Glucocorticoid effects on lipid lateral diffusion and membrane composition in lipopolysaccharide-activated B-cell...

We have investigated the effects of glucocorticoid treatment on lipid lateral diffusion and cholesterol:phospholipid ratios in the plasma membranes of the lipopolysaccharide (LPS)-responsive murine B-cell leukemia (BCL1). Exposure to LPS for 24 h at concentrations of 50 micrograms/ml, 100 micrograms/ml, and 250 micrograms/ml caused a 42% reduction in the lateral diffusion of the lipid probe 3,3'-dioctadecylindocarbocyanine iodide as measured by fluorescence photobleaching recovery techniques at 37 degrees C. In cells incubated with 50 micrograms/ml LPS, 3,3'-dioctadecylindocarbocyanine iodide diffusion is reduced to 2.2 x 10(-9) cm2s-1 compared to freshly isolated BCL1 cells where 3,3'-dioctadecylindocarbocyanine iodide diffused at a rate of 3.8 x 10(-9) cm2s-1. In BCL1 cells activated by LPS for 24 h and recultured with a pharmacological concentration of the synthetic glucocorticoid triamcinolone acetonide (TA, 10(-6) M) for 6 h, lipid lateral diffusion increased to 3.5 x 10(-9) cm2s-1. Concentrations of TA lower than 10(-6) M had no effect on lipid lateral diffusion. Six-h treatment with 10(-6) M TA had no effect on freshly isolated BCL1 cells (Time 0). However, lipid lateral diffusion increased in cells incubated 24, 48, and 72 h with LPS, and an additional 6 h with 10(-5) M TA suggested that activated BCL1 cells were more glucocorticoid sensitive than cells at Time 0. Plasma membrane cholesterol and phospholipid content, analyzed at 0 and 24 h, indicated that LPS activation was associated with a 7% increase in the cholesterol:phospholipid ratio in BCL1 membranes and that glucocorticoid treatment of these LPS-activated cells for 6 h decreased the membrane cholesterol:phospholipid ratio perhaps through inhibition of de novo cholesterol synthesis.