S100P促进结直肠癌细胞增殖和迁移的机制探讨

Mechanism of S100P promoting the proliferation and migration of the colorectal cancer cells

目的:初步探讨S100P对结直肠癌细胞生物学行为的影响及作用机制。方法:构建S100P真核细胞表达载体转染不表达S100P的野生型SW480结肠癌细胞,细胞生长曲线法、MTT法检测S100P对SW480细胞生长增殖的影响;流式细胞术分析S100P基因转染后细胞周期分布的变化;细胞划痕实验评价S100P对SW480细胞迁移能力的影响;免疫印迹法检测S100P对SW480胞外信号调节蛋白激酶1/2磷酸化水平的影响。结果:稳定转染S100P基因的SW480-S100P细胞生长增殖速度加快,细胞群体倍增时间由20h左右缩短至15h。细胞周期发生明显改变,野生型及转染空载体SW480细胞的S期比例分别为(21.9±0.8)%、(21.4±1.8)%,而转染S100P的SW480细胞为(30.6±3.8)%,差异有显著性(P=0.007)。细胞划痕24h后,野生型SW480、空载体SW480和SW480-S100P 3株细胞的平均迁移细胞数分别为(11.3±3.1)个、(10.7±3.7)个、(19.3±3.5)个,组间有明显差异(P=0.035)。S100P基因转染SW480细胞后,胞外信号调节蛋白激酶1/2的磷酸化水平升高,分别是野生型SW480和空载体SW480细胞的4.2和3.7倍(P=0.001)。结论:S100P可显著影响结直肠癌细胞的生物学行为,通过受体影响胞外信号调节蛋白激酶1/2信号转导通路可能是其发挥促细胞增殖和迁移作用的机制之一。

Objective： To investigate the effects of S100P protein on the biological behaviors of colorectal cancer cell lines. Methods： The mammalian expression vector of S100P was constructed and transfected into colorectal cancer cell line SW480 which was S100P negative in wild type. The effects of S100P on the proliferation of SW480 cells were studied using MTT assay and cell growth curve. The cell cycle was assessed by flow cytometry and the ability of cell migration was evaluated by wound healing assay. Phosphorylated extracellular signal regulated protein kinase 1 /2（ ERK1 /2） was detected by Western blot. Results： MTT assay and cell growth curve analysis showed that SW480-S100P cells proliferated more rapidly than wide type SW480 and SW480- vector cells. Average doubling time of cell population was reduced from about 20 h in wild type SW480 and SW480- vector cells to 15 h in SW480- S100P cells. Flow cytometry analysis showed a higher proportion of S phase in SW480- S100 P cells compared with SW480-vector cells as well as wild type SW480 cells（ P = 0. 007）. Twenty- four hours after wounding,the cell number of SW480- S100 P migration were 19. 3 ± 3. 5,much more than the number of migrated wild type SW480 and SW480-vector cells（ 11. 3 ± 3. 1 and 10. 7 ± 3. 7）,respectively（ P = 0. 035）. The levels of phospho- ERK1 /2 in SW480-S100 P cells were up- regulated,being 4. 2 or 3. 7 times more than that in wild SW480 or SW480- vector cells（ P =0. 001）. Conclusion： S100 P could promote proliferation and migration of colorectal cancer cells and regulate cell cycle. Increasing the level of phospho- ERK1 /2 was a possible mechanism underlying these processes.