新型聚合物脂质体靶向血管内皮生长因子小干扰RNA转染抑制视网膜新生血管形成

[Targeting VEGF siRNA transfection by new polymeric liposomes to inhibit retinal neovascularization].

作者信息

Gao Yan, Liu Xinling, Li Chunhui, Peng Yao, Yang Jizhong, Wang Xiaowu, Li Xiaorong

机构信息

Department of Ophthalmology, Shanxi Eye Hospital, Taiyuan 030002, China.

Email:

出版信息

Zhonghua Yan Ke Za Zhi. 2015 May;51(5):344-50.

PMID:26311694

Abstract

OBJECTIVE

To formulate and evaluate polymeric liposomes (PL) nanoparticles as a novel non-viral gene delivery. To explore its applicability and feasibility as a non-viral vector for gene transportation.

METHOD

Experimental study. To construct the orongoxygen induced retinopathy (OIR) mouse (C57BL/6J) model on the basis of improved Smith's methods. Western blot was used to measure VEGF protein expression in retinal tissue at P17 and P22. HE staining and fluorescein-dextran angiography of retinal vascular were performed to observe the morphologic alterations of retinal neovascularization. Frozen-section was used to show the membrane translocation of PL.

RESULTS

In fluorescence angiograms, irregular neovascularization and fluoresce leakage were observed in OIR model. The results of Western blot showed that VEGF protein in retinal tissues were significantly different among groups (F = 158.207, P = 0.000) at P17 and P22 (F = 25.695, P = 0.000). The protein level was lower in both PL (0.70 ± 0.03) and Lipo group (0.66 ± 0.04) at P17 (P = 0.092), and the lower level was presented at P22 in PL group (0.50 ± 0.03) than in Lipo group (0.53 ± 0.05) (P < 0.05). HE staining were performed to observe the significantly improvements of retinal neovascularization in PL (28.0 ± 3.44) and Lipo group (24.50 ± 3.06) at P17. Moreover, inhibitory effects maintained at P22 in PL group (11.70 ± 3.09) as HE staining showed. Fluorescein angiography of retinal vascular showed retinal non-perfusion and neovascularization areas were smaller in both PL and Lipo group at P17. Frozen-section examination showed the property of membrane translocation. GFP expression could be seen in vitreous cavity just at first day post-intravitreal administration in PL and Lipo group, which could reach their peaks in external retina nearby RPE layer at P17, remaining at P22 in PL group.

CONCLUSION

The PL performed excellent ability of membrane translocation and it was a kind of slow steady released gene vector.

摘要

目的

制备并评估聚合物脂质体（PL）纳米粒作为一种新型非病毒基因递送载体。探索其作为基因运输非病毒载体的适用性和可行性。

方法

实验研究。在改良的史密斯方法基础上构建氧诱导视网膜病变（OIR）小鼠（C57BL/6J）模型。采用蛋白质免疫印迹法检测P17和P22时视网膜组织中血管内皮生长因子（VEGF）蛋白表达。进行视网膜血管苏木精-伊红（HE）染色和荧光素-葡聚糖血管造影，观察视网膜新生血管的形态学改变。采用冰冻切片显示PL的膜转位情况。

结果

在荧光血管造影中，OIR模型中观察到不规则新生血管形成和荧光素渗漏。蛋白质免疫印迹结果显示，P17和P22时视网膜组织中的VEGF蛋白在各组间有显著差异（P17时F = 158.207，P = 0.000；P22时F = 25.695，P = 0.000）。P17时PL组（0.70±0.03）和脂质体组（0.66±0.04）的蛋白水平均较低（P = 0.092），P22时PL组（0.50±0.03）的蛋白水平低于脂质体组（0.53±0.05）（P < 0.05）。HE染色显示，P17时PL组（28.0±3.44）和脂质体组（24.50±3.06）的视网膜新生血管有显著改善。此外，HE染色显示PL组在P22时仍保持抑制作用（11.70±3.09）。视网膜血管荧光素血管造影显示，P17时PL组和脂质体组的视网膜无灌注区和新生血管区均较小。冰冻切片检查显示了膜转位特性。在玻璃体腔内注射后第1天，PL组和脂质体组的玻璃体内均可看到绿色荧光蛋白（GFP）表达，在P17时可在视网膜色素上皮（RPE）层附近的外视网膜达到峰值，PL组在P22时仍有表达。