Molecular and Cell Therapy Program, Division of Experimental Hematology & Cancer Biology, Cancer and Blood Diseases Institute, Cincinnati Children's Hospital Medical Center, Cincinnati, OH, USA.
Mol Biotechnol. 2013 Jan;53(1):63-73. doi: 10.1007/s12033-012-9519-6.
A naturally secreted Gaussia luciferase (Gluc) has been utilized as a reporter for bioluminescence imaging (BLI) evaluation. However, the potential application of Gluc for in vivo monitoring of systemic protein delivery, as well as its natural biodistribution, has not been studied. To examine Gluc secretion and uptake profile, we injected Gluc-encoding plasmids into mice by hydrodynamic tail-vein injection. Whole-body BLI showed that imaging quantification obtained at pawpad was directly correlated to blood Gluc activities. When gene expression was restricted to the liver by the use of a hepatic promoter, in vivo Gluc biodistribution analysis revealed the kidney/bladder, stomach/intestine, and lung as the major uptake organs. Three-dimensional BLI identified liver/stomach and lung as the main internal luminescent sources, demonstrating the feasibility of detecting major uptake organs in live animals by 3D BLI with high-background signals in circulation. Notably, Gluc levels in capillary-depleted brain samples from Gluc-injected mice were comparable to controls, suggesting that Gluc may not cross the blood-brain barrier. Gluc uptake kinetics and intracellular half-life were assessed in various types of cell lines, implicating the involvement of non-specific pinocytosis. These results suggest that Gluc-based system may provide a useful tool for in vivo evaluation of protein/agent biodistribution following systemic delivery.
一种天然分泌的海肾荧光素酶（Gluc）已被用作生物发光成像（BLI）评估的报告基因。然而，Gluc 用于体内监测系统蛋白递送以及其天然生物分布的潜在应用尚未得到研究。为了研究 Gluc 的分泌和摄取情况，我们通过尾静脉注射的方式将 Gluc 编码质粒注入小鼠体内。全身 BLI 显示，爪垫的成像定量与血液 Gluc 活性直接相关。当通过使用肝启动子将基因表达限制在肝脏中时，体内 Gluc 生物分布分析显示肾脏/膀胱、胃/肠和肺是主要的摄取器官。三维 BLI 确定了肝脏/胃和肺是主要的内部发光源，表明通过 3D BLI 可以在有背景信号的情况下检测活体动物中的主要摄取器官。值得注意的是，来自注射 Gluc 的小鼠的毛细血管耗竭脑样本中的 Gluc 水平与对照相似，这表明 Gluc 可能不会穿过血脑屏障。在各种类型的细胞系中评估了 Gluc 的摄取动力学和细胞内半衰期，这表明非特异性胞吞作用的参与。这些结果表明，基于 Gluc 的系统可能为体内评估系统给药后蛋白质/药物的生物分布提供有用的工具。
