Abstract 429: The Effects of Fluvastatin and Methyl-β-Cyclodextrin-Mediated Cholesterol Depletion on the Biomechanics of Vascular Smooth Muscle Cells in Atherosclerosis

Abstract

Atherosclerosis is a leading cause of death worldwide. Phenotypic shifting, alteration in cell adhesion, and migration toward inflammatory site of blood vessel wall are all critical contributions of vascular smooth muscle cells (VSMC) to the progression of atherosclerosis. Knowing that cholesterol is a main participant of fatty deposition in atherosclerotic lesions, cholesterol’s role in orchestrating cell migration, mechanics and spreading has grown more apparent. Growing evidences suggests that cholesterol loaded into VSMC in atherosclerosis is much larger than previously known, and about 40% of the total foam cells in the atherosclerotic plaque were VSMC-derived. Emerging studies have increasingly categorized cholesterol as major player in regulating VSMC stiffness and N-Cadherin mediated cell-cell adhesions. Modulating their activity or expression may block VSMC migration during the progression of atherosclerosis. In this study, the effects of a 3-hydroxy-3-methylglutaryl-coenzyme-A reductase inhibitor, fluvastatin and Methyl-β-Cyclodextrin-Mediated (MβCD) cholesterol depletion on VSMCs N-cadherin adhesion and cellular stiffness were addressed. VSMCs cholesterol quantification and lactate dehydrogenase assays demonstrated significant reduction in total cellular cholesterol content by approx. 67% for fluvastatin and 40% for MβCD. The atomic force microscope (AFM) was used to investigate real time adhesion and stiffness of living VSMCs. A proprietary software package written in Matlab was used to estimate Young’s modulus of the cell cortex. Cell adhesion was measured for 50-70% confluent cells with N-Cadherin coated stylus AFM probes on an AFM mounted on an inverted microscope. Our results suggested that fluvastatin and MβCD mediated cholesterol depletion increased N-cadherin mediated cell adhesion force by approx. 27% and 17% respectively. In addition, fluvastatin and MβCD respectively reduced VSMCs stiffness by approx. 24% and 29 % compared to control VSMCs. Altogether, the knowledge that we obtained in this project may lead to a novel therapeutic strategy that could potentially control and block VSMC migration and prevent atherosclerotic plaque formation by deciphering and regulating N-cadherin mediated adhesion