RUNX3参与抑制人前列腺癌迁移和血管发生

目的研究RUNX3基因对前列腺癌的发生与迁移的影响，并探讨其分子作用机制。方法运用肿瘤组织芯片（tissue microarray，TMA）技术评估RUNX3在前列腺癌发生过程中的作用；分别用RUNX3的真核表达载体pFlag—RUNX3和对照载体pFlag—control转染人前列腺癌细胞系PC3和DU145细胞；细胞迁移实验、细胞侵袭实验和微血管形成实验分别检测RUNX3基因高表达对2种前列腺癌细胞迁移、侵袭和促血管生成能力的影响；利用RT—PCR和Western blot检测RUNX3基因表达后对2种前列腺癌细胞中基质金属蛋白酶抑制因子-2（tissue inhibitor of metalloproteinase-2，TIMP-2）和基质金属蛋白酶-2（metalloproteinase-2，MMP-2）mRNA和蛋白表达的影响；明胶酶谱实验和ELISA实验检测RUNX3基因表达后对2种前列腺癌细胞分泌活性MMP-2和血管内皮生长因子（vascular endothelial growth factor，VEGF）的影响。结果相较于癌旁组织，RUNX3在前列腺癌组织中异常低表达，且与肿瘤分级有关。过表达RUNX3会上调TIMP-2、抑制MMP-2的表达与激活，从而抑制前列腺肿瘤细胞的迁移与侵袭。在前列腺细胞中抑制RUNX3表达会破坏TIMP-2和MMP-2之间的平衡，沉默TIMP-2会抑制MMP-2的表达。恢复RUNX3表达会降低VEGF的分泌，从而抑制内皮细胞生长和血管生成。结论RUNX3通过TIMP-2和VEGF通路对前列腺癌产生抑制作用。

The role of RUNX3 in suppressing metastasis and angiogenesis of human prostate cancer

Objective To investigate the role of RUNX3 gene in cell migration, invasion and its molecular mechanisms in PC3 and DU145 prostate cancer cell lines. Methods We used tissue microarray （TMA） to determine the significance of RUNX3 in prostate cancer progession. The plasmids pFlag - control or pFlag - RUNX3 was transfected into PC3 and DU145 prostate cancer cells. Then we studied the migration, invasion and tube formation abilities in these cells by performing cell migration, invasion, and tube formation assay. RT - PCR and Western blot were used to determine the mRNA and protein expressions of TIMP - 2 and MMP - 2 after RUNX3 restoration in prostate cancer cells. Gelatin zymography was performed to detect the MMP -2 activities and VEGF concentration was determined using Quantikine ELISA kits. Results The expression of RUNX3 in prostate cancer tissues was significantly lower than in tumor adjacent normal prostate tissues, and reduced RUNX3 staining was significantly correlated with TNM stage. Moreover, RUNX3 overexpression inhibited prostate cancer Cell migration and invasion resulting from the elevated upregnlation of TIMP - 2, which subsequently inhibited MMP -2 expression and activity in vitro. Knock down of RUNX3 expression broke up the balance of TIMP - 2/MMP - 2, whereas silence of TIMP - 2 resulted in the inhibition of MMP - 2 expression in prostate cells. Restoration of RUNX3 decreased VEGF secretion and suppressed endothelial cell growth and tube formation. Conclusions Our results support the tumor suppressive role of RUNX3 in human prostate cancer through the MMP -2 and VEGF pathway.