Imaging dynamic cell-cell junctional coupling in vivo using Trojan-LAMP
Imaging dynamic cell-cell junctional coupling in vivo using Trojan-LAMP
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DOI：
10.1038/nmeth.1238
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发表时间：
2008-09-01
期刊：
影响因子：
48
通讯作者：
Li, Wen-hong
中科院分区：
文献类型：
--
作者：
Guo, Yan-Ming;Chen, Shiuhwei;Li, Wen-hong
为了研究体内细胞间缝隙连接通讯的生理调节和功能，我们开发了一种笼状染料的生物缀合物，命名为dextran-CANPE-HCC，用于在小模型生物中成像细胞偶联。在体外，该化合物被有效地光解，具有稳健的荧光增强。递送到秀丽隐杆线虫卵母细胞中的右旋糖酐-CANPE-HCC在整个发育过程中保留在细胞中。使用局部释放，我们光解葡聚糖-CANPE-HCC以释放小HCC染料，并通过间隙连接通道成像细胞间染料转移的动力学，我们将该技术命名为特洛伊局部激活分子荧光探针（LAMP）。在胚胎发育的早期，细胞偶联的模式经历了戏剧性的重塑和成像显示，生殖细胞前体，P2，P3和P4，是从体细胞通讯室分离。由于右旋糖酐-CANPE-HCC在化学和代谢上是稳定的，标记的蠕虫在孵化后光活化时显示出非常明亮的信号，这使得我们能够非侵入性地检查活蠕虫中的细胞偶联。
To study the physiological regulation and function of cell-cell gap junction communication in vivo, we developed a bioconjugate of caged dye, named dextran-CANPE-HCC, for imaging cell coupling in small model organisms. In vitro, the compound was photolyzed efficiently with robust fluorescence enhancement. Dextran-CANPE-HCC delivered into Caenorhabditis elegans oocytes was retained in cells throughout development. Using local uncaging, we photolyzed dextran-CANPE-HCC to release the small HCC dye and imaged the dynamics of intercellular dye transfer through gap junction channels, a technique we named Trojan-local activation of molecular fluorescent probes (LAMP). Early during embryonic development, the pattern of cell coupling undergoes dramatic remodeling and imaging revealed that the germ cell precursors, P2, P3 and P4, were isolated from the somatic cell communication compartment. As dextran-CANPE-HCC is chemically and metabolically stable, labeled worms showed very bright signal upon photoactivation after hatching, which allowed us to examine cell coupling in living worms noninvasively.