5 - IL-33 PROMOTES CARDIOVASCULAR LESION DEVELOPMENT IN A MURINE MODEL OF KAWASAKI DISEASE BY BOOSTING IL-1 BETA PRODUCTION

Background: IL-1b plays a key role in the development of Kawasaki Disease (KD), a pediatric systemic vasculitis. Circulating levels of IL-33, an alarmin released upon inflammation and tissue damage that belongs to the IL-1 cytokine family and exerts its effects by binding to its receptor ST2 (Il1rl1), are elevated in KD patients during the acute phase of the disease. However, if and how IL-33 contributes to cardiovascular lesion development in KD remains unknown.



Methods: We used the well-established Lactobacillus casei cell wall extract (LCWE) murine model of KD vasculitis to determine the role of IL-33 on the development of cardiovascular lesions. To quantify Il33 and Il1rl1 mRNA transcripts and assess their cellular expression, we performed bulk RNA sequencing, single-cell RNA sequencing, spatial transcriptomics, and flow cytometric analysis on abdominal aorta and heart tissues collected from PBS control and LCWE-injected mice. We also performed immunofluorescent staining on tissue sections from control and LCWE-injected mice to quantify IL-33 and ST2. We investigated the role of IL-33 in vivo using either Il-33 deficient mice or a neutralizing antibody against IL-33 in this experimental model of KD. Finally, we studied the effect of IL-33 on macrophages in vitro using LCWE-stimulated bone marrow-derived macrophages (BMDMs) and measured IL-1β levels in the supernatant of these cells by ELISA.



Results: We observed increased Il33 mRNA expression in hearts and abdominal aorta lesions of LCWE-injected mice. While ST2 was detected by immunofluorescence in the hearts of both PBS control and LCWE-injected mice, its expression was increased in the abdominal aortas of LCWE-injected mice. In contrast, IL-33 expression was only increased in inflamed cardiovascular tissues of LCWE-injected mice. Single-cell RNA-sequencing and flow cytometric analysis of abdominal aortas, as well as spatial transcriptomics of heart tissues from LCWE-injected mice, indicated that Il33 transcripts were expressed primarily by stromal cells, such as fibroblasts, endothelial cells, and vascular smooth muscle cells. On the other hand, Il1rl1 transcripts were highly expressed by tissue infiltrating immune cells, such as macrophages, eosinophils, and neutrophils. Blocking IL-33, using either Il33^–/– mice or an anti-IL-33 antibody, significantly attenuated LCWE-induced cardiovascular lesions in this murine model of KD vasculitis. In vitro, IL-33 treatment of LCWE-stimulated BMDMs boosted their IL-1β production.




Conclusions:  Our results indicate that IL-33, produced by stromal cells, may promote LCWE-induced KD vasculitis by increasing the release of IL-1β by tissue infiltrating immune cells, and this axis could therapeutically be targeted in KD vasculitis.