VCAM1 regulates Fra1/JunB mediated IL‐8 expression and pathological retinal neovascularization in a murine model of oxygen‐induced retinopathy

Purpose Pathological retinal neovascularization is a clinical manifestation of various proliferative retinopathies, including retinopathy of prematurity (ROP), diabetic retinopathy, and the wet form of macular degeneration. Over the last decade, numerous studies have reported an increased infiltration of hematopoietic stem cells (HSCs), macrophage/microglia in patients with intraocular angiogenic diseases. Methods Using a murine model of oxygen‐induced retinopathy (OIR) and human retinal microvascular endothelial cells (HRMVECs), we are trying to understand the significance of IL‐33/Hypoxic induced VCAM1 expression in retinal neovascularization. Results Here, using a murine model of oxygen‐induced retinopathy, we found that NFkB and VCAM1 levels were highly up regulated in retina when new vessels were developing in the retina. Genetic deletion of IL‐33 resulted in reduced NFkB activation and VCAM1 levels in hypoxic retina, resulting in reduced neovascularization. These findings suggest a role for IL‐33‐NFkB‐VCAM1 signaling in pathological neovascularization. Our results demonstrate that IL‐33 induce tube formation in human retinal microvascular endothelial cells and depletion of VCAM1 levels significantly blocked it. Protein profiler analysis showed that IL‐33 regulates VCAM1 dependent IL‐8 expression in HRMVECs. In order, to understand how IL‐33 induced VCAM1 regulates the expression of IL‐8, we did sequence analysis of IL‐8 promoter and observed that IL‐8 promoter has AP1 binding site. We observed that IL‐33 induced the expression of Fra1 and JunB significantly in HRMVECs. Cloning and site directed mutagenesis of IL‐8 promoter revealed that the AP1 binds to IL‐8 promoter in response to IL‐33 and regulate the expression of IL‐8 in HRMVECs. Consequently, down regulation of VCAM1 levels significantly reduced IL‐33 induced Fra1 and JunB expression as well as IL‐8 expression. Conclusions These results strongly suggest that IL‐33/ST2L‐NFkB‐VCAM1‐AP1 regulates IL‐8 expression, playing a key role in retinal neovascularization and thus represents a new pharmacological target for the treatment of diseases where excessive neovascularization is the underlying pathology.