohn's Disease (CD) is a chronic inflammatory condition characterized by intestinal inflammation, especially in the progression of postoperative anastomotic recurrence. Recent evidence implicates mesenteric adipose tissue (MAT) in CD pathogenesis, particularly through its exosome secretion, which may influence inflammation pathways. The molecular mechanisms driving this inflammation remain inadequately understood. Methods: Exosomes were isolated from MAT of the diseased bowel and macroscopically normal MAT from the surgical margins of patients with CD. We induced chronic intestinal inflammation in mice using dinitrobenzene sulfonic acid (DNBS), simulating CD-like MAT. Using a surgical model of IL10-knockout mice, we performed a series of experiments in vitro and in vivo to assess the effects of exosomes on ileocolonic anastomosis inflammation and macrophage M1 polarization. We performed microRNA microarray analysis, colonoscopy, Western blotting, luciferase assays, and immunofluorescence to investigate the underlying mechanisms. Results: Hypertrophic MAT-Exosomes (Ht-exos) promoted ileocolonic anastomotic inflammation by activating macrophage M1 polarization in CD. In vivo, injection of Ht-exos induced inflammatory tissue damage and macrophage M1 polarization in an IL-10-/- mouse model of ileocecal resection. In vitro, Ht-exos was found to promote macrophage inflammatory response and M1 polarization through the activation of the p38-MAPK pathway. Further, exosomal miR-26b-3p was enriched in MAT-Exosomes and involved in exosome-mediated inflammation activation. Mechanistically, hypertrophic MAT released exosomal miR-26b-3p and promoted inflammation by targeting tripartite motif-containing 33 (TRIMM33) via the p38-MAPK signaling pathway and promoting macrophage M1 polarization. Furthermore, miR-26b-3p expression was positively correlated with the degree of ileocolonic anastomosis inflammation in CD. Conclusion: Our findings reveal that exosomal miR-26b-3p drives widespread macrophage inflammation and M1 polarization in hypertrophic MAT-induced ileocolonic anastomosis inflammation via the MAPK pathway.