FACTORS UNDERLYING THE VARIABILITY OF LIPID DROPLET FLUORESCENCE IN MA-10 LEYDIG TUMOR-CELLS

Neutral lipids accumulate in cellular lipid droplets. These droplets v ary remarkably in number and amount between cells. In the present stud ies, the variability in lipid content was quantified by comparing the coefficient of variation of fluorescence histograms of nile red lipid- stained cells to the variability of cell size or cell protein distribu tions. This measure of lipid droplet variability persisted through a w ide range of cell lipid content and averaged 4.4-fold more variability than cell size and 2.6-fold more variability than cell protein conten t. While looking for possible explanations for this variability, it wa s determined that cell to cell variability could not be explained by m ultiple clonal populations of cells or the confluence of the cell mono layer. Analysis of lipid variability using a more droplet-specific flu orescent dye, bodipy, reduced variability by about 44%. Cell cycle ana lysis revealed that G(2) + M cells contained more lipid than S-phase c ells, which in turn contained more lipid than G(0) + G(1) cells, but t hat variability was equally large throughout the cell cycle. The chole steryl ester hydrolase inhibitor, diethylumbelliferyl phosphate, inhib ited hydrolysis of both cholesteryl esters and triglycerides. Lipid co ntent of diethylumbelliferyl phosphate-treated cells increased while t he variability in lipid staining decreased by an average of 72%. Thus, the excess lipid fluorescence variability compared to cell size or pr otein fluorescence could in part be explained by variability in cellul ar hydrolysis of triglyceride and cholesteryl ester. Excess lipid fluo rescence variability could be reduced by an average of 44% when a more lipid droplet-specific stain was used instead of nile red. (C) 1994 W iley-Liss, Inc.