caveolin-1过表达抑制胰腺癌细胞的增殖和侵袭

Over-expression of caveolin-1 inhibits proliferation and invasion of pancreatic carcinoma cells in vitro

目的体外研究caveolin-1对胰腺癌细胞生物学行为及相关信号通路的影响。方法caveolin-1基因脂质体法稳定转染胰腺癌细胞株panc-，筛选出稳定高表达caveolin-1的克隆，并用实时荧光定量PCR和免疫印迹法鉴定。应用四甲基偶氮唑蓝（MTI＂）法检测细胞的生长能力，软琼脂克隆生长实验检测细胞锚定非依赖性生长能力，流式细胞仪检测细胞周期和凋亡情况，细胞侵袭实验检测细胞侵袭能力，免疫印迹法检测表皮生长因子受体（EGFR）、c—Raf、Mek、Erk、p38和SAPK／JNK的表达。结果转染panc1细胞后筛选到3株稳定高表达caveolin-1的克隆。MTT法检测结果显示，转染后的细胞生长速度明显慢于对照组，软琼脂中克隆生长能力较对照组也明显下降。流式细胞术检测结果显示，高表达caveolin-1可使细胞阻滞在G0／G1期，并且促进细胞的凋亡。细胞侵袭实验显示，转染caveolin-1后，细胞的侵袭能力明显下降，此外，caveolin-1过表达降低了EGFR、C—Raf、Mek和Erk的磷酸化水平。结论caveolin-1基因过表达抑制了胰腺癌细胞panc1的生长和侵袭能力，对EGFR—C—Raf-Mek—Erk信号级联的抑制与其抑制胰腺癌细胞恶性表型的作用相关。

Objective To investigate the effects of caveolin-1 on the biologic behavior of pancreatic carcinoma cell line panel cells in vitro. Methods Eukaryotic expression vectors containing human caveolin-1 gene was stably transfeeted into panel cells with Lipofectamine2000. The clones stably overexpressing caveolin-1 were identified by real-time PCR and Western bloting. The cell growth activity was examined by MTT assay. Anchorage-independent growth was detected by colony formation assay in soft agar. Flow cytometry was used to analyze the cell cycle and apoptosis. Cell invasion assay was used for evaluating cell invasion capacity. The relative phosphorylation level of EGFR, c-Raf, Mek, Erk, p38 and SAPK/JNK were detected by Western blotting. Results Three transfected eel] clones overexpressing caveolin-1 were obtained. Comparing with the panel cells, the transfected cells exhibited a slower growth rate and formed fewer colonies in soft agar. The results of flow cytometry showed that over-expression of caveolin-1 resulted in the cell cycle arrest at G0/Gl phase and increased the apoptotic cell fraction. Cell invasion assay showed that overexpression of caveolin-1 significantly inhibited the panel cell invasion. Western blotting results showed that overexpression of caveolin-1 reduced the phosphorylation of EGFR, c-Raf, Mek and Erk while did not affect the activity of p38 and SAPK/JNK. Conclusion Over-expression of caveolin-1 inhibits the growth and invasion of pancreatic carcinoma cells in vitro. Thesephenotypes may be correlated with the inhibition of EGFR-c-Raf-Mek-Erk signaling pathway.