The effects of membrane cholesterol on the mechanics of vascular smooth muscle cell

Abstract

As the major cellular component of blood vessel wall, vascular smooth muscle cell (VSMC) plays important roles at each stage of atherosclerosis by proliferation and migration toward the inner layer of blood vessel wall. It is also known that cholesterol is not only the major factor involved in fatty deposition in the atherosclerotic lesion, but also plays an important role as a regulator of cell mechanics, spreading, and migration. A further complication is the mechanical environment of the vascular wall, which undergoes progressive stiffening with aging and hypertension and thus accelerates development of atherosclerosis. Therefore, it is important to investigate the combined effect of cholesterol and mechanical environment on VSMC and determine how it alters VSMC functional characteristics such as cell stiffness and adhesion. VSMCs used in this study were enzymatically isolated from the descending aorta of male Sprague‐Dawley rats and cultured in DMEM with 10% FBS in a humidified incubator at 37°C and 5% CO2. Cell adhesion was measured for 50–70% confluent cells with an atomic force microscope (AFM) system and the fibronectin (FN) or N‐cadherin coated stylus AFM probes at 100 nm/sec retraction speed. Cell stiffness was tested with spherical AFM probe (5μm diameter) at 1 μm/sec indentation speed. Cells were treated with 5mM Methyl‐β‐cyclodextrin for 1 h for cholesterol depletion prior to the AFM experiment. Our results showed that cholesterol depletion significantly increased N‐cadherin mediated cell adhesion by ~20% and adhesion probability by ~35%, respectively. While α5β1/FN adhesion force significantly decreased upon membrane cholesterol depletion. Moreover, our results suggested that cholesterol depletion significantly reduced VSMCs stiffness by 47% compared to control cells. All together, these results reveal that membrane cholesterol is a key regulator for cellular stiffness and integrin/cadherin‐mediated cell adhesion. 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 the formation of atherosclerotic plaque by regulating integrin and/or cadherin mediated cell adhesion.Support or Funding Information15SDG25420001 (AHA)