上海交通大学学报（医学版）

›› 2012, Vol. 32 ›› Issue (10): 1288-.doi: 10.3969/j.issn.1674-8115.2012.10.002

• 论著（基础研究） • 上一篇    下一篇

超声辐照与丁酸钠对腺相关病毒转染大鼠视网膜色素上皮细胞的影响

李红丽1, 杜联芳2, 李凤华1, 王慧萍3   

  1. 1.上海交通大学 医学院附属仁济医院超声医学科, 上海 200127； 2.上海交通大学附属第一人民医院超声科, 上海 200080； 3.上海交通大学附属第一人民医院中心实验室, 上海 200080
  • 出版日期:2012-10-28 发布日期:2012-11-05
  • 通讯作者: 杜联芳, 电子信箱: Du_lf@163.com。
  • 作者简介:李红丽（1975—）, 女, 博士, 主治医师；电子信箱: lihliwl@163.com。
  • 基金资助:

    国家自然科学基金（30772369）；上海市自然科学基金（11ZR1421100）；上海交通大学医学院附属仁济医院科研培育基金（RJPY10-003）

Effects of ultrasound exposure and sodium butyrate on transfection of adeno-associated virus to rat retinal pigment epithelial cells

LI Hong-li1, DU Lian-fang2, LI Feng-hua1, WANG Hui-ping3   

  1. 1.Department of Ultrasound, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200127, China；2.Department of Ultrasound, 3. Department of Experimental Research Center, the First People´s Hospital, Shanghai Jiaotong University, Shanghai 200080, China
  • Online:2012-10-28 Published:2012-11-05
  • Supported by:

    National Natural Science Foundation of China,30772369；Natural Science Foundation of Shanghai, 11ZR1421100；Foundation of Renji Hospital, Shanghai Jiaotong University School of Medicine, RJPY10-003

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摘要：

目的 探讨超声辐照与丁酸钠在携带增强型绿色荧光蛋白的2型重组腺相关病毒(rAAV2-EGFP)感染大鼠视网膜色素上皮（RPE）细胞中的应用价值。方法 大鼠RPE细胞接种于24孔板中，分别以rAAV2EGFP（对照组）、rAAV2-EGFP+超声辐照（超声辐照组，辐照时间60 s，声强0.5、1 W/cm2）、rAAV2-EGFP+不同终浓度丁酸钠（丁酸钠组）转染大鼠RPE细胞。转染后48 h，倒置荧光显微镜观察转染细胞EGFP表达情况，流式细胞仪检测细胞转染效率（EGFP阳性细胞百分率）；转染后24 h，MTS法检测细胞增殖。结果 流式细胞仪检测结果显示：超声辐照组和丁酸钠组EGFP阳性细胞百分率显著高于对照组（P<0.05），丁酸钠组转染效率提高的百分率显著高于超声辐照组（P<0.05）。细胞增殖检测结果显示，超声辐照组转染细胞的吸光度值显著高于丁酸钠组（P<0.05）。结论 丁酸钠和超声辐照均能提高rAAV2-EGFP 对大鼠RPE 细胞的转染效率，其中丁酸钠的效果显著，而超声辐照对细胞增殖的抑制程度较轻。

关键词: 超声辐射, 丁酸钠, 腺相关病毒, 视网膜色素上皮细胞

Abstract:

Objective To investigate the role of ultrasound exposure and sodium butyrate in transfection of type 2 recombinant adeno-associated virus carrying enhanced green fluorescent protein (rAAV2-EGFP) to rat retinal pigment epithelial (RPE) cells. Methods Rat RPE cells were incubated in 24-well plates, and were transfected with rAAV2-EGFP (control group), rAAV2-EGFP+ultrasound exposure (ultrasound exposure group, ultrasound exposure for 60 s with ultrasound intensity of 0.5 and 1 W/cm2) and rAAV2-EGFP+different concentrations of sodium butyrate (sodium butyrate group) respectively. Forty-eight hours after transfection, the expression of EGFP in the transfected cells was observed by inverted microscopy, and the transfection efficiency (percentage of EGFP positive cells) was detected by flow cytometry. Twenty-four hours after transfection, the cell proliferation was determined by MTS method. Results Flow cytometry indicated that the percentages of EGFP positive cells in ultrasound exposure group and sodium butyrate group were significantly higher than that in control group (P<0.05), and the increase in transfection efficiency in sodium butyrate group was significantly higher than that in ultrasound exposure group (P<0.05). Cell proliferation detection revealed that the absorbance of the transfected cells in ultrasound exposure group was significantly higher than that in sodium butyrate group (P<0.05). Conclusion Both sodium butyrate and ultrasound exposure can increase the transfection efficiency of rAAV2-EGFP to rat RPE cells, and the effect of sodium butyrate is more significant, while ultrasound exposure has less inhibition effect on cell proliferation.

Key words: ultrasound, sodium butyrate, adeno-associated virus, retinal pigment epithelial cells