Activation of human lymphocytes by concanavalin A or purified protein derivative results in no alteration of fluorescence polarization of lipid probes although the electrophoretic mobility of the cells is changed

Abstract

Upon stimulation with either concanavalin A or the tuberculin antigen, purified protein derivative, human peripheral blood lymphocytes, purified on Ficoll-Hypaque, did not exhibit a concomitant lipid fluidity alteration as measured by fluorescence polarization (P) of the lipid probe, 1,6-diphenyl-1,3-5-hexatriene (DPH). This result was independent of the incubation period, ranging from 10 min to 72 h. However, a general reduction in polarization value, from P = 0.287 (maintained for up to 2 h of incubation) to P = 0.225 after 20 h was observed for both experimental and control samples. Moreover, fluorescence polarization studies of the nonpenetrating modified DPH cationic lipid probe, 1-[4'-trimethylaminophenyl]-6-phenyl-1,3-5-hexatriene (TMA-DPH), also failed to show any change in lipid fluidity subsequent to a 1-3 h incubation of lymphocytes with concanavalin A. Cell electrophoretic mobility, however, was altered (mean cell mobility increased by 10-15%) in a fast response to stimulation and was observed within several hours of in vitro application of concanavalin A and purified protein derivative. This initial response disappeared with further incubation at 37 degrees C (greater than 3 h) and was followed by a decline of cellular mobility of the concanavalin A-exposed cells after 48 and 72 h of incubation. The unstimulated control cells did not change in mobility as a function of incubation time. The slow decline in mean cell mobility of the experimental cells is believed to be associated with blastogenesis. It is concluded that neither blastogenic transformation nor short term membrane alterations associated with human lymphocyte activation lead to lipid fluidity changes as measured in steady state by the fluorescence polarization of both DPH and TMA-DPH.