. Emerging clinical importance of innate immunity and Toll-like receptors (TLRs) in vascular development and diseases, and discoveries of new endogenous TLRs ligands generated by lipid oxidation and their expanding functions in human pathologies underscore importance of TLRs (TLR2 in particular) in cardiovascular and lung diseases and vascular pathologies associated with inflammation, such as retinal diseases of aging and diabetes. TLR2 expression is not restricted to immune system; it is functional on other cells, including endothelium and can be activated by bacterial and endogenous ligands, i.e. products of lipid peroxidation. The knowledge of endothelial TLR2 is limited to studies with inhibitors or ubiquitous TLR2 KO. Accordingly, we propose to establish a role for endothelial TLR2 in angiogenesis using postnatal and developmental models. We will test that endothelial TLR2 regulates angiogenesis by two mechanisms: directly by integrin-mediated migration and indirectly, by promoting recruitment of myeloid cells. TLR2-dependent angiogenesis is augmented in the presence of exogenous bacterial TLR2 ligands as well as endogenous TLR2 ligands, i.e. CEP, especially in tissues where its endogenous ligand is highly prevalent (i.e. in retinas).
Our specific Aims are:
Aim 1. To define the cell autonomous function of TLR2 on endothelium by using endothelial-specific (Cdh5-CreERT) KO mice in postnatal (wound, tumors) models and developmental (retinal) angiogenesis.
Aim 2. To assess the role of TLR2- dependent myeloid cells recruitment in angiogenesis. To establish the role of EC TLR2 as we as the role of myeloid TLR2 using KO (CX3CR1-CreERT) in myeloid cells recruitment during postnatal and developmental angiogenesis.
Aim 3. To define the proangiogenic pathways activated by endogenous and exogenousTLR2 ligands in endothelial cells. The regulatory function and mechanism of TLR2/CD36 complex will be addressed. Role for TLR2/Myd88/TIRAP/IRAK3/4/TRAF6 and Src family kinases in regulation of proangiogenic integrin functions, i.e. activation, adhesion and migration will be established. Main targets of TLR2 pathway novel to EC biology, e.g. TRAF6 will be verified in vivo using novel reagents. These studies will establish for the first time a direct role of endothelial TLR2 in angiogenesis and will link innate immunity and vascularization, especially in pathologies associated and exacerbated by infection, inflammation and oxidative stress.
Increasing number of clinical studies link innate immunity Toll like Receptors with cardiovascular, lung and eye diseases and diabetes. This proposal will show how these receptors and its ligands operate and how they control blood vessels development in physiology in contrast to pathological angiogenesis combined with infection, inflammation and excessive oxidative stress. New models will be used to identify potential targets for these conditions and key compounds will be used to prevent excessive vascularization in pathologies.
