血管内皮生长因子反义寡核苷酸联合低分子肝素治疗小鼠膀胱癌

Effect of VEGF Antisense Oligonucleotide Combined with Low Molecular Weight Heparin on the Growth and Metastasis of Mice Bladder Cancer

目的探讨血管内皮生长因子(VEGF)反义寡核苷酸和低分子肝素法安明单用或联合对小鼠膀胱癌、转移的抑制作用。方法制备小鼠膀胱癌模型40只,随机分为对照组、VEGF反义寡核苷酸(ASODN)组、VEGF错义寡核苷酸(MSODN组)、低分子肝素(LMWH组)及联合治疗组,每组8只,分别给予生理盐水、VEGF-ASODN、VEGF-MSODN、法安明及VEGF-ASODN+法安明皮下注射,隔日1次,共15次。检测皮下移植瘤变化和盆腔淋巴结转移率;应用免疫组织化学的方法检测肿瘤组织微血管密度(MVD);应用Western印迹的方法检查肿瘤组织VEGF蛋白表达。结果MSODN、ASODN、LMWH和联合治疗组的抑瘤率分别为2.1%、33.6%、20.1%和45.3%;盆腔淋巴结转移率分别为62.5%、25%、25%和12.5%;Western印迹法检测显示,对照组和MSODN组表达较高水平的VEGF;ASODN组和联合治疗组MVD分别为(11.34±3.03)和(10.01±1.25),与对照组(21.23±4.01)和MSODN组(18.21±3.36)相比,差异有统计学意义(P<0.01);LMWH组MVD为(13.04±6.53),较对照组和MSODN组降低,差异有统计学意义(P<0.05)。结论VEGF-ASODN和法安明具有下调VEGF表达,抗血管生成作用;两者联合应用有协同抗肿瘤作用。

Objective To investigate the inhibitory effect of vascular endothelial growth factor（VEGF） antisense oligonucleotides or/and daheparin sodium（fragmin） on tumor growth and metastasis of mice bladder cancer. Methods Forty mice with bladder cancer were randomizedly divided into five groups：control group, VEGF antisense oligonucleotides （ASODN） group, VEGF mismatch sense oligonucleotides （MSODN） group,low molecular weight heparin （LMWH） group, and combined group. Sodium chloride, VEGF-ASODN, VEGF-MSODN, fragmin, and VEGF-ASODN plus fragmin were given respectively（ once every two days, 15 times altogether）. The volume and weight of subcutaneous tumors were measured, and the rate of pelvic lyrnphonode metastasis were detected by HE staining. The microvessel density（MVD） in tumor mass was measured by immunohistochemistry staining. VEGF protein level in tumor tissue was detected by Western blot assay. Results After treatment, the tumor growth inhibitory rates were 2. 1%, 33.6% ,211. 1% and 45.3% ,and the rate of pelvic lymphonode metastasis were 62. 5%, 25% ,25% and 12. 5 % in MSODN, ASODN, LMWH, and the combined group, respectively. Western Blot indicated that there was high level VEGF in MSOND and Control group. MVD of MSODN, ASODN, LMWH, the combined group and Control group was （ 18. 21 ± 3.36）, （ 11.34± 3. 113）, （ 13.04 ± 6.53 ）, （ 10.01 ± 1.25） and （21.23 ± 4. 01 ）. Difference was significant. Conclusion VEGF-ASODN and fragmin may down-regulate VEGF gene expression and inhibit angiogenesis. Combined use of fragmin can enhance anti-umor effect of VEGF-ASODN.