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 liquidordered (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-C₁₈ in endothelial cells subjected to physiological fluid shear stress. Under static conditions, DiI-C₁₂ fluorescence lifetime was less than DiI-C₁₈ lifetime and the diffusion coefficient of DiI-C₁₂ was greater than the DiI-C ₁₈ diffusion coefficient, confirming that DiI-C₁₂ probes l_d, a more fluid membrane environment, and DiI-C₁₈ probes the l_o phase. Domains probed by DiI-C₁₂ exhibited an early (10 s) and transient decrease of fluorescence lifetime after the onset of shear while domains probed by DiI-C₁₈ 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-C₁₈ increased after shear imposition, while that of DiI-C₁₂ 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-C₁₈-stained membrane transiently decreased. These results demonstrate that membrane microdomains are differentially sensitive to fluid shear stress.