小RNA干扰MMP-2基因表达对结直肠癌细胞生物学特性的影响

刘秀兰; 陈世杰

doi:10.13898/j.cnki.issn.1000-2200.2019.10.003

小RNA干扰MMP-2基因表达对结直肠癌细胞生物学特性的影响

Effect of the expression of siRNA targeting MMP-2 on the biological characteristics of colorectal cancer cells

摘要

摘要:
目的探讨小RNA干扰基质金属蛋白酶2（MMP-2）基因表达对结直肠癌细胞生物学特性的影响。
方法收集结直肠癌病人的结直肠癌组织及正常的癌旁组织，Western blotting检测MMP-2表达水平。取结直肠癌细胞SW620作为对照组，将MMP-2 siRNA、siRNA control转染至结直肠癌细胞中，Western blotting检测转染48 h后MMP-2 siRNA组、siRNA control组和对照组细胞中MMP-2水平，MTT法检测各组细胞存活情况，流式细胞术检测各组细胞凋亡变化，Western blotting检测各组细胞中B细胞淋巴瘤/白血病-2（Bcl-2）、β-连环蛋白（β-catenin）、Bcl-2相关X蛋白（Bax）、下游靶基因C-myc、细胞周期蛋白D1（cyclin D1）水平。结直肠癌细胞与Wnt/β-catenin信号通路抑制剂FH535作用后，测定抑制剂组和未处理组（不加抑制剂）细胞增殖、凋亡变化，同时检测细胞中Bcl-2、β-catenin、Bax、C-myc、cyclin D1蛋白变化。
结果结直肠癌组织中MMP-2蛋白表达水平明显高于癌旁组织（P < 0.01）。MMP-2 siRNA组MMP-2水平均低于对照组和siRNA control组（P < 0.05），细胞存活率低于对照组（P < 0.05），细胞凋亡率均高于对照组和siRNA control组（P < 0.05）。Bcl-2、β-catenin、C-myc、cyclin D1水平均低于对照组和siRNA control组（P < 0.05），Bax水平均高于对照组和siRNA control组（P < 0.05）。Wnt/β-catenin信号通路抑制剂作用后，抑制剂组细胞存活率低于未处理组（P < 0.05），细胞凋亡率高于未处理组（P < 0.05），Bcl-2、β-catenin、C-myc、cyclin D1水平均明显低于未处理组（P < 0.01），Bax水平明显高于未处理组（P < 0.01），与转染MMP-2 siRNA的结直肠癌细胞一致。
结论MMP-2在结直肠癌组织中表达上调，抑制MMP-2的结直肠癌细胞增殖受到抑制，凋亡增多，其作用机制与Wnt/β-catenin信号通路有关。

Abstract:
ObjectiveTo investigate the effects of the expression of siRNA targeting MMP-2 on the biological characteristics of colorectal cancer cells.
MethodsThe expression levels of MMP-2 in colorectal cancer tissues and normal adjacent tissue was detected using Western bloting.The SW620 colorectal cancer cell was set as the control, and the MMP-2 siRNA and siRNA control were transfected into colorectal cancer cells.After 48h of transfection, the expression levels of MMP-2 in MMP-2 siRNA group, siRNA control group and control group were detected using Western bloting, and the survival rate and apoptosis rate of colorectal cancer cells in three groups were detected using MTT assay and flow cytometry, respectively.The expression levels of Bcl-2, Bax, β-catenin, C-myc and cyclin D1were detected using Western bloting.After colorectal cancer cells and Wnt/β-catenin signaling pathway were treated with the inhibitor FH535, the proliferation and apoptosis of cells in inhibitor group and untreated group were detected, and the expression levels of Bcl-2, Bax, β-catenin, C-myc and cyclin D1 were detected.
ResultsThe expression level of MMP-2 in colorectal carcinoma tissue was significantly higher than that in adjacent tissues(P < 0.01).The expression level of MMP-2 in MMP-2 siRNA group was lower than that in control group and siRNA group(P < 0.05), the survival rate of colorectal cancer cells transfected with MMP-2 siRNA was lower than that in control group(P < 0.05), and the apoptosis rate in MMP-2 siRNA group was higher than that in control group and siRNA group(P < 0.05).The levels of Bcl-2, β-catenin, C-myc and cyclin D1 in MMP-2 siRNA group were significantly lower than those in control group and siRNA control group(P < 0.05), the level of Bax in MMP-2 siRNA group was higher than that in control group and siRNA group(P < 0.01).After the Wnt/β-catenin signaling pathway was treated with the inhibitor, the survival and apoptosis rates of cells in inhibitor group were lower and higher than that in untreated group, respectively(P < 0.05), the levels of Bcl-2, β-catenin, C-myc and cyclin D1 in inhibitor group were significantly lower than that in untreated group(P < 0.01), the level of Bax in inhibitor group was significantly higher than that in untreated group(P < 0.01), which was consistent with that in colorectal cancer cells transfected with MMP-2 siRNA.
ConclusionsThe MMP-2 expression is up-regulated in colorectal cancer tissue.The proliferation of colorectal cancer cells with inhibiting MMP-2 expression is inhibited, the apoptosis of colorectal cancer cells increase, and the mechanism of which is related to Wnt/β-catenin signaling pathway.

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