CD47 regulates CD11b‐dependent neutrophil transepithelial migration during intestinal inflammation

Inflammatory bowel disease (IBD), including Crohn's disease and Ulcerative colitis, is a chronic, relapsing disorder of the gastrointestinal tract characterized by neutrophil (PMN) associated bystander tissue damage, crypt abscess formation and mucosal ulceration. It is well appreciated that neutrophil transepithelial migration (TEpM) in the intestine correlates with disease flares in individuals with IBD. While the molecular events regulating PMN TEpM are far from understood, it is now clear that the integrin CD11b/CD18 and CD47 play crucial roles in this process. CD47 is a highly glycosylated protein that associates in cis with β1, β2 and β3 integrins in different cell types while also serving as a receptor for Signal Regulatory Proteins (SIRP‐α and ‐γ) and thrombospondin (TSP). CD47 is involved in regulation of innate and adaptive immune responses. For example, it has been reported that PMN recruitment in CD47−/− mice is reduced in models of peritonitis, lung injury and dermal air pouch inflammation, but the exact mechanism(s) behind CD47 regulation of PMN infiltration into inflamed mucosal tissues is unclear. We have previously reported that CD47 expressed in T‐cells regulates transendothelial migration through its physical and functional association with the β2 integrin CD11a/CD18. In the gut, it has been shown that PMN transmigration across intestinal epithelium is dependent on CD11b/CD18 but not CD11a/CD18. Thus, we hypothesized that CD47 interacts with CD11b/CD18 to regulate PMN chemotaxis and TEpM in the intestine. Here we report that CD47 co‐associates with CD11b/CD18 in PMN as determined by co‐immunoprecipitation and proximal ligation assay (PLA). Flow cytometry analyses of murine PMN deficient in CD47 showed that, despite expressing similar levels of CD11b/CD18 at baseline, there was defective upregulation of CD11b/CD18 upon stimulation with the chemoattractants fMLF or LTB 4 compared with WT PMN. Additionally, loss of CD47 in murine PMN resulted in a 25% reduction in LTB 4 driven chemotaxis. Furthermore, antibody mediated inhibition of CD11b/CD18 reduced WT PMN migration to similar levels as CD47−/− PMN. In contrast, blocking CD11b on CD47−/− PMN did not further reduce migration, suggesting a functional regulation of integrin‐dependent chemotaxis by CD47. In vivo studies modeling PMN TEpM into the intestinal lumen in response to LTB 4 revealed decreased migration in CD47−/− mice (>60% reduction vs. WT; p<0.05). Taken together, our data suggest that CD47 plays an important role in PMN intestinal recruitment through regulation of CD11b/CD18 adhesive function. We suggest that CD47 represents a promising new candidate for therapeutic approaches aimed at dampening pro‐inflammatory signals, reducing PMN influx and improving mucosal wound healing in IBD. Support or Funding Information Supported by NIH 5R01DK079392‐12 and R01HL125780