超声微泡介导bcl-xl基因抗视网膜神经节细胞凋亡作用的实验研究  

Anti-apoptosis effect of bcl-xl on retinal ganglion cells by ultrasound-mediated microbubble destruction

作　　者：李伟[1] 刘苏[1] 王志刚[2] 

机构地区：[1]重庆医科大学附属第二医院眼科,重庆400010 [2]重庆医科大学超声影像学研究所,重庆400010

出　　处：《中国医学影像技术》2006年第8期1147-1150,共4页Chinese Journal of Medical Imaging Technology

基　　金：国家自然科学基金重点项目资助(30430230)

摘　　要：目的评价超声微泡介导bclxl基因抗视网膜神经节细胞(RGCs)凋亡的作用。方法体外混合培养LongEvans大鼠RGCs,并建立N甲基D天冬氨酸(NMDA)损伤的RGCs凋亡模型。将体外培养的RGCs分为A组(正常对照组),B组(NMDA组),C组(bclxl转染+NMDA组);其中C组在加入NMDA前48h用超声微泡介导bclxl转染RGCs。转染48h后采用免疫组化分析转染细胞和未转染细胞的bclxl蛋白水平;加入NMDA36h后采用吖啶橙/溴化乙锭(AO/EB)双荧光染色检测细胞凋亡的形态特征,琼脂糖电泳检测细胞凋亡DNA片断。结果免疫组化检测表明转染细胞与未转染细胞的bclxl蛋白表达水平有差异,AO/EB检测发现B组可见大量凋亡小体,C组可见少许凋亡细胞。琼脂糖电泳检测亦发现B组呈典型的DNA“梯度”条带,而A组和C组无明显的DNA“梯度”条带。结论超声微泡介导bc1xl基因抗视网膜神经节细胞凋亡有一定作用,有可能为视网膜视神经疾病的基因治疗提供一种新的方法。Objective To assess the effect of bcl-x1 gene on N-methyl-D-aspartate （NMDA）-induced apoptosis on cultured retinal ganglion cells （RGCs） by ultrasound-mediated microbubble destruction. Methods RGCs were cultured in 24-well plates. NMDA-induced apoptosis in cultured RGCs was established. The experiment was divided into three groups： control, NMDA treatment and bcl-x1 transfection＋NMDA treatment group （bcl-x1 gene was transfected into RGCs 48 h by ultrasound-mediated microbubble destruction before NMDA-induced apoptosis was established）. The expression of bcl-x1 protein in transfected and non-transfected RGCs was assessed by immunohistochemistry assay. The morphotic character of RGCs was revealed by acridine orange and ethidium bromide staining. DNA fragment was detected by agarose gel electrophoresis. Results The expression of bcl-x1 protein in transfected and non-transfected RGCs was different. Lots of apoptotic bodies was found in NMDA treatment group, less were found in the other two groups. Representative DNA fragment was detected in the NMDA treatment group. Conclusion Transfection of bcl-x1 has anti-apoptosis effect on cultured RGCs from apoptosis induced by NMDA with ultrasound-mediated microbubble destruction. It may be a promising gene therapy for retinal and optic nerve diseases.

关 键 词：超声造影剂 视网膜神经节细胞 凋亡 BCL-X1 基因疗法 

分 类 号：R445.1[医药卫生—影像医学与核医学] Q95-33[医药卫生—诊断学]