Abstract
Differential scanning calorimetry and polarizing light microscopy have been used to investigate kinetic and thermodynamic properties of the phase behavior of cholesteryl ester contained in Fu5AH rat hepatoma cells and J774 murine macrophages. These cultured cells store cholesteryl esters as cytoplasmic inclusions of approximately 1-micron diameter and thus are models of the foam cells characteristic of atherosclerotic plaque. Simple binary mixtures of cholesteryl palmitate and cholesteryl oleate, the predominant cholesteryl esters in cellular inclusions in both cell types serve as models to explain important aspects of the phase behavior of these inclusions. Although inclusions should exist as stable crystals at 37 degrees C under conditions of thermodynamic equilibrium, microscopic examination of cells indicates that inclusions exist as metastable liquid crystals at 37 degrees C for extended periods of time. Using an analytical model based on nucleation theory, we predict that the cholesteryl ester inclusions should be liquid-crystalline in the cytoplasm of living cells. This may not be true either for lysosomal cholesteryl ester or for extracellular cholesteryl ester present in advanced atherosclerotic plaque where fusion of droplets can enhance the possibility of crystallization. The enhanced metastability of the relatively fluid liquid-crystalline state in cellular inclusions should result in increased activity of the neutral cholesteryl ester hydrolase in living cells.
Highlights
The results showed that during the subsequent scan only a liquid-crystalline transition was observed, which indicates that thseample had notcrystallized when cooled to this intermediate temperature, as was the case for both hepatoma cells and isolated inclusions
We found that after being exposed to one such thermal history, defined as 1 h at 25 “C, both hepatoma and macrophage inclusions were in a liquid-crystalline state
We indicated in an earlier report [6]that microcrystals of CP would exist in the CE-rich inclusions in living macrophages if the phase behavior were under thermodynamic control
Summary
From the Departmentof Chemistry, PhiladelphiaCollege of Pharmacy and Science, Philadelphia, Pennsylvania19104-4495. The presence of an extracellular acand thermodynamic properties of the phase behavior ceptorparticle such as high density lipoprotein promotes of cholesteryl ester contained iFnuSAH rat hepatoma cells and 5774 murine macrophages These cultured cells store cholesterylesters as cytoplasmic inclusions of approximately 1-pm diameter and are models ofthefoam cells characteristic of atherosclerotic plaque. The results indicate that cytoplasmic inclusions are in a metastable liquid-crystalline state in living cells This conclusion is in accord with earlier findings on intracellular cholesteryl ester-rich droplets. For cholesterol enrichment,cells were exposed to 10 ml of RPMI 1640 medium containing acetylatedLDL a t a concentration of 50 pg/ml LDL protein, 225 pg/ml FC as FC/PC dispersions, and 0.1% bovine serum albumin for 3 days at 37 "C Under these conditions, 5774 macrophages accumulated 75-100pgof esterified cholesterol/mg of cell protein. Light Microscopy-Evidence for lipid anisotropy was obtained visually by examining whole cells using a Zeiss polarizing microscope (X1250 magnification) equipped with ahotstage (OPT0 systems, Jenkintown, PA)
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