选择性iNOS抑制剂1400W对缺氧培养人肝癌SMMC-7721细胞株iNOS和VEGF表达的影响

Effects of Selective iNOS Inhibitor 1400W in Hypoxic on the Expression of iNOS and VEGF in Hepatocarcinoma Cell Line SMMC-7721

目的探讨选择性诱导型一氧化氮合酶(inducible nitric oxide synthase,iNOS)抑制剂1400W对缺氧条件培养的人肝癌SMMC-7721细胞株iNOS和血管内皮生长因子(vascular endothelial growth factor,VEGF)表达的影响。方法实验组分别加入选择性iNOS抑制剂1400W 50(50μmol/L组)、100(100μmol/L组)、200(200μmol/L组)μmol/L的培养液培养;对照组不加选择性iNOS抑制剂1400W培养液培养,缺氧条件下培养24 h后,采用硝酸还原酶法检测细胞培养上清液NO含量;RT-PCR检测iNOSmRNA和VEGFmRNA水平。结果50μmol/L组、100μmol/L组、200μmol/L组NO含量分别为(28.77±2.1)、(21.63±3.0)、(17.23±2.4)μmol/L,均明显低于对照组[(43.68±4.3)μmol/L](P均<0.01);50μmol/L组、100μmol/L组、200μmol/L iNOSmRNA水平分别为0.212±0.009、0.169±0.012、0.136±0.006,均明显低于对照组0.246±0.009(P<0.05或P<0.01);50μmol/L组、100μmol/L组、200μmol/L VEGFmRNA水平分别为0.429±0.038、0.309±0.022、0.249±0.014,均明显低于对照组0.598±0.040(P<0.05或P<0.01)。结论选择性iNOS抑制剂1400W可抑制缺氧条件下培养的人肝癌SMMC-7721细胞株iNOS和VEGF表达。提示选择性iNOS抑制剂1400W对肝癌新生血管的形成具有预防和潜在的治疗价值。

Objective To explore the effects of selective iNOS inhibitor 1400W in hypoxic on the expressions of iNOS and VEGF in Hepatocarcinoma cell line SMMC-7721. Methods The experimental groups were cultured in medium with different concentrations of 1400W （50,100, 200 μmol/L）,control group without 1400W. All groups were cultured in hypoxia for 24 h. The levels of NO in the supernatants were determined by nitrate reducase method and the expressions of iNOS and VEGF mRNAs were examined by Real-time polymerase chain reaction. Results The NO levels of 50 μmol/L group, 100 μmol/L group, and 200 μmol/L group were （28.77 ± 2.1）, （21. 63 ± 3. 0）, （17. 23 ± 2. 4）μmol/L, respectively. Compared to control group [（43. 68 ± 4.3）μmol/L] ,the levels of NO was decreased（all P〈0.01） ;the expressions of iNOS mRNA in a- bove treatment groups were 0. 212±0. 009,0. 169±0. 012,0. 136±0. 006,respectively,and compared to control group （0. 246± 0. 009）, the expressions of iNOSmRNA were down-regulated significantly （P〈0.05 or P〈0.01） ;the expressions of VEGF mRNAs in the treatment groups were 0. 429±0. 038, 0. 309 ± 0. 022, 0. 249 ±0. 014, respectively, and compared to control group （0. 598±0. 040） ,the expressions of VEGF mRNA were down-regulated significantly （P〈0.05 or P〈0.01）; Conclusion Selective iNOS inhibitor in hypoxic could inhibit the expressions of iNOS and VEGF in Hepatocarcinoma cell line SMMC-7721. It suggests that 1400W might play a latent role in the prevention and treatment of hepatocarcinoma neovascularization.