缺氧微环境中Bcl-2干扰对食管癌Ec9706细胞血管生成拟态的影响

目的 探讨缺氧微环境对食管癌Ec9706细胞血管生成拟态的影响及其可能的分子机制。方法 应用CoCl2化学法诱导人食管癌细胞形成缺氧微环境，在缺氧微环境中以Bcl-2-siRNA干扰人食管鳞癌细胞系Ec9706，三维培养体系模拟人体内环境并置于荧光倒置显微镜下观察管道结构形成数量及形态。流式细胞术检测干扰前后细胞的增殖情况，采用RT-PCR法和Western blot法检测Bcl-2、VEcadherin(VE-cad)及MMP2的mRNA及蛋白表达情况。结果 缺氧对照组比常氧对照组形成更多的网络样管状结构（P＜0.000），凋亡减少（P＜0.05），Bcl-2、VE-cad和MMP2的mRNA和蛋白表达较常氧下显著增多（P＜0.05）。而在缺氧培养下进行Bcl-2-siRNA干扰后，缺氧实验组对比缺氧对照组网络样管状结构显著减少（P＜0.000），凋亡增多（P＜0.05）， 而Bcl-2、VE-cad及MMP2的mRNA和蛋白表达也较缺氧对照组细胞的表达显著降低（P＜0.05）。缺氧空载体组与缺氧对照组网络样管状结构未见明显差异（P＞0.05），凋亡率差异未见统计学意义（P＞0.05），Bcl-2、VE-cad和MMP2的mRNA和蛋白表达两组之间差异没有统计学意义（P＞0.05）。结论 缺氧能够诱导食管癌Ec9706细胞血管生成拟态的形成，且Bcl-2依赖的VE-cad过表达可能是缺氧诱导血管生成拟态形成的重要分子机制。

Effects of Bcl-2 Interference on Esophageal Cancer Cell Ec9706 Vasculogenic Mimicry in Hypoxic Microenvironment

Objective To investigate hypoxic microenvironment on esophageal cancer cells Ec9706vasculogenic mimicry and its possible molecular mechanism. Methods Human esophageal cancer cells wereinduced by CoCl2 chemical method to form hypoxia-microenvironment. While in hypoxic microenvironment,human esophageal squamous cell carcinoma cell lines Ec9706 was interfered by Bcl-2-siRNA, and humanbody environment was simulated by three-dimensional culture system and placed in the inverted fluorescencemicroscope to observe the number of pipeline structure formation and morphology. Flow cytometry wasapplied to detect cell proliferation. RT-PCR and Western blot were used to assay mRNA and proteinexpression of Bcl-2, VE-cad and MMP2. Results Hypoxia group formed more network-like tubular structure(P<0.000) than normoxic control group, decreased apoptosis (P<0.05), the mRNA and protein expression ofBcl-2, VE-cad and MMP2 compared with normoxic significantly increased (P<0.05) lower. After training inhypoxia carried out under Bcl-2-siRNA interference, the tubular network-like structure of experimental groupsignificantly reduced than hypoxic hypoxia control group (P<0.000), apoptosis increased (P<0.05), whereasthe mRNA and protein expression of Bcl-2, VE-cad and MMP2 was also significantly lower (P<0.05). Thenetwork-like tubular structures between hypoxia and anoxia empty vector control was no significant difference(P>0.05), apoptosis rate was no statistically significant (P>0.05), the mRNA and protein of Bcl-2, VE-cadand MMP2 was not statistically significant between groups(P>0.05).Conclusion Hypoxia could induce theformation of esophageal cancer cells Ec9706 vasculogenic mimicry, and the important molecular mechanismmay be related to Bcl-2-dependent VE-cad overexpression.