Centre National de la Recherche Scientifique-Institut de Pharmacologie et de Biologie Structurale, Unité Mixte de Recherche 5089, Université de Toulouse, Université Paul Sabatier, F-31077 Toulouse, France.
Proc Natl Acad Sci U S A. 2010 Dec 7;107(49):21016-21. doi: 10.1073/pnas.1007835107. Epub 2010 Nov 16.
Podosomes are unique cellular entities specifically found in macrophages and involved in cell-matrix interactions, matrix degradation, and 3D migration. They correspond to a core of F-actin surrounded at its base by matrix receptors. To investigate the structure/function relationships of podosomes, soft lithography, atomic force microscopy (AFM), and correlative fluorescence microscopy were used to characterize podosome physical properties in macrophages differentiated from human blood monocytes. Podosome formation was restricted to delineated areas with micropatterned fibrinogen to facilitate AFM analyses. Podosome height and stiffness were measured with great accuracy in living macrophages (578 ± 209 nm and 43.8 ± 9.3 kPa) and these physical properties were independent of the nature of the underlying matrix. In addition, time-lapse AFM revealed that podosomes harbor two types of overlapping periodic stiffness variations throughout their lifespan, which depend on F-actin and myosin II activity. This report shows that podosome biophysical properties are amenable to AFM, allowing the study of podosomes in living macrophages at nanoscale resolution and the analysis of their intimate dynamics. Such an approach opens up perspectives to better understand the mechanical functionality of podosomes under physiological and pathological contexts.
足突是一种独特的细胞实体，仅在巨噬细胞中发现，参与细胞-基质相互作用、基质降解和 3D 迁移。它们对应于一个 F-肌动蛋白核心，其基底周围有基质受体。为了研究足突的结构/功能关系，使用软光刻、原子力显微镜（AFM）和相关荧光显微镜来表征从人血液单核细胞分化而来的巨噬细胞中的足突物理特性。足突的形成仅限于用微图案化纤维蛋白原限定的区域，以方便 AFM 分析。在活巨噬细胞中，足突的高度和刚度可以非常精确地测量（578 ± 209nm 和 43.8 ± 9.3kPa），这些物理特性与底层基质的性质无关。此外，延时 AFM 揭示了足突在其整个生命周期中存在两种类型的重叠周期性刚度变化，这取决于 F-肌动蛋白和肌球蛋白 II 的活性。本报告表明，足突的生物物理特性适合 AFM，可以在纳米尺度分辨率下研究活巨噬细胞中的足突，并分析其紧密动力学。这种方法为在生理和病理环境下更好地理解足突的机械功能提供了新的视角。
