Arora Neha, Syed Aleem, Sander Suzanne, Smith Emily A
Department of Chemistry, Iowa State University, 1605 Gilman Hall, Ames, Iowa 50011, USA.
Phys Biol. 2014 Oct 7;11(6):066001. doi: 10.1088/1478-3975/11/6/066001.
A combination of sterol modulation with cyclodextrins plus fluorescence microscopy revealed a biophysical mechanism behind cholesterol's influence on the diffusion of a ubiquitous class of receptors called integrins. The heterogeneous diffusion of integrins bound to ligand-coated quantum dots was measured using single particle tracking (SPT), and the ensemble changes in integrin diffusion were measured by fluorescence recovery after photobleaching (FRAP). A 25 ± 1% reduction of membrane cholesterol resulted in three significant changes to the diffusion of ligand-bound αPS2CβPS integrins as measured by SPT. There was a 23% increase in ligand-bound mobile integrins; there was a statistically significant increase in the average diffusion coefficient inside zones of confined diffusion, and histograms of confined integrin trajectories showed an increased frequency in the range of 0.1-1 μm(2) s(-1) and a decreased frequency in the 0.001-0.1 μm(2) s(-1) range. No statistical change was measured in the duration of confinement nor the size of confined zones. Restoring the cholesterol-depleted cells with exogenous cholesterol or exogenous epicholesterol resulted in similar diffusion properties. Epicholesterol differs from cholesterol in the orientation of a single hydroxyl group. The ability of epicholesterol to substitute for cholesterol suggests a biophysical mechanism for cholesterol's effect on integrin diffusion. Influences of bilayer thickness, viscosity and organization are discussed as possible explanations for the measured changes in integrin diffusion when the membrane cholesterol concentration is reduced.
将甾醇调节与环糊精相结合，并运用荧光显微镜技术，揭示了胆固醇对一类名为整合素的普遍存在的受体扩散产生影响背后的生物物理机制。使用单粒子追踪（SPT）测量与配体包被量子点结合的整合素的异质扩散，并通过光漂白后荧光恢复（FRAP）测量整合素扩散的整体变化。通过SPT测量，膜胆固醇降低25±1%导致与配体结合的αPS2CβPS整合素的扩散发生三个显著变化。与配体结合的可移动整合素增加了23%；在受限扩散区域内，平均扩散系数有统计学意义的增加，并且受限整合素轨迹的直方图显示，在0.1 - 1μm² s⁻¹范围内频率增加，在0.001 - 0.1μm² s⁻¹范围内频率降低。在受限持续时间和受限区域大小方面未测量到统计学变化。用外源性胆固醇或外源性表胆固醇恢复胆固醇耗尽的细胞会导致类似的扩散特性。表胆固醇与胆固醇的区别在于单个羟基的取向。表胆固醇替代胆固醇的能力表明了胆固醇对整合素扩散影响的生物物理机制。讨论了双层厚度、粘度和组织的影响，作为膜胆固醇浓度降低时整合素扩散测量变化的可能解释。
