This is a resubmission of K99 HL114648-01. Dr. Csanyi is establishing himself as an independent investigator in the field of cardiovascular science with specific focus on the role of thrombospondin-1 (TSP1)-mediated CD47 signaling in atherosclerosis and vascular free radical biology. This grant will be critical to achieve the following short- and long-term objectives: 1) to acquire additional scientific training in skills ad knowledge; 2) to merge the yet distinct NADPH oxidase (Nox) and TSP1 fields with the goals toward opening up new avenues of discovery and establishing Dr. Csanyi's independent research program; and 3) to become an independently funded scientist at the forefront of cardiovascular and TSP1-related research. Dr. Csanyi has assembled a multidisciplinary team, including his mentor, collaborators, and advisors to guide his career towards independence and assist with the completion of the research proposed in this application. As Dr. Csanyi moves towards his scientific independence he will greatly benefit from attending the following seminars: Course in Scientific Management and Leadership, Managing the Direction of Your Career, and Grant Writing workshop. By the end of the funding period of this K99/R00 award, it is expected that Dr. Csanyi will have published several high-impact first and last author papers and successfully competed for subsequent NIH funding. The overall objective of the research plan is to investigate a new mechanistic pathway linking pathological TSP1-mediated CD47 activation in macrophages with Nox activation and foam cell formation, contributing to the evolution of atherosclerotic plaques, plaque destabilization and thrombosis. The grant's overarching hypothesis is that CD47 activation by TSP1 promotes increased Nox-derived reactive oxygen species (ROS) production in macrophages, increasing oxidation of LDL and its phagocytosis, resulting in foam cell formation (Aim 1) and initiation of atherosclerosis (Aim 2). It is also proposed that CD47-driven ROS in macrophages increases matrix metalloproteinase activity leading to plaque destabilization and ultimate rupture of atherosclerotic plaques and thrombosis (Aim 3). ROS will be measured using cytochrome c, electron paramagnetic resonance, and quantification of 2-hydroxyethidium. The role of CD47 will be investigated using CD47 binding sequences, CD47 null macrophages, and CD47 gene silencing morpholinos. Nox null cells and siRNA technology will be used to characterize the source of TSP1-stimulated ROS. Unique CD47 null ApoE null bone marrow chimeras will be created to investigate the contribution of macrophage CD47 to plaque rupture and thrombosis. The therapeutic potential of CD47 morpholinos to attenuate plaque progression and prevent rupture will be evaluated. This endeavor is highly significant because it has the potential to a) open up an entirely new field of study; b) provide new evidence that TSP1-CD47 signaling is a stimulator of macrophage ROS, foam cell formation and unstable plaque; and c) identify the TSP1-CD47-Nox nexus as a potential therapeutic target in atherosclerosis and other cardiovascular disorders.
The proposed research plan seeks to provide a better understanding of the mechanisms that contribute to the atherosclerotic process, plaque destabilization and coronary heart disease. Atherosclerosis and its cardiovascular consequences are the number one killer of both men and women in the United States, placing a considerable burden on US health care delivery and its costs. The proposed studies are significant because they lend new insight into the key inflammatory events leading to atherosclerosis. Identification of new targets holds significant promise for the development of effective cutting-edge therapies aimed at preventing this devastating disease.
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