单纯疱疹病毒Ⅰ型扩增子载体转移脑啡肽基因在神经细胞的表达  

Experimental study on the expression of enkephalin gene transferred into neural cells by Herpes Simplex Virus type I amplicon vector

作　　者：王锷[1] 郭曲练[1] 白念岳[1] 王云姣[1] 曹晖[2] 吴小兵[2] 毕好生[3] 

机构地区：[1]中南大学湘雅医院麻醉科,长沙市410008 [2]中国预防医学科学院病毒基因工程国家重点实验室 [3]华中科技大学同济医学院同济医院麻醉科

出　　处：《中华麻醉学杂志》2002年第4期214-216,共3页Chinese Journal of Anesthesiology

基　　金：国家863基因治疗重大关键技术项目资助(863-Z20-05-02)

摘　　要：目的 研究采用单纯疱疹病毒Ⅰ型扩增子载体将脑啡肽基因导入神经细胞,观察载体lacZ基因和外源脑啡肽基因的表达。方法 实验共分4组:A组为对照组,未加病毒悬液的神经细胞;B组,加病毒悬液的神经细胞培养2d;C组,加病毒悬液的神经细胞培养3d;D组,加病毒悬液的神经细胞培养至第10d,留培基用放射免疫法检测外源脑啡肽基因的表达。用X-gal原位检测lacZ基因表达反映扩增子病毒的存在和滴度。结果 L-脑啡肽放射免疫检测证实转基因神经细胞L-脑啡肽含量较对照组明显升高（P<0.05）,X-gal染色证实加入假病毒悬液的神经细胞有30％以上的细胞蓝染。结论　HSV-I扩增子假病毒能将外源脑啡肽基因导入神经细胞,并使之有效表达。ve Neurotropic Herpes Simplex Virus type I(HSV-I) amplicon vector was adopted to transfer the enkephalin gene into neural cells. The purpose of the present study was to determine whether the express of LacZ gene in the vector and the transferred enkephalin gene could be effectively detected. Methods The mixed stock of the packaged and amplified recombinant plasmid was used to infect the cultured embryonic forebrain cortex cells. The study consisted of 4 groups: in group A in vitro cultured neural cells without the mixed virus stock served as control; in group B the neural cells were added to the mixed virus stock and cultured for 2 days; in group C and D the neural cells were added to the mixed virus stock as in group B but were cultured for 3 days and 10 days respectively. The medium was collected to detected HPPE expression by radioimmunoassay. The lacZ gene expression was detected by X-gal staining. Results L-enkephalin radioimmunoassay showed that the neural cell containing transferred gene had significantly higher L-enkephalin content as compared with those in control group(P<0.05) .The rate of cells expressing beta-galactosidase was up to 30% . Conclusions The amplicon virus we packaged and amplified has high infective ability and can transfer enkephalin gene into neural cells and have it expressed efficiently.

关 键 词：基因表达 疼痛 基因治疗 实验研究 单纯疱疹病毒I型扩增子载体 转移脑啡肽 基因 神经细胞 

分 类 号：R346[医药卫生—基础医学]