载脂蛋白E对结直肠癌细胞侵袭迁移能力的影响

Effect of Apolipoprotein E on Invasion and Migration of Colorectal Cancer Cells

目的观察载脂蛋白E(apolipoprotein E,Apo E)对结直肠癌细胞侵袭迁移能力的影响。方法在结直肠癌细胞系SW48中转染ApoE过表达质粒及其特异性siR NA,实时荧光定量PCR(Real timeq PCR)检测转染Apo E的表达,利用Transwell小室观察Apo E对细胞侵袭迁移能力的影响,通过免疫印迹检测Apo E对基质金属蛋白酶(matrix metalloproteinase,MMP)家族蛋白表达的影响。结果 RTPCR检测结果显示:过表达Apo E组细胞中Apo E m RNA水平显著高于空白对照白组和阴性对照组,差异有统计学意义(P=0.008),Apo E沉默组中Apo E m RNA水平与阴性对照组比,差异有统计学意义(P=0.013);Transwell侵袭实验结果显示,穿膜细胞数:空白对照组、阴性对照组、过表达Apo E组、si-ApoE组依次为:(209±17)、(228±11)、(67±9)、(428±15)个/视野,过表达ApoE组和si-ApoE组与阴性对照组相比,差异有统计学意义(P<0.05);Transwell迁移实验结果显示,空白对照组(326±18)个/视野,阴性对照组(338±21)个/视野,过表达ApoE组(133±11)个/视野,与阴性对照组相比,差异有统计学意义(P<0.05),si-Apo E组(518±13)个/视野与阴性对照组相比,差异有统计学意义(P<0.05)。同时免疫印迹实验显示,过表达Apo E组细胞中MMP-2及MMP-9的表达下降,组织基质金属蛋白酶抑制剂2(tissue inhibitors of metalloproteinase-2,TIMP-2)的表达升高,而si-ApoE组MMP-2及MMP-9表达升高,TIMP-2的表达降低。结论 Apo E可影响结直肠癌细胞的侵袭、迁移能力,并且可能通过影响TIMP-2、MMP-2及MMP-9的表达发挥上述功能。

Objective To observe the effect of apolipoprotein E(Apo E) on the invasion and migration ability of colorectal cancer(CRC) cells. Methods The Apo E was over-expressed or silenced by plasmids or si RNAs transfection in CRC cell line SW48, and the expression of Apo E was tested by Real-time-q PCR; then the effect of Apo E on the invasion and migration ability of SW48 cells was detected by Transwell assay; and the effect of Apo E on the expression of matrix metalloproteinase(MMP) family proteins was detected by Western blot. Results RT-PCR test result showed that Apo E m RNA expression in overexpressed Apo E group was significantly higher that those in blank control group and negative control group(P=0.008); there was significant difference in Apo E m RNA expression between silenced Apo E group and negative control group(P=0.013).Transwell invasion assay result showed that cells count in blank control group was(209±17)/field, negative control group(228±11)/field, overexpressed Apo E group(67±9)/field, silenced Apo E(428±15)/field, compared with negative control group(P<0.05); Transwell migration assay result showed that cells count in blank control group was(326±18)/field, negative control group(338±21)/field, overexpressed Apo E group(133±11)/field, silenced Apo E group(518±13)/field, compared with negative control group(P<0.05). Meantime, the overexpression of Apo E could reduce the expression of MMP-2 and MMP-9, and elevate the expression of tissue inhibitors of metalloproteinase-2(TIMP-2); the silenced Apo E could promote the expression of MMP-2 and MMP-9, and decline the expression of TIMP-2. Conclusion Apo E could regulate the invasion and migration ability of CRC cells, which may be via regulating TIMP-2, MMP-2 and MMP-9 expression.