Department of Pharmaceutics, Faculty of Pharmaceutical Sciences, Hokkaido University of Science, 7-Jo 15-4-1 Maeda, Teine, Sapporo, Hokkaido 006-8585, Japan; Division of Pharmaceutics, Hokkaido Pharmaceutical University School of Pharmacy, 7-Jo 15-4-1 Maeda, Teine, Sapporo, Hokkaido 006-8585, Japan.
Division of Pharmaceutics, Hokkaido Pharmaceutical University School of Pharmacy, 7-Jo 15-4-1 Maeda, Teine, Sapporo, Hokkaido 006-8585, Japan.
Int J Pharm. 2019 May 1;562:218-227. doi: 10.1016/j.ijpharm.2019.03.032. Epub 2019 Mar 19.
To develop a three-dimensional visualization method for evaluating the distribution of pulmonary drug delivery systems and compare four tissue-clearing techniques (Clear, CUBIC, ScaleS, and SeeDB2) using intrapulmonary liposomes as drug carriers.
Rhodamine B-labeled liposomes were administered intrapulmonarily to mice using a MicroSprayer, and then fluorescent-labeled tomato lectin was administered intravenously to visualize the general lung structure. Tissue-clearing treatment of the mouse lungs was performed using the standard protocols of the Clear, CUBIC, ScaleS, and SeeDB2 techniques. Lung clearing was clarified using laser-scanning confocal microscopy, and three-dimensional images were reconstructed.
Fluorescent-labeled tomato lectin was preserved using Clear and SeeDB2 but not using CUBIC and ScaleS. In addition, the liposomes were stable in Clear reagent, but they were mostly degraded in other reagents by surface-active agents. Clear treatment enabled the three-dimensional visualization of intrapulmonary rhodamine B-labeled liposomes at the alveolar scale.
These results suggest that the Clear tissue-clearing technique was appropriate for the three-dimensional visualization of intrapulmonary liposomes at the alveolar scale. This study provides important information for selecting and optimizing suitable optical tissue-clearing techniques in lungs for evaluating the distribution of pulmonary drug delivery systems.
开发一种三维可视化方法，用于评估肺部药物传递系统的分布，并使用肺部内脂质体作为药物载体比较四种组织透明化技术（Clear、CUBIC、ScaleS 和 SeeDB2）。
使用 MicroSprayer 将罗丹明 B 标记的脂质体经肺部给药，然后静脉内给予荧光标记的番茄凝集素以可视化肺部的一般结构。使用 Clear、CUBIC、ScaleS 和 SeeDB2 技术的标准方案对小鼠肺部进行组织透明化处理。使用激光扫描共聚焦显微镜阐明肺部透明化，并重建三维图像。
荧光标记的番茄凝集素使用 Clear 和 SeeDB2 得以保留，但 CUBIC 和 ScaleS 则不行。此外，脂质体在 Clear 试剂中稳定，但在其他试剂中由于表面活性剂的作用大部分降解。Clear 处理使肺部内罗丹明 B 标记的脂质体在肺泡尺度上的三维可视化成为可能。
这些结果表明，Clear 组织透明化技术适合于肺泡尺度上肺部内脂质体的三维可视化。本研究为选择和优化用于评估肺部药物传递系统分布的肺部光学组织透明化技术提供了重要信息。
