VEGF165反义RNA对人食管鳞癌细胞影响的实验研究

Experimental study on the effect of VEGF165 antisense RNA on human squamous cell carcinoma of esophagus

目的 :观察血管内皮生长因子16 5 (VEGF16 5 )反义RNA对人食管鳞癌细胞EC10 9的影响 ,探讨其治疗食管癌的可行性。方法 :采用亚克隆技术 ,构建并鉴定VEGF16 5 反义RNA的真核表达载体。以重组质粒转染人食管鳞癌细胞EC10 9后 ,将其接种于裸鼠皮下 ,分别利用原位杂交、激光共聚焦、图象分析及微血管计数等方法 ,观察转染前后EC10 9细胞的生物学性状和致瘤性。结果 :成功地构建了VEGF16 5 反义RNA的真核表达载体 ,并在EC10 9细胞中获得表达。转染细胞中VEGF16 5 的表达下降 75% ,其生物学性状不受外源基因表达的影响 ,但其在裸鼠皮下的致瘤性和和瘤组织中血管的生成明显下降。VEGF16 5 反义RNA转染组、空载体转染组和对照组中肿瘤的体积 ,分别为 (82 0± 112 .5)mm3 、(793 0± 10 3 5)mm3 和 (7850± 950 )mm3(P <0 .0 1) ;微血管的密度分别为 (8.5± 1.2 ) /mm2 、(44.3± 9.4) /mm2 和 (46.4± 12 .6) /mm2 (P <0 .0 1)。结论 :VEGF16 5反义RNA能够明显减少食管鳞癌细胞内VEGF16 5 的表达 ,具有抑制肿瘤生长和血管生成的作用 ,可望用于实体肿瘤的辅助治疗。

AIM: To investigate the effect of antisense RNA against vascular endothe lial growth factor 165 (VEGF 165 ) on human esophagus squamous cell carcinom a cell line EC109. METHODS: Eukaryotic expression vector for VEG F165 antisense RNA was constructed and identified. Recombinant plasmid was trans fected into EC109 cells and the transfected EC109 cells were inoculated subcuta neously to nude mice. The biological characteristics and tumorigenicity of trans fected EC109 cells were observed by in situ hybridization, laser confocal m icroscope, transmission electron microscopy and flow cytometry. RESULTS: The eukaryotic expression vector pCEP-AVEGF 165 was successfull y constructed and expressed in transfected EC109 cells. The rate of VEGF 165 expression dropped by 75% in transfected cells. The morphology and cell cycle of transfected EC109 cells were not affected by the antisense RNA, but the tum origenicity and angiogenesis of transfected EC109 cells were greatly reduced in nude mice. The volume of tumors in pCEP-AVEGF 165 -transfected group, empty vector-transfected gro up and control group were (820±112.5) mm 3, (7 930±1 035) mm 3 and (7 850±9 50) mm 3, respectively. The microvessel density of the three groups were (8.5 ±1.2)/mm 2, (44.3±9.4)/mm 2 and (46.4±12.6)/mm 2, respectively. C ONCLUSION: The angiogenesis and tumorigenicity of human esophageal squam ous cell carcinoma were effectively inhibited by VEGF 165 antisense RNA, w hich may be applied to treat solid tumor in the future.