Laboratory of Biomaterials, Department of Regeneration Science and Engineering, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan.
Tissue Eng Part A. 2019 Nov;25(21-22):1527-1537. doi: 10.1089/ten.TEA.2019.0017. Epub 2019 May 2.
The objective of this study is to prepare cationized gelatin nanospheres incorporating molecular beacon (MB) (cGNS) and a cationized gelatin-molecular beacon complex, and evaluate the time period of messenger RNA (mRNA) visualization. The cGNS with different degradabilities were prepared to incorporate MB. There was no difference in the apparent size and zeta potential between the cGNS and the complex, while the MB release from the complex was faster than that from the cGNS. When mouse mesenchymal stem cells were incubated with the complex and cGNS, the amount of MB internalized into cells and cytotoxicity of complex were higher compared with cGNS. However, in the amount range of noncytotoxicity, the amount of MB internalized was at a similar level among them. The intracellular fluorescence of cGNS was observed over 14 days, whereas that of complex disappeared within 5 days. Moreover, the time period of cGNS remaining in the cells prolonged with the increase of glutaraldehyde amount used in cGNS preparation. As the result, it is likely that the intracellular fluorescence was retained at a high level for longer time periods. It is concluded that the intracellular controlled release of MB is promising, to achieve long-term and continuous visualization of mRNA. Impact Statement Molecular beacon (MB) is a versatile activatable imaging probe to detect messenger RNA (mRNA). Cationized gelatin nanospheres incorporating MB (cGNS) with different degradabilities and a cationized gelatin-MB complex were prepared. The intracellular fluorescence of cGNS was observed over 14 days, whereas that of complex disappeared within 5 days. This is because the intracellular decrease of cGNS was slower than that of complex. It is concluded that the intracellular controlled release of MB is promising, to achieve long-term and continuous visualization of mRNA.
本研究旨在制备结合分子信标（MB）的阳离子化明胶纳米球（cGNS）和阳离子化明胶-分子信标复合物，并评估信使 RNA（mRNA）可视化的时间。制备了具有不同降解性的 cGNS 以结合 MB。cGNS 和复合物的表观粒径和zeta 电位没有差异，而复合物中 MB 的释放速度快于 cGNS。当间充质干细胞与复合物和 cGNS 孵育时，细胞内内化的 MB 量和复合物的细胞毒性均高于 cGNS。然而，在非细胞毒性的量范围内，它们内化的 MB 量处于相似水平。cGNS 的细胞内荧光可观察到 14 天，而复合物在 5 天内消失。此外，cGNS 在细胞内的保留时间随着用于制备 cGNS 的戊二醛量的增加而延长。因此，细胞内荧光很可能在较长时间内保持在较高水平。结论是，MB 的细胞内控制释放具有很大的潜力，可以实现 mRNA 的长期和连续可视化。
分子信标（MB）是一种多功能的可激活成像探针，可用于检测信使 RNA（mRNA）。制备了具有不同降解性的结合了 MB 的阳离子化明胶纳米球（cGNS）和阳离子化明胶-MB 复合物。cGNS 的细胞内荧光可观察到 14 天，而复合物在 5 天内消失。这是因为 cGNS 在细胞内的减少速度比复合物慢。结论是，MB 的细胞内控制释放具有很大的潜力，可以实现 mRNA 的长期和连续可视化。
