Department of Biotechnology, The Catholic University of Korea, Wonmi-gu, Gyeonggi-do 420-743, South Korea.
Biomaterials. 2012 Sep;33(27):6485-94. doi: 10.1016/j.biomaterials.2012.05.040. Epub 2012 Jun 9.
To improve the transfection efficiency of non-viral gene vectors to human mesenchymal stem cells (hMSCs), a photosensitizer (PS)-induced gene delivery system was designed by using pheophorbide-a (pheo-a) as a PS. In FACS results, this system showed excellent gene transfection efficiency depending on irradiation power. The result was strongly supported by western blot and real-time quantitative PCR (RT-qPCR) assays. The protein and mRNA expression of enhanced green fluorescent protein (EGFP) in hMSCs treated with 0.9 J/cm(2) irradiation increased 9.8- and 8.7-fold compared with non-irradiated hMSCs, respectively. Furthermore, the internalization of PEI/pDNA complexes in hMSCs was enhanced by light irradiation even under conditions that inhibited endocytosis. The hemolytic activity of PS with irradiation (0.9 J/cm(2)) significantly increased to 55%. Thus, PS with light irradiation facilitated both the internalization and endosomal escape of gene complexes. For osteogenic induction, the Runt-related transcription factor 2 (Runx2) gene was transferred to hMSCs via PS-induced transfection. Von Kossa staining indicated that Runx2 overexpression significantly enhanced the osteogenesis of hMSCs. Therefore, this PS-induced gene delivery method has potential value for stem cell therapy via gene delivery.
为提高非病毒基因载体对人骨髓间充质干细胞（hMSCs）的转染效率，设计了一种以原卟啉 IX（pheo-a）为光敏剂（PS）的基因传递系统。在流式细胞术结果中，该系统显示出优异的基因转染效率，这与辐照功率有关。Western blot 和实时定量 PCR（RT-qPCR）检测结果强烈支持了这一结果。与未辐照的 hMSCs 相比，用 0.9 J/cm(2)辐照处理的 hMSCs 中增强型绿色荧光蛋白（EGFP）的蛋白和 mRNA 表达分别增加了 9.8 倍和 8.7 倍。此外，即使在抑制细胞内吞作用的条件下，光照也能增强 hMSCs 中 PEI/pDNA 复合物的内化。辐照（0.9 J/cm(2))后的 PS 的溶血活性显著增加至 55%。因此，光照后的 PS 促进了基因复合物的内化和内涵体逃逸。对于成骨诱导，通过 PS 诱导的转染将 runt 相关转录因子 2（Runx2）基因转导至 hMSCs。Von Kossa 染色表明，Runx2 过表达显著增强了 hMSCs 的成骨作用。因此，这种 PS 诱导的基因传递方法在通过基因传递进行干细胞治疗方面具有潜在价值。
