Tian Shouqin, Cao Duanwen, Zou Haijuan, Bai Feng, Wang Zhongjuan, Pan Shirong, Feng Min
School of Pharmaceutical Sciences, Department of Pharmacy, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou, People's Republic of China.
Department of Pharmaceutical Sciences, Nanfang Hospital, Southern Medical University, Guangzhou, People's Republic of China.
Int J Nanomedicine. 2015 Sep 10;10:5751-68. doi: 10.2147/IJN.S88109. eCollection 2015.
Upregulation of vascular endothelial growth factor (VEGF) expression can inhibit intimal thickening after vascular injury. However, the lack of efficient gene delivery systems leads to insufficient VEGF expression, which prevents its application in gene therapy. In the present study, to improve the delivery of the plasmid vector with the VEGF gene (pVEGF165) to the injured vessel wall, we explored the potentially important difference between endothelial cell-targeted and nontargeted polymeric carriers. The αvβ3 integrin is overexpressed on activated endothelial cells but not on normal quiescent vessels. In this study, CDG2-cRGD, synthesized by conjugating an αvβ3 integrin-binding cyclic arginylglycylaspartic acid (cRGD) peptide with the Generation 2 polycation polyamidoamine (PAMAMG2)-g-cyclodextrin (termed as CDG2), was developed as a targetable carrier. It was observed that the specific integrin-ligand interactions greatly enhanced cellular internalization of CDG2-cRGD in human umbilical vein endothelial cells (HUVECs), which are notoriously difficult to transfect. Consequently, HUVECs were found to show remarkably high levels of VEGF165 expression induced by the CDG2-cRGD polyplex. Interestingly, VEGF165 overexpression in vivo was more complex than that in vitro, and in vivo assays demonstrated that the stimulus response to balloon injury in arteries could obviously upregulate VEGF165 expression in the saline-treated group, although it was not enough to prevent intimal thickening. In gene-transfected groups, intravascular delivery of pVEGF165 with the CDG2-cRGD polyplex into rabbits after vascular injury resulted in a significant inhibition of intimal thickening at 4 weeks, whereas the low therapeutic efficacy in the nontargeted CDG2-treated group was only comparable to that in the saline-treated group. It is becoming clear that the conflicting results of VEGF165 gene therapy in two gene-transfected groups are reflective of the pivotal role of the cRGD-conjugated carriers in achieving the beneficial therapeutic effects of vascular gene therapy.
血管内皮生长因子（VEGF）表达上调可抑制血管损伤后的内膜增厚。然而，缺乏高效的基因递送系统导致VEGF表达不足，这阻碍了其在基因治疗中的应用。在本研究中，为了改善携带VEGF基因的质粒载体（pVEGF165）向受损血管壁的递送，我们探讨了内皮细胞靶向和非靶向聚合物载体之间潜在的重要差异。αvβ3整合素在活化的内皮细胞上过度表达，但在正常静止血管上不表达。在本研究中，通过将αvβ3整合素结合环精氨酸甘氨酰天冬氨酸（cRGD）肽与第2代聚阳离子聚酰胺胺（PAMAM G2）-g-环糊精（称为CDG2）偶联合成的CDG2-cRGD被开发为一种可靶向载体。据观察，特异性整合素-配体相互作用极大地增强了CDG2-cRGD在人脐静脉内皮细胞（HUVECs）中的细胞内化，而HUVECs notoriously difficult to transfect。因此，发现HUVECs显示出由CDG2-cRGD多聚体诱导的显著高水平的VEGF165表达。有趣的是，VEGF165在体内的过表达比在体外更复杂，体内试验表明，动脉球囊损伤的刺激反应在生理盐水处理组中可明显上调VEGF165表达，尽管这不足以防止内膜增厚。在基因转染组中，血管损伤后将携带pVEGF165的CDG2-cRGD多聚体经血管内递送至兔体内，4周时可显著抑制内膜增厚，而在非靶向CDG2处理组中低治疗效果仅与生理盐水处理组相当。越来越清楚的是，两个基因转染组中VEGF165基因治疗的矛盾结果反映了cRGD偶联载体在实现血管基因治疗有益治疗效果中的关键作用。
