Chemokine Ligand-3 (CCL3) as a Novel Mediator of Inflammatory Bowel Disease Activity

Abstract

Inflammatory bowel disease (IBD) is characterized by dysregulation of the immune system, driven in part by pro-inflammatory macrophage polarization. Chemokine ligand-3 (CCL3) is a chemokine known to mediate Th1 responses in inflammatory conditions and act to polarize M1 macrophages at the site of inflammation. CCL3 is upregulated in patients with IBD but its contribution to disease pathophysiology is unknown. We hypothesized that CCL3 expression increases at sites of inflammation in biopsies of IBD patients and that it mediates inflammation through chemoattraction of M1 macrophages. To investigate the relationship between CCL3 and IBD, gastrointestinal mucosa biopsies of patients with IBD were immunostained for CCL3, CCR1, and CCR5. Imaging mass cytometry (IMC) was performed to identify CCL3 cell type expression and CCL3-expressing cell spatial relationships. Immunohistochemistry found that there was no correlation between the severity of inflammation and percent of CCL3 positive cells in the lamina propria of IBD patient gut mucosa biopsies. However, IMC analysis highlighted a positive correlation between CCL3 expression and the severity of inflammation of IBD pediatric biopsies. The majority of CCL3 positive cells were neutrophils and macrophages (in particular, proinflammatory M1 macrophages), but there was also some expression on eosinophils and epithelial cells. Spatial proteomics identified that CCL3 expression was highest in the neutrophil and M1 macrophage cell types. Finally, there was no significant difference between the fluorescent intensity of CCL3, CCR1 and CCR5 across IBD gut mucosa biopsies. However, the co-occurrence and correlation of CCR1 with CCL3 was significantly higher than that of CCR5 with CCL3. This suggested that the CCL3 mediated effect may have occurred primarily through the CCL3-CCR1 signalling pathway. Overall, CCL3 expression was positively associated with severity of inflammation in pediatric biopsies, and it is thought to primarily mediate its effects through M1 macrophage and neutrophil polarization in the context of IBD.