Relation between membrane fluidity and signal transduction in the human megakaryoblastic cell line MEG-01

Abstract

The fluidity of the plasma membrane is thought to affect the responsiveness of blood platelets. We measured membrane fluidity in a single cell by Fluorescence Recovery after Photobleaching (FRAP) of the lipophilic probe DiIC 14. Since platelets are too small for this technique, we used the human megakaryoblastic cell-line MEG-O1, which shares many properties with platelets. MEG-01 cells were cultured for 44 h with simvastatin or mevalonate to change the cholesterol content, enabling analysis of signal processing at cholesterol /phospholipid ratios (C/P) between 0.20 and 0.31. The diffusion of DiIC,4 correlated inversely with the C/P ratio with lateral diffusion coefficients (D) of 3.28 · 10 −9 cm 2/s at a low C/P decreasing to 2.55 · 10 −9 cm 2/s at a high C/P ratio. The mobile fraction was 65% and constant at the different C/P ratios. The relation between lipid diffusion and signal processing was measured following stimulation with 10 U/ml thrombin at 22°C There were only little differences in phosphatidylinositol metabolism, Ca 2+ influx or mobilization and prostaglandin I 2-induced formation of cyclic AMP. At 37°C cells with a high C/P ratio showed increased phosphatidylinositol metabolism, but these differences had no major effect on the Ca 2+ responses. These data demonstrate that in megakaryoblasts the lateral diffusion of lipids is inversely correlated with the C/P ratio, but within the range of 0.20–0.31 the influence on signal processing is minor.