This K22/R00 application describes a career development program designed to facilitate my transition to research independence. My goals during the mentored phase of this award are to cultivate a variety of new research skills as well as receive the career training necessary to acquire a tenure-track assistant professorship. To broaden my research expertise I aim to master new techniques in the fields of ectopic calcification, induced- pluripotent stem cells, and murine models of chronic kidney disease; gaining expertise in these areas will pro- vide me with the background necessary to successfully transition to and sustain an independent research pro- gram. During the mentored phase I also intend to continue to develop career skills in such areas as trainee mentorship, teaching, and the effective communication of my work to colleagues and the general public. I plan on participating in a variety of seminars and workshops offered by the NIH Office of Intramural Training and Education to enhance my leadership and lab management skills. I have assembled an Advisory Committee made up of my primary mentor, three scientific consultants, and a career development advisor, all of whom have pledged their support and guidance throughout the award period of this proposal. During the extramural phase I plan to recruit to my independent laboratory molecular and cellular biologists, as well as clinical fellows interested in conducting patient-based research. My long-term career plan is to pair my expertise in molecular biology, biochemistry, and genetic approaches with clinical collaborations to advance the study of vascular calcification and vessel integrity disorders, with the ultimate goal of identifying drug targets for the development of therapeutics to treat affected patients. Vascular calcification often manifests as a secondary complication to diseases such as chronic kidney disease and diabetes, and is a predictor of premature death. I was part of a team that discovered a novel, monogenetic disease in which affected individuals develop medial arterial calcifications in their lower-extremity arteries. Affected arteries are tortuous and have significantly reduced blood flow, resulting in pain and lack of patient mobility. This disease-Arterial Calcification due to Deficiency of CD73 (ACDC)-results from biallelic inactivating mutations in the gene encoding for the CD73 protein. CD73 converts extracellular AMP to adenosine, and we showed that the vascular calcification in ACDC stems from a lack of extracellular adenosine signaling via adenosine receptors (ARs). The link between vascular calcification and AR signaling is unprecedented. The research I propose in this application will seek to elucidate the mechanisms and processes regulating vascular calcification and vascular homeostasis.
Aim 1 will identify the mechanisms regulating calcification in ACDC cells using an in vitro patient-specific fibroblast calcification model as well as an in vivo patient-specific induced pluripotent stem cell (iPSC) disease model.
Aim 2 will define the role of CD73 in vessel wall homeostasis, specifically 1) the role of CD73-mediated adenosine signaling in keeping vascular smooth muscle cells (VSMCs) in their fully differentiated state, and 2) the role of CD73-mediated adenosine signaling in regulating extracellular matrix cross-linking proteins.
Aim 3 will use an established CD73 knockout mouse to assess the ability of this genetic line to be used as a model for the human disease. Additionally, the role of CD73 and adenosine signaling will be assessed in a murine model of chronic kidney disease, as the lower-extremity medial calcification accompanying this disease is histologically similar to that seen in ACDC patients, and may share common mechanisms and respond to common therapeutic strategies. The significance of this study is best exemplified by the number of people affected by vascular calcification; in addition to ACDC, individuals with chronic kidney disease and diabetes often develop lower-extremity medial arterial calcifications. The innovation of this study is the use of novel, monogenetic disease as a model to study processes regulating vascular calcification, and the creation of an in vivo iPSC-based calcification model that can be used in drug screening. The identification of adenosine signaling as a mechanism that regulates vascular calcification opens up a new avenue of research in the field of ectopic calcification. Elucidating the pathways underlying this monogenetic disease will provide important insight into the process of medial calcification in other disorders, and may lead to better interventions and therapeutics.
Vascular calcification afflicts millions of individuals, and often leads to pain, loss of limbs, and premature death. This research proposal aims to investigate and uncover the underlying causes of vascular calcification, and could help scientists and physicians identify new treatments to prevent, halt, or even reverse the condition in diseases such as chronic kidney disease and diabetes.
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