高尔基磷酸化蛋白-3基因过表达对胰腺癌细胞迁移的影响

Effect of golgi phosphorylation protein 3 gene overexpression on migration of pancreatic cancer cells

目的构建高尔基磷酸化蛋白-3（GOLPH-3）基因过表达的胰腺癌稳定细胞株BxPC-3，观察GOLPH-3基因过表达对胰腺癌细胞迁移能力的影响。方法制备逆转录病毒pMSCV—GOLPH-3和pMSCV—vector，并分别感染胰腺癌细胞株BxPC-3，用0．5mg／L嘌呤霉素筛选得到GOLPH-3过表达及空载对照稳定细胞株。Westernblot技术在蛋白水平上检测BxPC-3稳定细胞株中GOLPH．3及β-连环蛋白（13-catenin）的蛋白表达水平，细胞迁移实验（Transwell）检测GOLPH-3对胰腺癌细胞迁移能力的影响。结果胰腺癌细胞BxPC-3表达GOLPH-3蛋白；空白对照组、空载对照组及过表达组的GOLPH．3蛋白相对含量分别为0．32±0．13、0．46±0．12、1．02±0．30，过表达组较空载对照组上调50．9％，其差异有统计学意义（P〈0．05）。β-catenin蛋白相对含量分别为0．42±0．06、0．63±0．34、1．77±0．19，与空载对照组比较，上调GOLPH．3能明显促进过表达组β-catenin蛋白的表达，且上调64．4％（P〈0．01）。每200倍随机计数5个视野下空白对照组、空载对照组、过表达组穿膜胰腺癌细胞数分别为（14．33±6．30）个、（16．25±10．45）个、（83．08-4-62．57）个，过表达组的迁移细胞数明显多于空载对照组（P〈0．001），而两对照组间差异无统计学意义（P〉0．05）。结论上调GOLPH-3能影响胰腺癌细胞的迁移能力，GOLPH-3过表达增强胰腺癌细胞的迁移机制可能是上调Wnt／β-eatenin信号通路B—eatenin表达的结果。

Objective To establish stable overexpression of golgi phosphorylation protein 3 （GOLPH-3） gene in the pancreatic cancer cell line BxPC-3 and observe the effect of GOLPH-3 gene over- expression on migration of pancreatic cancer cells. Methods The BxPC-3 cells were transfected by retro- viruses of pMSCV-GOLPH and pMSCV-vector respectively. The stable cell lines were obtained after screen- ing by 0. 5 mg/L puromyein. The protein expression levels of GOLPH-3 and ^-eatenin were detected by u- sing Western blotting. The migration of BxPC-3 stable cells was tested by transwell assay. Results BxPC-3 cells could express GOLPH-3 protein. The relative protein expression of GOLPH-3 was 0. 32 ± 0. 13, 0. 46 ±0. 12 and 1.02 ± 0. 30 in the blank, vector control and overexpression groups, respectively. As compared with the vector control group, the protein levels of the GOLPH-3 in overexpression group was sig- nificantly up-regulated by 50. 90% （P 〈 0. 05 ）. The gray value of β-catenin protein expression in the blank, vector control and overexpression groups was 0. 42 ± 0. 06, 0. 63 ± 0. 34 and 1.77 ± 0. 19 respec- tively. As compared with the vector control group, up-regulated GOLPH-3 could promote the protein expression of β-catenin in the overexpression group which was up-regulated by 64. 4% （P 〈 0. 01 ）. The mean migrating cell number was 14. 33 ±6. 30 in the blank control group, 16. 25 ± 10.45 in the vector con- trol group and 83.08 ± 62. 57 in the overexpression group. The number of cells positive for GOLPH-3 in the overexpression group was significantly greater than that in the vector control group （ P 〈 0. 001 ）. Conclu- sion Overexpresssion of GOLPH-3 could increase the migration of pancreatic cancer cells, which might be related to the up-regulated expression of β-catenin in Wnt-β--eatenin signal pathway.