Effect of Apolipoprotein E on Invasion and Migration of Colorectal Cancer Cells
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摘要: 目的 观察载脂蛋白E（apolipoprotein E, ApoE）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siRNA，实时荧光定量PCR（Real timeqPCR）检测转染ApoE的表达，利用Transwell小室观察ApoE对细胞侵袭迁移能力的影响，通过免疫印迹检测ApoE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ApoE组细胞中ApoE mRNA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），ApoE沉默组中ApoE mRNA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ApoE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-ApoE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ApoE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 ApoE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。Abstract: Objective To observe the effect of apolipoprotein E (ApoE) on the invasion and migration ability of colorectal cancer (CRC) cells. Methods The ApoE was over-expressed or silenced by plasmids or siRNAs transfection in CRC cell line SW48, and the expression of ApoE was tested by Real-time-qPCR; then the effect of ApoE on the invasion and migration ability of SW48 cells was detected by Transwell assay; and the effect of ApoE on the expression of matrix metalloproteinase(MMP) family proteins was detected by Western blot. Results RT-PCR test result showed that ApoE mRNA expression in overexpressed ApoE group was significantly higher that those in blank control group and negative control group (P=0.008); there was significant difference in ApoE mRNA expression between silenced ApoE 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 ApoE group (67±9)/field, silenced ApoE (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 ApoE group (133±11)/field, silenced ApoE group (518±13)/field, compared with negative control group (P<0.05). Meantime, the overexpression of ApoE could reduce the expression of MMP-2 and MMP-9, and elevate the expression of tissue inhibitors of metalloproteinase-2 (TIMP-2); the silenced ApoE could promote the expression of MMP-2 and MMP-9, and decline the expression of TIMP-2. Conclusion ApoE could regulate the invasion and migration ability of CRC cells, which may be via regulating TIMP-2, MMP-2 and MMP-9 expression.
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