Date of Award

12-2023

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Food Science and Human Nutrition

Committee Chair/Advisor

Alexis Stamatikos

Committee Member

Vivian Haley-Zitlin

Committee Member

Elliot Jesch

Committee Member

Kimberly Paul

Abstract

Atherosclerotic cardiovascular disease is the leading cause of mortality worldwide due to being the predominant factor in heart attack and stroke deaths. Atherosclerotic cardiovascular disease is caused by cholesterol accumulating in arteries. It is historically thought that the cell type most responsible for causing atherosclerosis via excessive cholesterol accumulation is intimal monocyte-derived macrophages. However, macrophage-like cells (MLC) of vascular smooth muscle cell (VSMC) origin have now been proposed as another cell type to trigger atherosclerosis, since VSMC cholesterol accumulation results in these cells to trans-differentiate into a MLC phenotype. Three major genes involved in cholesterol metabolism are ABCA1, ABCG1, and miR-33a-5p. ABCA1 and ABCG1 remove cellular cholesterol upon interaction with apoAI and HDL respectively, while miR-33a-5p can silence ABCA1 expression. Therefore, manipulating expression of these genes precisely in MLC may influence atherogenesis, but this has not been rigorously tested in this cell type. In the first part of my dissertation project, I cultured ABCA1-deficient, ABCG1-deficient, and ABCA1/ABCG1-deficient MLC and measured cholesterol efflux in these cells and in MLC with endogenous ABCA1/ABCG1 expression. As expected, there was a dramatic reduction in apoAI-mediated cholesterol efflux in ABCA1-deficient MLC. However, the reduction in HDL-mediated cholesterol efflux in ABCG1-deficient MLC was not as drastic as anticipated, which made me infer SR-BI may preserve HDL-mediated cholesterol efflux when ABCG1-dependent cholesterol efflux is impaired in MLC, since SR-BI has been shown to participate in HDL-mediated cholesterol efflux. When I knocked down SR-BI expression in ABCG1-deficient MLC, I did observe decreased HDL-mediated cholesterol efflux when compared to ABCG1-deficient MLC without inhibited SR-BI expression, which provides mechanistic evidence that SR-BI may maintain HDL-mediated cholesterol efflux in MLC that have impaired ABCG1 expression. In the second part of my dissertation, I measured ABCA1 protein expression and apoAI-mediated cholesterol efflux in cultured MLC that expressed endogenous miR-33a-5p or had miR-33a-5p expression inhibited. Inhibiting MLC miR-33a-5p expression resulted in increasing ABCA1 protein expression and enhancing apoAI-mediated cholesterol efflux, which suggests miR-33a-5p inhibition in MLC may be atheroprotective. Based on my results, future in vivo studies should directly assess the athero-modulatory effects of miR-33a-5p, ABCA1, ABCG1, and SR-BI expression precisely in VSMC/MLC.

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