Endothelial Cell Membrane Sensitivity to Shear Stress is Lipid Domain Dependent

Blood flow-associated shear stress causes physiological and pathophysiological biochemical processes in endothelial cells that may be initiated by alterations in plasma membrane lipid domains characterized as liquid-ordered (l(o)), such as rafts or caveolae, or liquid-disordered (l(d)). To test for domain-dependent shear sensitivity, we used time-correlated single photon counting instrumentation to assess the photophysics and dynamics of the domain-selective lipid analogues DiI-C-12 and DiI-C-18 in endothelial cells subjected to physiological fluid shear stress. Under static conditions, DiI-C-12 fluorescence lifetime was less than DiI-C-18 lifetime and the diffusion coefficient of DiI-C-12 was greater than the DiI-C-18 diffusion coefficient, confirming that DiI-C-12 probes l(d), a more fluid membrane environment, and DiI-C-18 probes the l(o) phase. Domains probed by DiI-C-12 exhibited an early (10 s) and transient decrease of fluorescence lifetime after the onset of shear while domains probed by DiI-C-18 exhibited a delayed decrease of fluorescence lifetime that was sustained for the 2 min the cells were subjected to flow. The diffusion coefficient of DiI-C-18 increased after shear imposition, while that of DiI-C-12 remained constant. Determination of the number of molecules (N) in the control volume suggested that DiI-C-12-labeled domains increased in N immediately after step-shear, while N for DiI-C-18-stained membrane transiently decreased. These results demonstrate that membrane microdomains are differentially sensitive to fluid shear stress.