Painter R G, Ginsberg M H
Exp Cell Res. 1984 Nov;155(1):198-212. doi: 10.1016/0014-4827(84)90781-x.
We have examined the F-actin and myosin distribution in resting and thrombin-activated platelets by double label immunofluorescence microscopy. In resting, discoid platelets, F-actin and myosin staining was distributed in a diffuse pattern throughout the interior of the cell with slight accentuation at the cell periphery. In contrast, platelet factor 4 antigen (PF4) was more centrally localized in a fine punctate distribution which is consistent with its localization in alpha-granules. Within 5 sec after thrombin stimulation both F-actin and myosin staining were increased at the periphery of the now spherical platelets. Subsequently, a myosin-containing spherical structure decreased in diameter closely surrounding a phase-dense central zone. In contrast, F-actin staining continued to be accentuated at the cell periphery and was prominent in filopodia and blebs. As previously shown, PF4 staining was localized after 30 sec within large intracellular masses that corresponded to closed vacuolar structures at the ultrastructural level. Morphometric analysis of electron micrographs showed that formation of these vacuolar structures kinetically paralleled alpha-granule disappearance and preceded PF4 release. These PF4-containing structures translocated to the cell periphery after 1-3 min, where they appeared to fuse with the plasma membrane. Ultrastructural analysis of thin sections showed that the myosin-rich spherical structure spatially and temporally correlated with a band of microfilaments that closely surrounded the organelle-rich central zone of the cell. Morphometric analysis of these micrographs showed that the absolute volume of this central zone decreased with time after thrombin addition, showing a significant change after 15 sec and reaching a maximum value after 3-5 min. Changes in the volume of this compartment kinetically preceded PF4 release. On the basis of these data, we propose that an actomyosin contractile force is generated which centripetally redistributes the myosinrich structure and organelle zone. Conceivably this inward force may not only accelerate granule-granule fusion to form intracellular secretory vacuoles, but may also provide aid in their extrusion toward the platelet plasma membrane.
我们通过双标记免疫荧光显微镜检查了静息和凝血酶激活血小板中的F-肌动蛋白和肌球蛋白分布。在静息的盘状血小板中，F-肌动蛋白和肌球蛋白染色以弥漫模式分布于整个细胞内部，在细胞周边略有增强。相比之下，血小板因子4抗原（PF4）更集中地定位于精细的点状分布中，这与其在α-颗粒中的定位一致。凝血酶刺激后5秒内，F-肌动蛋白和肌球蛋白染色在现在呈球形的血小板周边均增加。随后，一个含肌球蛋白的球形结构直径减小，紧密围绕一个相致密的中央区。相比之下，F-肌动蛋白染色在细胞周边持续增强，在丝状伪足和小泡中很突出。如先前所示，PF4染色在30秒后定位于大的细胞内团块中，这些团块在超微结构水平上对应于封闭的液泡结构。电子显微镜照片 的形态计量分析表明，这些液泡结构的形成在动力学上与α-颗粒消失平行，并先于PF4释放。这些含PF4的结构在1-3分钟后转移到细胞周边，在那里它们似乎与质膜融合。薄切片 的超微结构分析表明，富含肌球蛋白的球形结构在空间和时间上与紧密围绕细胞富含细胞器的中央区的微丝带相关。这些显微镜照片 的形态计量分析表明，添加凝血酶后该中央区的绝对体积随时间减少，在15秒后显示出显著变化，并在3-5分钟后达到最大值。该区室体积的变化在动力学上先于PF4释放。基于这些数据，我们提出产生了一种肌动球蛋白收缩力，该力向心重新分布富含肌球蛋白的结构和细胞器区。可以想象，这种向内的力不仅可以加速颗粒-颗粒融合以形成细胞内分泌泡，还可以帮助它们向血小板质膜挤出。
