KAI1基因转染对乏氧培养下胰腺癌MiaPaCa-2细胞增殖、迁移及VEGF表达的影响

Effects of KAI1 gene transfection on proliferation, migration, invasion and VEGF expression of pancreatic cancer MiaPaCa-2 cells under hypoxic condition

目的 观察转染KAI1基因的人胰腺癌MiaPaCa-2细胞在乏氧条件下培养后细胞增殖、迁移、侵袭能力的变化,探讨其可能机制.方法 应用KAl1基因过表达质粒转染乏氧条件培养后的人胰腺癌MiaPaCa-2细胞,采用蛋白质印迹法检测转染细胞KAI1、VEGF-C、VEGF-A蛋白的表达,四甲基偶氮唑蓝（MTT）法检测转染细胞的增殖,细胞划痕及Transwell小室实验观察转染细胞的迁移及侵袭能力,酶联免疫吸附测定法检测培养上清液中人VEGF-C、VEGF-A含量.结果 转染KAI1基因后的MiaPaCa-2-K细胞的KAI1蛋白表达量较未转染细胞显著增加[（0.549 ±0.021）比0].乏氧条件培养后转染细胞的增殖无明显变化,但它的迁移距离明显缩短,穿膜细胞数显著减少[（14.0±5.8）比（43.0±14.4）个,P＜0.05];细胞的VEGF-C表达显著降低[（0.218±0.043）比（0.745±0.069）.P＜0.05],但VEGF-A表达变化不显著;细胞培养上清液中VEGF-C含量显著减少[（1236±247）比（2045±221） pg/ml,P＜0.01].结论 转染KAl1基因的MiaPaCa-2细胞在乏氧条件下培养后的细胞迁移、侵袭能力减弱,其机制可能是通过下调VEGF-C的表达来抑制胰腺癌淋巴转移的.

Objective To investigate the effects of KAll gene transfection on proliferation,migration,invasion and VEGF expression of pancreatic cancer MiaPaCa-2 cells under hypoxic condition,and explore possible mechanism.Methods The pcMV-KAI1 vector which contained the full length of KAI1 cDNA was transfected into pancreatic cancer cells MiaPaCa-2,and KAI1,VEGF C,VEGF A protein expressions were determined by Western blot.The proliferation of pancreatic cancer cells was evaluated by MTT method.Wound-healing assay and cell invasion assay were used to detect the migration and invasion of pancreatic cancer cells.The expression of VEGF C,VEGF A in supernatant of culture was measured by ELISA method.Results The expression of KAI1 protein in MiaPaCa-2 K transfected with KAI1 was significantly higher than that in nontransfected cells [（0.549 ± 0.021） vs 0,P〈0.05].The proliferation under hypoxic condition was not significantly different,but the migration distance was significantly shorter and the number of transmembrane cell was significantly decreased [（14.0 ± 5.8） vs （43.0 ± 14.4）,P 〈 0.05].The expression of VEGF-C in cell was significantly decreased [（0.218 ± 0.043） vs （0.745 ± 0.069）,P 〈0.05],but the expression of VEGF-A was not significantly different; the expression of VEGF-C in cell culture supernatants was significantly decreased [（1236 ± 247） vs （2045 ± 221） pg/ml,P 〈 0.01].Conclusions The migration,invasion ability of pancreatic cancer MiaPaCa-2 cells with KAll transfection under hypoxic condition is decreased,and the possible mechanism of inhibiting lymphatic metastasis is down-regulation of VEGF-C expression.