目的： 探讨以黏着斑激酶（focal adhesion kinase，FAK）特异性抑制剂TAE226处理或siRNA沉默FAK基因对人脐静脉血管内皮细胞（human umbilical vein endothelial cells，HUVEC）的增殖、迁移、细胞凋亡及毛细血管样结构形成的影响。方法： 应用Real-time PCR检测HUVEC和胸膜间皮瘤（malignant pleural mesothelioma，MPM）细胞株Y-MESO-14、NCI-H290中FAK mRNA的表达。以FAK siRNA转染HUVEC细胞致FAK基因沉默或以TAE226处理，采用Western blotting法检测经/未经VEGF预处理的HUVEC中FAK蛋白的表达；采用MTT法检测TAE226或FAK siRNA处理对HUVEC增殖能力的影响，Annexin-V FITC/PI双染色流式细胞术检测两者对HUVEC细胞凋亡的作用，Transwell法检测TAE226及siRNA处理对HUVEC细胞迁移的影响，体外脉管生成实验检测HUVEC中毛细血管样结构形成的数量。结果： 在HUVEC中FAK mRNA的表达量显著高于Y-MESO-14和NCI-H290细胞\[(0.032±0.006) vs (0.014±0.001)、(0.006±0.002)，均P<0.05\]。HUVEC在VEGF刺激下，FAK和pFAK的表达量均有增高（P<0.05)，FAK siRNA转染可以抑制VEGF预刺激下的FAK蛋白表达\[(0011±0002) vs (0.036±0.004)，P<0.01\]。分别给予不同浓度TAE226或FAK siRNA处理，均可呈浓度依赖性地抑制HUVEC增殖、迁移和凋亡(P<0.01)。体外脉管生成实验发现，VEGF可促进HUVEC中毛细血管样结构的生成，TAE226\[(14.32±783) vs (4631±39.46)条，P<0.01\]而FAK siRNA\[(11.83±6.75) vs (42.86±27.63)、(48.32±18.19)条，均P<0.01\] 转染可明显抑制毛细血管样结构形成。结论：TAE 226处理或FAK siRNA转染可以抑制HUVEC增殖、迁移和诱导细胞凋亡，同时可显著抑制毛细血管样结构的形成，提示FAK可能成为靶向治疗恶性肿瘤的潜在靶点。

Objective:To investigate roles for focal adhesion kinase (FAK) in the regulation of proliferation, migration, apoptosis, and angiogenesis of human vascular endothelial cells. Methods: Human umbilical vein endothelial cells (HUVECs) were transfected with FAK siRNA or treated with TAE226. FAK mRNA abundance in siRNA-transfected or TAE226-treated HUVECs and malignant pleural mesothelioma cell lines Y-MESO14 and NCI-H290 was determined by Real-time PCR. FAK protein was assessed by Western blotting, cell viability by MTT, apoptosis by flow cytometry, migration by Transwell assay, and capillary structure formation by matrigel-based tube formation assay in control, TAE-treated and FAK siRNA-transfected HUVECs. Results: FAK mRNA was significantly more abundant in HUVECs than in both Y-MESO-14 and NCI-H290 cell lines(P<0.05). rhVEGF treatment significantly increased FAK and pFAK protein levels of HUVECs (P<0.05). FAK siRNA effectively silenced FAK gene expression in the presence of rhVEGF in HUVECs (P<0.01). TAE226 at all concentrations used and FAK mRNA significantly inhibited proliferation, migration, and apoptosis of HUVECs (P<0.01). While rhVEGF significantly promoted angiogenesis, TAE226 and FAK siRNA significantly inhibited capillary structure formation in HUVECs (P<0.01). Conclusion: FAK siRNA plays important regulatory roles in vascular endothelial cell proliferation, migration, apoptosis, and capillary structure formation, and thus offer a novel target for angiogenesis-based cancer therapy.

国家自然科学基金资助项目（No.81360350）