The long-term goal of the proposed studies is to understand the signal transduction mechanisms governing the development and function of the cardiovascular system. In the previous funding period, we focused on the role and mechanisms of focal adhesion kinase (FAK) signaling in ECs during embryonic angiogenesis and FAK- family interacting protein of 200 kDa (FIP200) in cardiac development. Using a functional reconstitution approach based on isolated primary FAK-null ECs, we identified a novel role for S732 phosphorylation of FAK in promoting EC proliferation and angiogenesis by the regulation of centrosome functions during mitosis. We have also created two different FAK mutant knockin mouse models. Using the EC-specific FAK kinase- defective (KD) mutant knockin mouse model, we demonstrated the role of kinase-independent and -dependent functions of FAK in EC survival and barrier function, respectively, during embryonic development. In addition to these and several others published studies that show the role of FAK and FIP200 in cardiac development, this grant also supported our studies on the role and mechanisms of FIP200 in fetal hematopoietic stem cells. In preliminary studies, we established new mouse models with the inducible EC-specific FAK knockout (KO) and KD mutant knockin mice and showed a role of FAK and its kinase activity in adult angiogenesis. Moreover, we observed increased Notch signaling upon FAK deletion or loss of its kinase activity, suggesting a potentially novel mechanism of FAK in the regulation of angiogenesis through the critical Notch pathway. We also generated and analyzed both embryonic and inducible EC-specific KO of TSC1 (tuberous sclerosis complex 1), which provided the first direct evidence for a role of TSC/mTOR signaling in the embryonic vascular development and angiogenesis of adult organisms in mouse models in vivo. Despite these progresses, still relatively little is known about the role and mechanisms o FAK signaling in the regulation of angiogenesis in adult organism in vivo. Likewise, the mechanisms of TSC/mTOR signaling in ECs during embryonic and adult angiogenesis have not been assessed directly in vivo. Based on our previous and preliminary studies, we propose to 1). Analyze the role and mechanism of FAK and its kinase activity in the regulation of Notch signaling in postnatal angiogenesis by using inducible EC-specific FAK KO and KD mutant knockin mouse models, 2). Investigate the role and mechanisms of TSC/mTOR signaling in vascular development and angiogenesis by using an EC-specific TSC1 KO mouse model, and 3). Study the role and mechanisms of TSC/mTOR signaling and its regulation of VEGF expression in postnatal angiogenesis. These studies will generate significant insights into the mechanisms of intracellular signaling in the regulation of angiogenesis in vivo and may also provide critical information for potential development of novel therapies for angiogenesis related diseases.
Angiogenesis plays an essential role in embryogenesis, homeostasis of adult organism, and contributes to various diseases including coronary heart disease, age-related macular degeneration, diabetes and cancer when it is not properly regulated. Analysis of molecular and cellular mechanisms by which key signaling molecules and their associated pathways regulate angiogenesis using mouse models in vivo will significantly advance our understanding of the basic mechanisms of angiogenesis that may contribute to novel therapies for cardiovascular and other diseases.
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