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目的:探讨重组人血管内皮抑制素(恩度)对人肝癌细胞SMMC-7721VEGF165b表达的影响,及上调VEGF165b表达后,SMMC-7721对恩度敏感性的变化。方法:采用RT-PCR法和蛋白质印迹法检测恩度干预SMMC-7721后VEGF165b的表达变化;稳定转染VEGF165b真核表达质粒到SMMC-7721,RT-PCR和免疫细胞荧光共聚焦显微镜检测VEGF165b、HIF-1α和下游靶基因VEGFA表达的变化;MTT法检测恩度对人肝癌细胞生长抑制率的影响。结果:100和400μg/mL恩度能够诱导VEGF165b mRNA和蛋白表达上调,而外源性VEGF165b下调HIF-1α及下游靶基因VEGFA的表达。转染空质粒后,500和1000μg/mL恩度引起SMMC-7721的生长移植率分别为1.6%和0.4%,转染VEGF165b后,500μg/mL和1000μg/mL恩度引起的生长移植率分别为8.5%(P=0.0000)和20.2%(P=0.0000)。结论:恩度抗肿瘤新生血管的作用部分是通过上调VEGF165b介导的,VEGF165b增加SMMC-7721细胞对恩度的敏感性,两者治疗肿瘤有协同作用。
Objective: To investigate the effect of recombinant human endostatin (Ende) on the expression of VEGF165b in human hepatoma cell line SMMC-7721 and the effect of SMMC-7721 on the sensitivity to Endostar after up-regulating the expression of VEGF165b. Methods: VEGF165b expression was detected by RT-PCR and Western blotting. The VEGF165b expression was stably transfected into SMMC-7721 cells. The expression of VEGF165b was detected by RT-PCR and immunofluorescence confocal microscopy. HIF-1αand the downstream target gene VEGFA expression; MTT assay was used to determine the effect of entecavir on the growth inhibition rate of human hepatoma cells. RESULTS: Endostar at 100 and 400 μg / mL could upregulate the expression of VEGF165b mRNA and protein, whereas exogenous VEGF165b down-regulated the expression of HIF-1α and downstream target gene VEGFA. After transfection with empty plasmid, the growth engraftment rates of SMMC-7721 at the doses of 500 and 1000μg / mL were 1.6% and 0.4%, respectively. After transfection with VEGF165b, the rates of engraftment induced by 500μg / mL and 1000μg / mL of Endeavor were 8.5% (P = 0.0000) and 20.2% (P = 0.0000). Conclusion: Endomethral neovascularization is partly mediated by upregulation of VEGF165b, and VEGF165b increases the sensitivity of SMMC-7721 cells to End degree. The two have synergistic effects on the treatment of tumors.