逆转录病毒介导人血管内皮抑制素基因对人肝癌细胞体内生长的抑制作用

目的：探讨宿主细胞表达的人血管内皮抑制素（endostatin)对人肝癌细胞在体内生长的影响。方法：利用逆转录病毒载体pLncx,将endostatin基因导入人肝癌细胞SMMC7721内建立转基因肝癌细胞株。应用PCR、免疫组化及Western blot检测人endostatin的转染、表达和分泌。血管内皮细胞增殖试验检测表达的endostatin生物学活性。同时对转染细胞株在体外及在裸鼠体内的生长情况进行观察。结果：PCR证实,转endostatin基因的肝癌细胞 基因组中存在有550bp人特异性endostatin片段,免疫组化及Western blot印迹分析示人endostatin在转染肝癌细胞株中获得稳定表达和分泌。转染细胞表达的endostatin能显著抑制人血管内皮生长,抑制率达48％（P＜0．01）。与对照组相比,转染人endostatin基因的肝癌细胞在体外的生长速度无明显改变,但在接种裸鼠皮下后,肿瘤生长明显受到抑制,在22d时,肿瘤缩小率达94．5％（P＜0．01）。结论：逆转录病毒介导的人endostatin基因疗法对人肝癌SMMC7721在裸鼠体内的生长有显著抑制作用。

Inhibitory effect of endostatin mediated by retroviral gene transfer on human liver carcinoma SMMC7721 in vivo

Objective To explore the effect of human endostatin expressed by host cells on the growth of human liver carcinoma in vivo Methods Human endostain gene was transferred into SMMC7721 cells by retroviral pLncx to build endostatin transfected cell line PCR, immunohistochemistry and Western blot analysis were applied to examine the transfection, expression and secretion of endostatin Endothelial cell proliferation assay was used to determine the biological activity of expressed endostatin The in vivo and in vitro growth rates of the endostatin transfected and control SMMC7721 cells were also observed Results PCR proved that the genome of endostatin transfected SMMC7721 cells contained a 550 bp specific fragment of endostatin The expression and secretion of human endostatin from endostatin transfected SMMC7721 cells were confirmed by immunohistochemistry and Western blot analysis Endostatin expressed by host cells could inhibit the proliferation of human umbilical vein endothelial cells by 48%( P <0 01) In vitro proliferation assay showed that endostatin transfected SMMC7721 cells had no change in proliferation rate compared to control SMMC7721 cells. In comparison with control group, however, tumor growth rate in vivo from endostatin transfected SMMC7721 cells was inhibited greatly by 94 5%, 22 days after inoculation into nude mice( P <0 01) Conclusion Human endostatin mediated by retroviral gene transfer can inhibit greatly the growth of human liver carcinoma SMMC7721 in vivo