Cellular cholesterol flux studies: methodological considerations

Reverse cholesterol transport (RCT) is the process in which peripheral cells release cholesterol to an extracellular acceptor such as high-density lipoprotein (HDL) which then mediates cholesterol delivery to the liver for excretion. RCT represents a physiological mechanism by which peripheral tissues are protected against excessive accumulation of cholesterol. The first step in RCT is the interaction of the cell with lipoprotein particles, a process that results in both the cellular uptake and release of cholesterol. The various components of this cholesterol flux can be viewed as efflux, influx and net flux. Experimental protocols for measuring each of these components of cholesterol flux are very different, and a number of considerations are required to design experimental approaches for the quantitation of flux parameters. Although many flux studies have been conducted in the past, the recent discoveries of the scavenger receptor B1 (SR-B1) and ATP binding cassette 1 (ABCA1), which mediate the movement of cholesterol between cells and extracellular acceptors, has led to increased interest in studies of cellular cholesterol flux. The aim of this review is to present a discussion of the methodological considerations that should be evaluated during the design and analysis of cellular cholesterol flux experiments.