In Vivo Nano-imaging of Membrane Dynamics in Metastatic Tumor Cells Using Quantum Dots
In Vivo Nano-imaging of Membrane Dynamics in Metastatic Tumor Cells Using Quantum Dots
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DOI：
10.1074/jbc.m109.075374
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发表时间：
2010-01-22
影响因子：
4.8
通讯作者：
Higuchi, Hideo
中科院分区：
文献类型：
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作者：
Gonda, Kohsuke;Watanabe, Tomonobu M.;Higuchi, Hideo
膜形态的改变和基于其流动性的膜蛋白动力学是肿瘤转移的关键。然而，由于以往的体内成像的空间精度仅限于微米水平，单分子成像是不可能的，这个问题一直不清楚。在这里，我们在共聚焦显微镜下以7-9 nm的空间精度对小鼠肿瘤细胞的膜动力学进行了成像。细胞膜上的转移促进因子蛋白酶激活受体1 （PAR1）被量子点标记为抗PAR1抗体。在体内清楚地观察到癌细胞和PAR1在转移过程中的运动。用于评估PAR1动态的图像取自四个转移阶段的代表性细胞；即拍摄肿瘤内远离血管的癌细胞、靠近血管的癌细胞、血流中的癌细胞以及肿瘤附近正常组织内粘附在血管内壁的癌细胞。PAR1在远离肿瘤血管的静止细胞中的扩散常数小于靠近血管和血流的运动细胞。正常组织中细胞粘附在血管内壁的扩散常数也很小。细胞在迁移过程中形成膜突。这些假足上的PAR1扩散常数大于同一细胞内其他膜区。因此，PAR1运动的动力学表明，膜流动性在内渗过程中增加，在血管内达到峰值，在外渗过程中减少，在局部形成的假足处也更高。
Changes in membrane morphology and membrane protein dynamics based on its fluidity are critical for cancer metastasis. However, this subject has remained unclear, because the spatial precision of previous in vivo imaging has been limited to the micrometer level and single molecule imaging is impossible. Here, we have imaged the membrane dynamics of tumor cells in mice with a spatial precision of 7-9 nm under a confocal microscope. A metastasis-promoting factor on the cell membrane, protease-activated receptor 1 (PAR1), was labeled with quantum dots conjugated with an anti-PAR1 antibody. Movements of cancer cells and PAR1 during metastasis were clearly observed in vivo. Images used to assess PAR1 dynamics were taken of representative cells for four stages of metastasis; i.e. cancer cells far from blood vessels in tumor, near the vessel, in the bloodstream, and adherent to the inner vascular surface in the normal tissues near tumor were photographed. The diffusion constant of PAR1 in static cells far from tumor blood vessels was smaller than in moving cells near the vessels and in the bloodstream. The diffusion constant of cells adhering to the inner vascular surface in the normal tissues was also very small. Cells formed membrane protrusion during migration. The PAR1 diffusion constant on these pseudopodia was greater than in other membrane regions in the same cell. Thus, the dynamics of PAR1 movement showed that membrane fluidity increases during intravasation, reaches a peak in the vessel, decreases during extravasation, and is also higher at locally formed pseudopodia.