过表达丝裂原诱导基因6对子宫内膜癌细胞凋亡和侵袭力的影响
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国家自然科学基金(81070471);江苏省“十二五”兴卫工程重点人才资助(RC201161)


Effects of mitogen-inducible gene 6 (MIG6) on apoptosis,proliferation and invasion of endometrial carcinoma
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    摘要:

    目的:观察过表达丝裂原诱导基因6(mitogen-inducible gene 6,MIG6)对子宫内膜腺癌细胞(Ishikawa cell)增殖-凋亡和侵袭能力的影响。方法:脂质体法介导MIG6过表达质粒转染Ishikawa细胞,采用实时荧光定量PCR和Western blot检测转染前-后MIG6表达变化;流式细胞仪检测细胞凋亡的变化;Western blot检测凋亡相关蛋白和周期蛋白的表达;5-溴脱氧尿嘧啶核苷(BrdU)标记法检测细胞增殖情况;Transwell检测细胞侵袭能力的变化。结果:转染MIG6过表达质粒后,Ishikawa细胞中MIG6 mRNA和蛋白的水平分别为对照组的24.2倍和2.8倍,差异具有统计学意义(P < 0.05)。增加Ishikawa细胞中MIG6表达后,裂解的半胱氨酰天冬氨酸蛋白酶-3(cleaved caspase 3)表达显著上升(P < 0.05);早期凋亡率-晚期凋亡率分别较对照组升高1.7倍(P < 0.01)-2.7倍(P < 0.05);Cyclin D1蛋白表达明显下降(P < 0.05),ERK蛋白磷酸化水平下降,pERK/ERK比值降低(P < 0.05);细胞增殖率由(43.4 ± 2.4)%下降至(30.5 ± 4.3)%(P < 0.05);穿过Transwell膜的细胞数从(36.2 ± 6.9)个下降至(26.2 ± 3.8)个(P < 0.05)。结论:MIG6表达上调能抑制Ishikawa细胞的增殖水平和侵袭能力,并促进其凋亡。

    Abstract:

    Objective:To investigate the effects of over-expressed mitogen-inducible gene 6 (MIG6)on apoptosis,proliferation and invasion of endometrial carcinoma cell. Methods:Ishikawa cells were transfected with MIG6 over-expressed plasmid by Lipofectamine 2000,and quantitative RT-PCR and Western Blotting were adopted to detect the expression level of MIG6 before and after transfection. Cell apoptosis and proliferation were measured by flow cytometry and BrdU incorporation assay,respectively. Expression of cleaved caspase 3 and Cyclin D1 was determined by Western Blotting. Invasive potential of Ishikawa cells was tested using Transwell assay. Results:Both mRNA and protein levels of MIG6 in Ishikawa cells transfected with MIG6 over-expression plasmid were significantly higher than those of negative control group. Fold changes were 24.2 and 2.8,respectively. Up-regulated MIG6 expression for 48 h,the expression of cleaved caspase 3 was significantly elevated(P < 0.05),and the early apoptosis rate and late apoptosis rate was 1.7 times and 2.7 times more than that of negative control group(P < 0.01,P < 0.05); the expressions of cyclin D1 and phosphorylation of ERK were both down-regulated significantly(P < 0.05),BrdU incorporation rate decreased from 43.4 ± 2.4% to 30.5 ± 4.3%(P < 0.05); meanwhile,the amount of trans-membrane cells decreased from 36.2 ± 6.9 to 26.2 ± 3.8(P < 0.05). Conclusion:Up-regulation of MIG6 inhibits proliferation and invasion,while promotes apoptosis of endometrial carcinoma Ishikawa cells in vitro.

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徐 炜,施 秀,晋一超,戴辉华,王秀丽.过表达丝裂原诱导基因6对子宫内膜癌细胞凋亡和侵袭力的影响[J].南京医科大学学报(自然科学版),2015,(5):626-630,643

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  • 收稿日期:2015-01-29
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  • 在线发布日期: 2015-05-22
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