Background: During intestinal inflammation, neutrophils aid in the recruitment of other immune cells and facilitate the immune response in the gut. However, during chronic inflammatory conditions, such as Inflammatory Bowel Disease (IBD, excessive neutrophil accumulation can lead to tissue damage, delayed tissue repair and loss of homeostasis. Thus, we aim to identify the role and function of neutrophils in the pathogenesis of colitis.
Methods: To block neutrophil chemoattraction and cytotoxicity in response to its ligand hepatocyte growth factor, we used the neutrophil-specific Mrp8-Cre line backcrossed with Metfl/fl. Acute and chronic colitis were induced in MRP8Cre/WT METfl/fl (KO) mice and MRP8WT/WT METfl/fl littermate controls (WT) by 1 or 3 cycles of 2.5% dextran sodium sulfate (DSS) in drinking water for 5 days followed by 2 weeks of water. Disease progression was assessed via a standardized disease activity index (DAI). Colonic immune cells were assessed by flow cytometry. Data are expressed as mean ± SEM; t-test was performed; p<0.05.
Results: During the third cycle of chronic DSS colitis, KO mice displayed a decreased DAI (p<0.01) and body weight loss (p<0.05) compared to WT mice. Moreover, flow cytometric analysis revealed a reduced amount of ROS+ neutrophils (WT; 3.17±0.84×105, KO; 0.69±0.26×105, p<0.05), eosinophils (WT; 3.00±0.62×105, KO; 0.80±0.20×105, p<0.05), and macrophages (WT; 9.98±0.59×105, KO; 6.46±0.73×105, p<0.05), implying a protective effect of MET deletion in MRP8+ neutrophils during chronic intestinal inflammation. In addition, KO mice subjected to acute DSS colitis showed an improved disease course with reduced body weight loss and DAI and a comparable decrease in the amount of neutrophils, eosinophils and macrophages. Moreover, the percentage of FoxP3+ T regulatory cells was increased in KO mice compared to their WT counterparts (WT; 29.27±2.14%, KO; 41.05±2.11%, p<0.05), pointing towards a return to homeostasis in the KO colon. Strikingly, analysis of CD3+ CD4+ T cells showed a predominant decrease of the percentage IL17A+ Th17 (WT; 26.87±1.85%, KO; 14.19±2.11%, p<0.01) and IL17A+ IFNg+ Th1-like Th17 (WT; 10.62±0.50%, KO; 4.70±1.51%, p<0.01) in KO mice compared to WT mice, while no differences were observed in the percentage of IFNg+ Th1 cells (WT; 12.03±0.61%, KO; 12.03±3.00%, ns).
Conclusions: In the present study, we showed that MET is required for neutrophil chemoattraction and cytotoxicity during colitis. In addition, MET deletion in neutrophils was associated with a specific reduction of Th17 cells. Further understanding the mechanisms underlying neutrophil function during colitis will aid in the development of novel therapeutic strategies to treat IBD patients.