Downregulation of VEGF mRNA expression by triamcinolone acetonide acetate-loaded chitosan derivative nanoparticles in human retinal pigment epithelial cells.
Downregulation of VEGF mRNA expression by triamcinolone acetonide acetate-loaded chitosan derivative nanoparticles in human retinal pigment epithelial cells.
复制标题
醋酸曲安奈德负载壳聚糖衍生物纳米颗粒下调人视网膜色素上皮细胞中 VEGF mRNA 表达
DOI：
10.2147/ijn.s29690
复制
发表时间：
2012
影响因子：
8
通讯作者：
Lan Y
中科院分区：
文献类型：
--
作者：
Zhou H;Yang L;Li H;Gong H;Cheng L;Zheng H;Zhang LM;Lan Y
本研究旨在探讨醋酸曲安奈德（TAA）壳聚糖纳米粒对人视网膜色素上皮细胞血管内皮生长因子（VEGF）mRNA表达的下调作用。方法采用自组装法制备TAA/DA-Chit纳米粒，并通过透射电镜和zeta电位分析对其形貌和zeta电位进行表征。DA-Chit和TAA/DA-Chit纳米颗粒毒性使用细胞计数试剂盒-8测定法进行评价。使用异硫氰酸荧光素标记的DA-Chit纳米颗粒代替TAA/DA-Chit纳米颗粒测定细胞摄取的效率，通过倒置荧光显微镜和流式细胞术评估。TAA/DA-Chit纳米颗粒对VEGF mRNA表达的下调通过经处理的人视网膜色素上皮细胞的实时逆转录聚合酶链反应（RT-PCR）测定进一步研究。结果TAA/DA-Chit纳米粒的载药量为12%~ 82%，TAA的水溶性由0.3 mg/mL提高到2.1 mg/mL。这些纳米粒子在透射电子显微镜图像中显示出尺寸为100-550 nm的扁球形，并且具有正zeta电位。细胞计数试剂盒-8测定表明，DA-Chit和TAA/DA-Chit纳米颗粒对人视网膜色素上皮细胞分别没有毒性和低毒性。通过倒置荧光显微镜和流式细胞术证实人视网膜色素上皮细胞对异硫氰酸黄绿素标记的DA-Chit纳米颗粒的摄取。荧光定量RT-PCR检测显示，TAA/DA-Chit纳米粒作用于人视网膜色素上皮细胞后，VEGF mRNA水平下降。结论TAA/DA-Chit纳米粒对人视网膜色素上皮细胞VEGF mRNA表达具有下调作用，且细胞毒性较低，这可能是开发糖尿病视网膜病变治疗药物的有益特点。
Background The purpose of this study was to investigate the downregulation of mRNA expression of vascular endothelial growth factor (VEGF) by triamcinolone acetonide acetate (TAA)-loaded chitosan nanoparticles in human retinal pigment epithelial cells. Methods TAA-loaded deoxycholic acid-modified chitosan (TAA/DA-Chit) nanoparticles were prepared via a self-assembly mechanism, and their morphology and zeta potential were examined by transmission electron microscopy and zeta potential analysis, respectively. DA-Chit and TAA/DA-Chit nanoparticle toxicity was evaluated using a Cell Counting Kit-8 assay. The efficiency of cellular uptake was determined using fluorescein isothiocyanate-labeled DA-Chit nanoparticles, in place of TAA/DA-Chit nanoparticles, assessed by both inverted fluorescence microscopy and flow cytometry. Downregulation of VEGF mRNA expression by TAA/DA-Chit nanoparticles was further investigated by real-time reverse transcription polymerase chain reaction (RT-PCR) assay of the treated human retinal pigment epithelial cells. Results TAA/DA-Chit nanoparticles were prepared with a TAA-loading capacity in the range of 12%–82%, which increased the water solubility of TAA from 0.3 mg/mL to 2.1 mg/mL. These nanoparticles showed oblate shapes 100–550 nm in size in transmission electron microscopic images and had positive zeta potentials. The Cell Counting Kit-8 assay indicated that the DA-Chit and TAA/DA-Chit nanoparticles had no toxicity and low toxicity, respectively, to human retinal pigment epithelial cells. Fluorescein isothiocyanate-labeled DA-Chit nanoparticle uptake by human retinal pigment epithelial cells was confirmed by inverted fluorescence microscopy and flow cytometry. Real-time RT-PCR assay showed that the VEGF mRNA level decreased after incubation of human retinal pigment epithelial cells with TAA/DA-Chit nanoparticles. Conclusion TAA/DA-Chit nanoparticles had a downregulating effect on VEGF mRNA expression in human retinal pigment epithelial cells and low cytotoxicity, which might be beneficial characteristics for the development of future treatment for diabetic retinopathy.