Anti-tumor effect of human endostatin mediated by retroviral gene transfer in nude mice  

逆转录病毒介导的人endostatin基因疗法抗肿瘤作用研究(英文)

作　　者：王轩[1] 刘福坤[1] 李希[2] 黎介寿 徐根兴 

机构地区：[1]Research Institute of General Surgery,Nanjing General Hospital,Clinical School of Medicine,Nanjing University,Nanjing 210002,China [2]Department of Life Sciences,Nanjing University,Nanjing 210002,China [3]Department of Molecular Biology,Nanjing Military Medical School,Nanjing 210002,China

出　　处：《Chinese Medical Journal》2002年第11期1664-1669,151,共6页中华医学杂志（英文版）

摘　　要：OBJECTIVE: To explore the inhibitory effect of human endostatin gene mediated by retroviral vector on the growth of human liver carcinoma. METHODS: A recombinant retroviral plasmid containing human endostatin gene and signal peptide was engineered and transferred into PA317 cells to produce retrovirus. Human liver carcinoma cells (SMMC7721) were infected with the above retrovirus to build a stable endostatin-transfected liver carcinoma cell line (SMMC-endo). The control liver carcinoma cell line (SMMC-pLncx) was developed in a similar way except that the plasmid was replaced by an empty retroviral vector. Immunohistochemistry and Western blot were used to test the expression and secretion of human endostatin. The biological activity of the expressed human endostatin was assessed by endothelial cell proliferation assay. The growth rates of SMMC-endo and control SMMC-pLncx cells in vivo and in vitro were also observed. RESULTS: The expression and secretion of human endostatin by endostatin-transfected SMMC-endo cells were confirmed by immunohistochemistry and Western blot. Compared with the control group, concentrated supernatant of SMMC-endo cells remarkably inhibited the proliferation of human umbilical vein endothelial cells by 48%, significantly higher than the inhibition by the control (10.2%; P目的　探讨逆转录病毒介导人血管内皮抑制素基因 (endostatin)对人肝癌细胞的生长抑制作用。方法　利用逆转录病毒载体pLncx构建含有人endostatin基因及信号肽的重组逆转录病毒质粒 ,并将其导入PA317细胞制备重组病毒。用重组病毒感染人肝癌细胞SMMC772 1获得稳定转人endostatin基因细胞株 (SMMC endo)。同样用空白病毒质粒制备对照细胞株SMMC pLncx。免疫组化及Westernblot检测人endostatin基因在转染细胞株中的表达和分泌。应用血管内皮细胞生长抑制试验验证转染细胞株表达的人endostatin的生物学活性。同时对转染细胞株在体内外的生长情况进行观察。结果　免疫组化及Westernblot印迹分析证实转染人endostatin基因的肝癌细胞能表达并分泌人endostatin。与转染空白对照组 (SMMC pLncx)相比 ,表达人endostatin的SMMC772 1浓缩上清可显著抑制人脐静脉血管内皮细胞生长 ,抑制率达 4 8% ,明显高于对照组 10 2 % (P <0 0 1)。体外生长实验显示 ,SMMC endo细胞与对照细胞SMMC pLncx体外生长速率无明显差别。而在转染细胞接种裸鼠 2 2d后 ,转染人endostatin基因的肿瘤细胞生长明显受到抑制 ,仅在部分裸鼠 (3/ 5只 )体内长出肿瘤 ,并且产生的腹侧肿瘤体积较对照组肿瘤缩小 94 5 % (P <0 0 0 1)。

关 键 词：Gene Therapy Animals Cell Division Cell Line Collagen ENDOSTATINS Endothelium  Vascular Gene Transfer Techniques MICE Mice  Nude Neoplasms  Experimental Peptide Fragments RETROVIRIDAE Transfection 

分 类 号：R73-36[医药卫生—肿瘤]