IL-33 and ST2 are important factors in the pathogenesis of IBD. The aim of our study was to characterize the precise contribution of IL-33/ST2 axis in the DSS and AOM/DSS model of colitis.Methods:
C57/BL6 wild-type (WT), IL-33 KO and ST2 KO mice were given a single dose of AOM followed by 2 cycles of DSS for 7 days in drinking water. Aged-matched WT mice, injected with vehicle and given regular drinking water were used as controls (CT). IHC, immunofluorescence (IF) and qPCR were done on full-thickness colons. FACS analysis was performed on resected, isolated polyps. DSS was also administered for 5 days to WT, IL-33 KO and ST2 KO mice. DSS was then replaced with drinking water for 2 weeks (recovery period). Another group of WT mice received DSS for 5 days and IL-33 or vehicle (VEH) during the recovery period. Mice were sacrificed either after DSS challenge or after 1 or 2 weeks of recovery.Results:
IL-33, ST2L, and sST2 mRNA transcripts were dramatically elevated in WT versus CT mice. IHC of treated WT mice revealed localization of IL-33 to the colonic epithelium and to cells within the LP morphologically consistent with tissue macrophages. ST2 staining was localized to the intestinal epithelium in tissues immediately adjacent to tumors, while within the tumors themselves, ST2+ cells displayed a spindle/fibroblast-like morphology with a unique distribution throughout the polyps. Little to no staining for both IL-33 and ST2 was present in CT. Using IF, ST2 co-localized with αSMA in polyps; ST2 staining was not exclusive for αSMA+ cells. FACS analysis showed a distinct population of CD45+ hematopoietic cells consisting of CD3/CD8+ cytotoxic T cells, CD19+ B-lymphocytes, CD11b+CD11c− and CD11b+CD11c+ myeloid cells. ST2 was mainly expressed by CTLs, and CD11b+CD11c− and CD11b+CD11c+ myeloid cells. Non-hematopoietic cells (CD45−) also expressed ST2. DSS challenge in WT mice resulted in increased body weight loss and DAI versus IL-33 KO and ST2 KO mice. At 5 weeks post AOM injection, experimental mice underwent survival colonoscopy. WT had developed pre-tumorous lesions, while IL-33 KO and ST2 KO mice showed their absence with a more impressive mucosal inflammation, likely due to reduced epithelial proliferation. At sacrifice, increased number and size of polyps were observed in WT versus IL-33KO and ST2KO mice. More severe colitis was observed following DSS+1 week recovery versus after 5 days of DSS, which decreased after DSS+2 weeks recovery. ST2 staining was more evident during the recovery phase following DSS, localized in close proximity to areas of re-epithelialization. Both IL-33 and ST2 KO mice showed increased colonic inflammation after 2 weeks recovery compared to after 5 days DSS and versus also WT. IL-33 treatment of WT mice resulted in increased body weight, reduced DAI, and decreased colonic inflammation after 2 weeks recovery versus VEH.Conclusions:
Our results suggest that activation of the IL-33/ST2 axis sustains mucosal healing and tumorigenesis in the murine model of colitis-associated CRC. Further studies are underway to determine mechanisms of action that support these findings.