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目的研究Runt相关转录因子3(RUNX3)对人骨肉瘤细胞株U2OS促血管形成能力的影响。方法 U2OS细胞转染RUNX3过表达质粒,免疫印迹(Western blot)法检测细胞内血管内皮生长因子(VEGF)、黏着斑激酶(FAK)蛋白表达变化,酶联免疫吸附(ELISA)法测定VEGF水平的变化。细胞计数(CCK-8)法检测RUNX3过表达对人脐静脉内皮细胞(HUVECs)增殖的影响,微血管形成实验测定RUNX3过表达对HUVECs微血管形成的影响。结果转染RUNX3质粒后,实验组U2OS细胞中RUNX3基因为2.20±0.22,RUNX蛋白0.25±0.04,对照组RUNX3基因为1.40±0.11,RUNX3蛋白为0.15±0.03,差异有统计学意义(P<0.05)。RUNX3过表达后,U2OS细胞中FAK和VEGF蛋白分别为0.15±0.02和0.22±0.03,对照组分别为0.37±0.05,和0.42±0.05,差异有统计学意义(P<0.05);实验组VEGF水平为(2381.04±166.07)pg·m L~(-1),对照组为(3177.74±322.76)pg·m L~(-1),差异有统计学意义(P<0.05)。RUNX3过表达后,U2OS细胞的促HUVECs血管形成个数为(5.00±1.00)个,对照组为(12.00±2.00)个;实验组U2OS促HUVECs增殖活性为0.66±0.02,对照组为1.05±0.09,差异有统计学意义(P<0.05)。结论 RUNX3可抑制骨肉瘤细胞的促血管形成能力,其机制可能是通过减少细胞中FAK的表达、抑制细胞对VEGF的分泌来实现的。
Objective To investigate the effect of Runt-related transcription factor 3 (RUNX3) on the angiogenesis of human osteosarcoma cell line U2OS. Methods RUNX3 overexpression plasmid was transfected into U2OS cells. The expression of vascular endothelial growth factor (VEGF) and focal adhesion kinase (FAK) protein was detected by Western blot. The level of VEGF was measured by enzyme - linked immunosorbent assay (ELISA) Variety. The effect of RUNX3 overexpression on the proliferation of human umbilical vein endothelial cells (HUVECs) was examined by CCK-8 assay. The effect of RUNX3 overexpression on the microvascular formation of HUVECs was examined by microvascular formation assay. Results The RUNX3 gene was 2.20 ± 0.22 in U2OS cells transfected with RUNX3 and 0.25 ± 0.04 in RUNX3. The RUNX3 gene in control group was 1.40 ± 0.11 and the RUNX3 protein was 0.15 ± 0.03 (P <0.05) ). After overexpression of RUNX3, the protein levels of FAK and VEGF in U2OS cells were 0.15 ± 0.02 and 0.22 ± 0.03 respectively, while those in control group were 0.37 ± 0.05 and 0.42 ± 0.05, respectively (P <0.05). The levels of VEGF, (2381.04 ± 166.07) pg · m L -1 in the control group, and (3177.74 ± 322.76) pg · m L -1 in the control group. The difference was statistically significant (P <0.05). After overexpression of RUNX3, the number of UUVIN-induced HUVECs was (5.00 ± 1.00) in the control group and (12.00 ± 2.00) in the control group, respectively. The proliferative activity of U2OS-induced HUVECs was 0.66 ± 0.02 in the experimental group and 1.05 ± 0.09 in the control group , The difference was statistically significant (P <0.05). Conclusion RUNX3 can inhibit the angiogenesis of osteosarcoma cells. The mechanism may be through reducing the expression of FAK in cells and inhibiting the secretion of VEGF by cells.