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Abstract

Background: Atherosclerosis, in the form of ischemic heart disease and stroke, is the leading cause of death globally. Its pathogenesis involves lipid accumulation in artery wall foam cells. Previous work indicates that foam cells in human and mouse atherosclerotic lesions are predominantly derived from smooth muscle cells (SMCs). Our laboratory has also characterized SMCs as being low in lysosomal acid lipase (LAL), the sole lysosomal cholesteryl ester hydrolase. Cholesteryl ester hydrolysis is critical in releasing cellular cholesterol stores for removal via cholesterol efflux. SMCs therefore represent a resistant pool of foam cells with reduced ability to efflux cholesterol. My dissertation investigates whether increasing LAL in SMCs can increase efflux for therapeutic effect in atherosclerosis. Methods and Results: In Chapter 1, I provided an overview of atherosclerosis, SMCs, and LAL. In Chapter 2, I tested whether increasing LAL activity in SMCs in vitro reduces lysosomal cholesteryl ester accumulation and increases cholesterol efflux, using adenoviral vectors, lipid nanoparticles (LNPs), and conditioned medium from SMCs treated with LNPs carrying LIPA mRNA, which encodes LAL. Increasing LAL activity by these methods reduced lysosomal cholesteryl esters. Treating SMCs with LNP LIPA conditioned medium increased efflux to ApoAI. In Chapter 3, I utilized a mouse model containing a tetracycline-response element promoter controlling human LIPA and a SMC specific promoter (SM22alpha) controlling a reverse-tetracycline transactivator, on a background of ApoE-deficiency, to specifically overexpress LAL in SMCs in response to doxycycline. I confirmed with preliminary data that increased SMC LAL reduced atherosclerosis progression. In Chapter 4, I tested whether increased circulating LAL stimulates atherosclerosis regression. LNPs carrying LIPA mRNA were injected into ApoE-deficient and AAV-PCSK9 atherosclerotic mice. I demonstrated increased serum LAL activity and aortic LAL protein, and preliminary evidence of reduced atherosclerosis, but also excess mortality in ApoE-deficient mice with prolonged LNP treatment. Conclusions: Increasing LAL in SMCs reduces lysosomal cholesteryl esters, freeing cholesterol stores to be effluxed from the cell. I determined that this reduces atherosclerotic lesion area in my SMC-specific LAL overexpression model. I also investigated LNP-mRNA as a method to increase LAL and target SMCs. These studies validate LAL and SMC foam cells as crucial therapeutic foci for atherosclerosis.