Membrane microviscosity and human platelet function

Abstract

An increased sensitivity to epinephrine-induced aggregation was observed in platelets obtained from patients with type IIa hyperlipoproteinemia and in normal platelets following incubation with cholesterol-rich lecithin dispersions. It was reported previously that the membrane fraction of platelets is enriched with cholesterol relative to phospholipid under each of these conditions. The fluidity (microviscosity) of whole platelets and platelet subcellular fractions was examined using a hydrophobic fluorescent probe, 1,6-diphenyl-1,3,5-hexatriene (DPH), under conditions in which the cholesterol-to-phospholipid mole ratio (C/PL) of platelets was varied by incubation with various cholesterol-lecithin sonicated dispersions. The C/PL of platelets directly influenced the rotational diffusion of DPH, as indicated by changes in fluorescence polarization. This was reflected in an increase in microviscosity at 37.degree. C from (.hivin..eta.37) from 2.84 P in normal platelets to 4.06 P in platelets with a 118% increase in C/PL. Conversely, platelets with a 43% decrease in C/PL had a 13% decrease in .hivin..eta.37. A strong correlation (r = 0.94) existed between C/PL and .hivin..eta.37 throughout this entire range. C/PL had no effect on the excited-state fluorescence lifetime of DPH. Both C/PL and .hivin..eta.37 were lower in isolated platelet membranes than in the platelet granule fraction. When platelets were incubated for 20 h with cholesterol-rich dispersions, there was an increase in C/PL and .hivin..eta.37 in both the membrane and granule fractions. This occurred more rapidly in membranes so that, at 5 h (a time when an increased sensitivity of whole platelets to epinephrine is evident), membrane C/PL had increased 55% and .hivin..eta.37 had increased 42%, but granule C/PL and had changed minimally. Cholesterol-rich platelets and subcellular fractions had a lower fusion (or flow) activation energy for viscosity (.DELTA.E), reflecting a higher degree of order, and the converse was true in cholesterol-poor platelets. A strong negative correlation existed between the percent change in .DELTA.E and the percent change in .DELTA.E and the percent change in .hivin..eta.37 induced either by cholesterol incorporation or depletion. Cholesterol apparently influences the fluidity and the degree of order within the hydrophobic core of platelet membranes. Changes induced in these physical properties by an excess of cholesterol relative to phospholipid may underlie the abnormal reception or transmission of the aggregation stimulus in cholesterol-rich platelets.