过表达人SNX3改变SW620结直肠腺癌细胞的形态

The morphological changes of SW620 cells induced by over-expression of human sorting nexin 3

目的通过过表达人分选连接蛋白3基因(h SNX3),以探讨h SNX3对结直肠腺癌细胞形态和迁移能力的影响。方法运用基因重组技术将h SNX3基因连接到含有GFP的慢病毒表达载体p EZ-Lv201(p EZ-SV40-e GFP-IRES-Puro)中,构建慢病毒载体p EZ-h SNX3-Lv201,感染结直肠腺癌SW620细胞,获得SNX3稳定过表达的结直肠腺癌细胞,并观察细胞形态和细胞迁移能力及相关蛋白分子的变化。结果经测序鉴定后,成功构建了慢病毒载体p EZ-h SNX3-Lv201,包装后慢病毒滴度测定为2.12×109拷贝/m L,对照慢病毒滴度为7.9×1010拷贝/m L。重组慢病毒感染SW620细胞后,经克隆筛选和Western blot法鉴定,获得了稳定过表达h SNX3的SW620h SNX3细胞。88%的SW620h SNX3细胞呈椭圆形而不是SW620原来的梭形,但TranswellTM实验和划痕试验显示,细胞迁移能力无显著变化,迁移相关蛋白上皮型钙黏素(E-cadherin)也无明显改变。结论h SNX3可改变SW620结直肠腺癌细胞的形态,但可能与细胞的迁移能力无关。

Objective To explore the effect of human sorting nexin 3（h SNX3） on the morphology and migration of colorectal cancer cells through over-expression of h SNX3 in SW620 cells. Methods A lentiviral vector p EZ-Lv201（ p EZ-SV40-e GFPIRES-Puro） inserted with h SNX3 gene was transferred into SW620 cells to construct and package recombinant lentiviral expression vector p EZ-h SNX3-Lv201. The cell clones stably over-expressing h SNX3（ SW620 h SNX3） were screened to observe cell morphology and migration ability using TranswellTMassay and wound scratch assay. E-cadherin was detected by Western blotting. Results Sequence analysis demonstrated that the lentiviral expression vector p EZ-h SNX3-Lv201 was successfully constructed. The packaging titers of the recombinant lentivirus and control virus were 2. 12 × 10^9 copies / m L and 7. 9 × 10^10 copies / m L,respectively. After SW620 cells were transfected with the lentivirus,we got stable h SNX3-overexpressed SW620 h SNX3 cells. Most of SW620 h SNX3 cells（88%） became oval instead of the original spindle-like shape. However,cell migration ability and expression of E-cadherin showed no significant differences between SW620 h SNX3 cells and SW620 cells. Conclusion Over-expression of h SNX3 alters the morphology of SW620 cells,but hardly affects cell migration.