Differential sensitivity of astrocyte primary cultures and derived spontaneous transformed cell lines to 70-hydroxycholesterol: effect on plasma membrane lipid composition and fluidity, and on cell surface protein expression

Summary

The cytotoxicity of 7β-hydroxycholesterol (7β-OHC) was investigated on rat astrocyte primary cultures and spontaneously transformed cell lines derived from them. Confluent astrocyte primary cultures (normal cells) were unaffected by 20 µM 7(3-OHC over a period of 72 h whereas 30 µM markedly affected the viability of the transformed cells within the first 72 h. Both cell types incorporated 18% of the total amount of 7β-OHC added to the cultures at concentrations of 20 µM or 30 µM. Cellular fractionation after incubation with 20 µM or 30 µM 7β-OHC indicated that the plasma membrane incorporated 2 or 6 fold more 7β-OHC than the intracellular one's respectively. Plasma membrane cholesterol (CH) and phospholipid (PL) analysis showed that 20 µM 7β-OHC did not affect CH/PL in normal cells; in contrast, plasma membranes of transformed cells displayed a significant CH/PL decrease, which was more pronounced with 30 µM 7β-OHC treatment. Fluorescence anisotropy measurements indicated that 20 µM 7β-OHC slightly fluidified the plasma membrane of normal cells whereas it has not effect on that of the transformed cells one; however, an increase in plasma membrane fluidity was observed when the transformed cells were treated with 30 µM 7β-OHC. Lactoperoxidase catalyzed radioiodination of cell surface proteins and subsequent autoradioelectrophoretic analysis demonstrated that the labelled protein pattern was unchanged when both cell types were incubated with 30 µM 7β-OHC.