Abstract
Purpose :
Human interleukin-33 (IL-33) is a 270 amino acid protein belonging to the IL-1 family cytokine with important roles in inflammatory diseases. Inflammatory proteases from neutrophils (Cathepsin G and Elastase) and mast cells (tryptase and chymase) regulate the activity of IL-33 by processing full-length IL-33 into mature forms. Despite recent advances in the role of IL-33 in allergic diseases, arthritis, cancer, and cardiovascular diseases, nothing is known regarding the role of these IL-33 mature forms in retinal endothelial cell signaling and angiogenesis.
Methods :
Here, we first cloned the various mature forms of human IL-33, such as IL-3395-270, IL-3399-270, IL-33109-270, and IL-33112-270 into pET30a vector, and expressed these mature IL-33 constructs in E.coli strain BL21(DE-3) and purified by immobilized metal affinity chromatography (IMAC). We next evaluated the role of these recombinant mature IL-33 proteins on human retinal microvascular endothelial cells (HRMVECs) proliferation, migration, tube formation, and sprouting angiogenesis. We also investigated the role of these recombinant mature IL-33 proteins on various cellular signaling involved in retinal endothelial cell angiogenic events by 3’-mRNA sequencing and western blotting.
Results :
We observed that these mature forms of IL-33 produced by inflammatory proteases potentiate HRMVEC proliferation, migration, tube formation, and sprouting angiogenesis. We also observed that these mature forms of IL-33 are more potent in inducing angiogenic signaling and endothelial cell activation. More specifically, IL-3399-270 produced by Cathepsin G (neutrophil protease) cleavage is more potent in inducing endothelial cell activation and angiogenesis.
Conclusions :
Our study suggests that blockage or inhibition of IL-33 cleavage by neutrophil proteases could be useful to limit IL-33-regulated proliferative retinopathies.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.