Li Qian, Hao Xuefang, Lv Juan, Ren Xiangkui, Zhang Kunyu, Ullah Ihsan, Feng Yakai, Shi Changcan, Zhang Wencheng
School of Chemical Engineering and Technology, Tianjin University, Yaguan Road 135, Tianjin 300350, China.
J Mater Chem B. 2017 Feb 28;5(8):1673-1687. doi: 10.1039/c6tb02212j. Epub 2017 Feb 9.
Gene delivery can enhance the endothelialization of biomaterial surfaces. However, the lack of efficient target function is still the major concern that hinders the clinical application of gene therapy. With the aim to develop a specific targeting gene carrier for endothelial cells (ECs), the Cys-Arg-Glu-Asp-Val-Trp (CREDVW) peptide was linked to the comb-like copolymer of poly(lactide-co-3(S)-methyl-morpholine-2,5-dione)-poly(poly(ethylene glycol) monomethacrylate) (PLMD-PPEGMA) to form the CREDVW modified copolymer PLMD-PPEGMA-CREDVW, which could enhance the special recognition of ECs. Mixed micelles were then prepared by co-assembling this comb-like copolymer and the amphiphilic grafting copolymer poly(lactide-co-3(S)-methyl-morpholine-2,5-dione)-g-polyethylenimine (PLMD-g-PEI). These mixed micelles with the CREDVW-functional peptide exhibited good pEGFP-ZNF580 (pDNA) binding ability and could condense it into complexes with proper size and positive zeta potential. The MTT results demonstrated the low cytotoxicity of the CREDVW-modified mixed micelle/pDNA complexes. The internalization efficiency of the CREDVW-modified complexes with targeting function was about two times higher than the dysfunctional CREVDW-modified complexes. Besides, the transfection efficiency of these complexes was more pronounced, compared to the control group, PEI(10 kDa)/pDNA, as detected by means of in vitro transfection studies. Western blot analysis demonstrated relatively high protein levels in the transfected cells by CREDVW-modified mixed micelle/pDNA complexes, up to 75%, in comparison to the control group (26%). In addition, the cell migration ability was significantly improved as demonstrated by the wound healing assay. These results indicated that the mixed micelles could act as an active targeting gene carrier, having both tunable gene transfection efficiency and low cytotoxicity, which are beneficial for the endothelialization of biomaterial surface.
基因递送可增强生物材料表面的内皮化。然而，缺乏有效的靶向功能仍是阻碍基因治疗临床应用的主要问题。为了开发一种针对内皮细胞（ECs）的特异性靶向基因载体，将半胱氨酸 - 精氨酸 - 谷氨酸 - 天冬氨酸 - 缬氨酸 - 色氨酸（CREDVW）肽与聚（丙交酯 - 共 - 3(S)-甲基 - 吗啉 - 2,5 - 二酮）- 聚（聚乙二醇单甲基丙烯酸酯）（PLMD - PPEGMA）的梳状共聚物相连，形成CREDVW修饰的共聚物PLMD - PPEGMA - CREDVW，其可增强对ECs的特异性识别。然后通过将这种梳状共聚物与两亲性接枝共聚物聚（丙交酯 - 共 - 3(S)-甲基 - 吗啉 - 2,5 - 二酮）- g - 聚乙烯亚胺（PLMD - g - PEI）共组装来制备混合胶束。这些带有CREDVW功能肽的混合胶束表现出良好的pEGFP - ZNF580（pDNA）结合能力，并能将其浓缩成具有合适尺寸和正ζ电位的复合物。MTT结果表明CREDVW修饰的混合胶束/pDNA复合物具有低细胞毒性。具有靶向功能的CREDVW修饰复合物的内化效率比无功能的CREVDW修饰复合物高约两倍。此外，通过体外转染研究检测，与对照组PEI(10 kDa)/pDNA相比，这些复合物的转染效率更显著。蛋白质免疫印迹分析表明CREDVW修饰的混合胶束/pDNA复合物转染的细胞中蛋白质水平相对较高，与对照组（26%）相比高达75%。此外，伤口愈合试验表明细胞迁移能力显著提高。这些结果表明，混合胶束可作为一种主动靶向基因载体，具有可调的基因转染效率和低细胞毒性，这有利于生物材料表面的内皮化。
