Sequestration and release of polycyclic aromatic hydrocarbons by vertebrate cells in vitro

Vertebrate fibroblasts grown in vitro and exposed to various concentrations of the mutagen/carcinogen benzo(a)pyrene (B(a)P) internalized the compound and recrystallized it in lysosomes by 6-18 h postexposure. This phenomenon occurred when B(a)P at or above 10 micrograms/ml was introduced to the culture medium in a solvent such as DMSO or acetone but not when introduced dissolved in serum. Likewise, high fetal bovine serum concentrations in the culture medium (greater than 20%) as well as human serum (10%) inhibited crystal formation, presumably owing to lipid competition for the compound. Electron-microscopic observations of the cells during the uptake and crystal forming periods revealed that the cell membrane became altered within 3 h of exposure to B(a)P. This was followed by a return to normal of the membrane and the appearance of vesicles within the cell by 6-8 h. The vesicles then became filled with crystals which continued to grow while in the presence of B(a)P and disappeared when the cells were exposed to B(a)P-free culture medium or serum lipids. Introduction of B(a)P into culture medium containing delipidated serum resulted in crystal formation indistinguishable from that which occurred when whole serum was present. Crystals were not formed when total lipoprotein and the lipoprotein components VLDL, LDL, HDL2, and HDL3 were used as solvents for B(a)P. The process of crystal formation was inhibited by the addition of 10(-3) M KCN, and removal of the crystals was dependent on the concentration of lipoprotein in the culture medium.