Cao Jing, Wang Meng, Wang Tao
Department of Pharmacy, Linyi People's Hospital of Shandong University, Linyi, Shandong 276000, P.R. China.
Department of Opthalmology, Linyi People's Hospital of Shandong University, Linyi, Shandong 276000, P.R. China.
Exp Ther Med. 2017 Aug;14(2):1554-1560. doi: 10.3892/etm.2017.4659. Epub 2017 Jun 23.
The aim of the present study was to examine the effect of CCAAT enhancer binding protein β (C/EBPβ) on human breast cancer cells. The plasmids pCDH-C/EBPβ and pLKO.1-shC/EBPβ were constructed and were infected into MDA-MB-468 cells, to provide C/EBPβ overexpressing and C/EBPβ knockdown cells, respectively. Cell viability, cell cycle and apoptosis were observed by MTT assay and flow cytometry analysis. Protein expression levels of C/EBPβ, TGF-β1, P-Smad3 and Smad3 were detected by western blotting. MTT assay showed that the absorbance of MDA-MB-468 cells in the pCDH-C/EBPβ group was increased, whereas that in the pLKO.1-shC/EBPβ group was decreased, compared with the respective control at 48 and 72 h. Flow cytometric analysis indicated that the percentage of cells in the G2 phase was significantly increased in the pCDH-C/EBPβ group (P<0.05) and decreased in the pLKO.1-shC/EBPβ group compared with the respective control group. The proportion of apoptotic cells was decreased in the pCDH-C/EBPβ group and increased in the pLKO.1-shC/EBPβ group compared with the controls. The scratch-wound assay revealed that MDA-MB-468 cells depleted of C/EBPβ exhibited reduced motility compared with the control cells. Moreover, western blotting demonstrated that pCDH-C/EBPβ increased transforming growth factor (TGF)β1 and P-Smad3 protein expression and decreased Smad3 protein expression, whereas pLKO.1-shC/EBPβ decreased TGFβ1 and P-Smad3 protein expression and increased Smad3 protein expression levels. The present study demonstrated that C/EBPβ has a crucial role in regulating breast cancer cell growth through activating TGF-β-Smad3 signaling. These findings suggest that C/EBPβ may be a potential therapeutic target for breast cancer; however, studies are required to confirm this.
本研究的目的是检测CCAAT增强子结合蛋白β（C/EBPβ）对人乳腺癌细胞的影响。构建了质粒pCDH-C/EBPβ和pLKO.1-shC/EBPβ，并将其感染MDA-MB-468细胞，分别获得C/EBPβ过表达和C/EBPβ敲低的细胞。通过MTT法和流式细胞术分析观察细胞活力、细胞周期和凋亡情况。通过蛋白质印迹法检测C/EBPβ、转化生长因子（TGF）-β1、磷酸化Smad3（P-Smad3）和Smad3的蛋白表达水平。MTT法显示，与各自的对照组相比，在48小时和72小时时，pCDH-C/EBPβ组MDA-MB-468细胞的吸光度增加，而pLKO.1-shC/EBPβ组的吸光度降低。流式细胞术分析表明，与各自的对照组相比，pCDH-C/EBPβ组G2期细胞的百分比显著增加（P<0.05），而pLKO.1-shC/EBPβ组则降低。与对照组相比，pCDH-C/EBPβ组凋亡细胞的比例降低，而pLKO.1-shC/EBPβ组则增加。划痕试验显示，与对照细胞相比，C/EBPβ缺失的MDA-MB-468细胞的运动能力降低。此外，蛋白质印迹法表明，pCDH-C/EBPβ增加了TGFβ1和P-Smad3蛋白表达，降低了Smad3蛋白表达，而pLKO.1-shC/EBPβ降低了TGFβ1和P-Smad3蛋白表达，并增加了Smad3蛋白表达水平。本研究表明，C/EBPβ通过激活TGF-β-Smad3信号通路在调节乳腺癌细胞生长中起关键作用。这些发现表明，C/EBPβ可能是乳腺癌的一个潜在治疗靶点；然而，需要进一步研究来证实这一点。
