线粒体融合蛋白-2基因增强人乳腺癌T47D细胞对小白菊内酯的敏感性

Mitofusin-2 gene enhances sensitivity of human breast cancer T47D cells to parthenolide

目的:探讨线粒体融合蛋白-2(mitofusin-2,Mfn-2)基因表达对人乳腺癌T47D细胞对小白菊内酯敏感性的影响。方法:Real-time PCR检测不同乳腺癌细胞系(T47D、MDA-MB-231、MCF-7、MDA-MB-435及HCC38)中Mfn-2 mRNA的表达。LipofectamineTM2000体外转染pEGFP和pEGFP-Mfn-2质粒至人乳腺癌T47D细胞,real-time PCR和Western blotting检测T47D细胞中Mfn-2 mRNA和蛋白的表达,MTT法检测T47D细胞的增殖,流式细胞术检测T47D细胞凋亡率及线粒体膜电位。结果:与正常乳腺细胞相比,Mfn-2 mRNA在乳腺癌HCC38细胞系中高表达,在T47D等其他细胞系中均低表达。pEGFP-Mfn-2转染48 h后,T47D细胞中Mfn-2 mRNA和蛋白的表达均明显上调。与pEGFP转染组相比,pEGFP-Mfn-2转染组T47D细胞在小白菊内酯(0.05 mol/L)作用下的存活率明显降低[(47.93±2.21)%vs(56.93±2.05)%,P<0.05]。流式细胞术检测结果显示:50 mmol/L小白菊内酯作用下,pEGFP-Mfn-2转染组与pEGFP转染组相比,T47D细胞凋亡率升高[(71.2±2.1)%vs(38.8±2.6)%,P<0.05],而线粒体膜电位明显降低[(1.6±0.1)%vs(5.0±0.5)%,P<0.05]。结论:pEGFP-Mfn-2转染可增强T47D细胞对小白菊内酯的敏感性。

Objective：To investigate the effect of mitofusin-2 （ Mfn-2 ） gene expression on sensitivity of human breast cancer T47D cells to parthenolide. Methods： The expressions of Mfn-2 mRNA in various breast cancer cell lines （T47D, MDA-MB-231, MCF-7, MDA-MB-435 and HCC38） were detected by real-time PCR. Plasmids pEGFP and pEGFP-Mfn-2 were transfected into human breast cancer T47D cells by LipofectamineTM 2000 in vitro. The expression levels of Mfn-2 mRNA and protein in T47D cells were detected by real-time PCR and Western blotting. MTT assay was used to detect the proliferation of T47D cells. The cell apoptotic rate and mitochondrial membrane potential of T47D cells were measured by flow cytometry. Results： Compared with that in normal breast cells, Mfn-2 mRNA was highly expressed in breast cancer HCC38 cell line, and lowly expressed in the other cell lines, such as T47D etc. After pEGFP-Mfn-2 transfection for 48 h, the expression levels of Mfn-2 mRNA and rotein were significantly up-regulated in T47D cells. Compared with the pEGFP transfection group, the pEGFP-Mfn-2 transfection group showed a significant decrease in surivival rate of T47D cells under the treatment of parthenolide （50 mmol/L） （／[47.93±2.21／]% vs ／[56.93±2.05／]%, P〈0.05）. Flow cytometry results showed that the apoptotic rate of T47D cells under the treatment of 0.05 mol/L parthenolide was significantly increased in pEGFP-Mfn-2 transfection group compared with that in pEGFP ransfection group （／[71.2±2.1／] % vs ／[38.8±2.6／] %, P〈005）. However, the mitochondrial membrane potential was significanly decreased in the pEGFP-Mfn-2 transfection group （／[1.6±0.1／] % vs ／[5.0±0.5／] %, P〈0.05）. Conclusion： pEGFP-Mfn-2 transfection can enhance the sensitivity of T47D cells to parthenolide.