重组人血管内皮生长因子165对K562白血病细胞生物学活性的影响

摘要

目的:探讨人重组血管内皮生长的因子165对人K562白血病细胞生物学活性的影响.方法:利用pcDNA3.1(+)质粒构建人重组含VEGF165的真核表达载体,脂质体转染至K562白血病细胞,并用G418筛选阳性克隆；以RT - PCR测定重组质粒载体VEGF165基因的表达；MTT法测定K562白血病细胞的生长；建立裸鼠皮下移植人K562肿瘤模型,测定肿瘤体积的大小；免疫组化检测肿瘤的微血管密度(MVD).结果:成功构建pcDNA3.1(+)- VEGF165表达质粒.K562白血病细胞转染重组质粒后,明显增加K562细胞中VEGF165 mRNA的表达,转染重组质粒的K562细胞增殖明显快于对照组；移植重组pcDNA3.1(+) -VEGF165质粒转染的K562细胞之后,荷瘤鼠移植瘤生长明显快于对照组；并且移植瘤组织微血管密度明显高于对照组(P＜0.05).结论:利用pc DNA3.1(+)真核表达质粒可成功构建含人VEGF165的重组质粒,重组质粒转染K562细胞后,体内外均明显促进K562细胞增殖.%Objective:To investigate the effect of recombinant human VEGF165 gene mediated by eukaryotic expression vector on biological activity of human K562 leukemia cells. Methods: Using eukaryotic expression vector pcDNA3.1( + ), recombinant humanVEGF165 gene clone was constructed, and huamanVEGF16S gene was trans-fected into K562 leukemic cells mediated by liposome and the positive clone was selected by G418. The recombinant VEGF165 gene was identified by RT - PCR, cell growth curve was detected by MTT assays. Subcutaneously transplanted human K562 carcinoma model was established in nude mice. Tumor size was detected regularly. Microvessel density in tumor was analyzed by imiminohistochemistry. Results: Plasmids pcDNA3. 1 ( + ) - VEGF165 was constructed successfully. The result showed that recombinant human VEGF165 was enhanced mRNA VEGF165 expression of K562 leukemic cells and the recombinant human VEGF165 could obviously promote the proliferation of K562 leukemic cells in virto. The volumes of transplantation tumors including recombinant human VEGF165 gene were significantly bigger than that of controls ( P < 0. 05 ). The number of microvessel density ( MVD) in experiment group was significantly more than that of control (P<0.05). Conclusion: VEGF165 cDNA gene was successfully cloned in K562 cells. And recombinant plasmid pcDNA3.1 ( + ) - VEGF165 gene can enhance proliferation of K562 leukemic cells in vitro and vivo.