Wu Han-Ping, Feng Gan-Sheng, Liang Hui-Min, Zheng Chuan-Sheng, Li Xin
Department of Interventional Radiology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China.
World J Gastroenterol. 2004 Mar 15;10(6):813-8. doi: 10.3748/wjg.v10.i6.813.
Transcatheter arterial embolization (TAE) of the hepatic artery has been accepted as an effective treatment for unresectable hepatocellular carcinoma (HCC). However, embolized vessel recanalization and collateral circulation formation are the main factors of HCC growth and recurrence and metastasis after TAE. Vascular endothelial growth factor (VEGF) plays an important role in tumor angiogenesis. This study was to explore the inhibitory effect of VEGF antisense oligodeoxynucleotides (ODNs) on VEGF expression in cultured Walker-256 cells and to observe the anti-tumor effect of intra-arterial infusion of antisense ODNs mixed with lipiodol on rat liver cancer.
VEGF antisense ODNs and sense ODNs were added to the media of non-serum cultured Walker-256 cells. Forty-eight hours later, VEGF concentrations of supernatants were detected by ELISA. Endothelial cell line ECV-304 cells were cultured in the supernatants. Seventy-two hours later, growth of ECV-304 cells was analyzed by MTT method. Thirty Walker-256 cell implanted rat liver tumor models were divided into 3 groups. 0.2 mL lipiodol (LP group, n=10), 3OD antisense ODNs mixed with 0.2 mL lipiodol (LP+ODNs group, n=10) and 0.2 mL normal saline (control group, n=10) were infused into the hepatic artery. Volumes of tumors were measured by MRI before and 7 d after the treatment. VEGF mRNA in cancerous and peri-cancerous tissues was detected by RT-PCR. Microvessel density (MVD) and VEGF expression were observed by immunohistochemistry.
Antisense ODNs inhibited Walker-256 cells' VEGF expression. The tumor growth rate was significantly lower in LP+ODNs group than that in LP and control groups (140.1+/-33.8%, 177.9+/-64.9% and 403.9+/-69.4% respectively, F=60.019, P<0.01). VEGF mRNAs in cancerous and peri-cancerous tissues were expressed highest in LP group and lowest in LP+ODNs group. The VEGF positive rates showed no significant difference among LP, control and LP+ODNs groups (90%, 70% and 50%, H=3.731, P>0.05). The MVD in LP+ODNs group (53.1+/-18.4) was significantly less than that in control group (73.2+/-20.4) and LP group (80.3+/-18.5) (F=5.44, P<0.05).
VEGF antisense ODNs can inhibit VEGF expression of Walker-256 cells. It may be an antiangiogenesis therapy agent for malignant tumors. VEGF antisense ODNs mixed with lipiodol embolizing liver cancer is better in inhibiting liver cancer growth, VEGF expression and microvessel density than lipiodol alone.
肝动脉导管栓塞术（TAE）已被公认为是不可切除肝细胞癌（HCC）的一种有效治疗方法。然而，栓塞血管再通及侧支循环形成是TAE术后HCC生长、复发和转移的主要因素。血管内皮生长因子（VEGF）在肿瘤血管生成中起重要作用。本研究旨在探讨VEGF反义寡脱氧核苷酸（ODNs）对培养的Walker-256细胞中VEGF表达的抑制作用，并观察动脉内注入与碘油混合的反义ODNs对大鼠肝癌的抗肿瘤作用。
将VEGF反义ODNs和正义ODNs加入无血清培养的Walker-256细胞培养基中。48小时后，用ELISA法检测上清液中VEGF浓度。将内皮细胞系ECV-304细胞培养于上清液中。72小时后，用MTT法分析ECV-304细胞的生长情况。将30只Walker-256细胞植入大鼠肝肿瘤模型分为3组。将0.2 mL碘油（LP组，n = 10）、3OD反义ODNs与0.2 mL碘油混合（LP + ODNs组，n = 10）和0.2 mL生理盐水（对照组，n = 10）注入肝动脉。治疗前及治疗后7天用MRI测量肿瘤体积。用RT-PCR法检测癌组织和癌周组织中的VEGF mRNA。用免疫组织化学法观察微血管密度（MVD）和VEGF表达。
反义ODNs抑制Walker-256细胞的VEGF表达。LP + ODNs组肿瘤生长率显著低于LP组和对照组（分别为140.1±33.8%、177.9±64.9%和403.9±69.4%，F = 60.019，P < 0.01）。癌组织和癌周组织中的VEGF mRNAs在LP组表达最高，在LP + ODNs组表达最低。LP组、对照组和LP + ODNs组的VEGF阳性率无显著差异（90%、70%和50%，H = 3.731，P > 0.05）。LP + ODNs组的MVD（53.1±18.4）显著低于对照组（73.2±20.4）和LP组（80.3±18.5）（F = 5.44，P < 0.05）。
VEGF反义ODNs可抑制Walker-256细胞的VEGF表达。它可能是一种恶性肿瘤抗血管生成治疗药物。与单纯碘油相比，VEGF反义ODNs与碘油混合栓塞肝癌在抑制肝癌生长、VEGF表达和微血管密度方面效果更好。
