Aletaha Mansoor, Mansoori Behzad, Mohammadi Ali, Fazeli Mehdi, Baradaran Behzad
Biotechnology, Department of Pathobiology, Shiraz University, Shiraz, Iran.
Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
Biomed Pharmacother. 2017 Apr;88:34-42. doi: 10.1016/j.biopha.2017.01.030. Epub 2017 Jan 13.
Despite the great improvements in clinical and therapeutic techniques in recent years, many advanced breast cancer patients still died of the postoperative recurrence and metastasis of disease. Bach1 plays a role in the development of the invasive phenotypes of cancer, cell division and apoptosis in tumor cells. The aim of this study was to investigate the effect of specific bach1 siRNAs, on the proliferation, migration, invasive, induction of apoptosis, cell cycle arrest and alter EMT related miRNA of MDA-MB-468 cells (breast cancer).
siRNA transfection was performed with transfection reagent. The expression levels of Bach1 mRNA and protein were measured by qRT-PCR and western blot analysis, respectively. The survival of cells was determined using MTT assay cells, apoptosis using Tunel assay, Cell migration using scratch assay and Cell cycle analysis by Propidium Iodide (PI) DNA staining method by using flow cytometry on the MDA-MB-468. The expression levels of MMP-9 and CXCR4 were measured by qRT-PCR.
Transfection with siRNA significantly suppressed the expression of bach1 gene in dose dependent manner after 48h (p<0.0001). Surprisingly, treatment with bach1 siRNA arrest cell cycle in S phases (p<0.0001). Moreover siRNA transfection had effects on breast adenocarcinoma cells and inhibits the migration (p<0.0001), proliferation (p<0.0001), cell cycle arrest (p=0.03) and induces apoptosis (p<0.0001) and reduces the expression of miR-21 (P=0.0014).
Our results suggest that the bach1 can be considered as a potent adjuvant in breast cancer therapy.
尽管近年来临床和治疗技术有了很大进步，但许多晚期乳腺癌患者仍死于疾病的术后复发和转移。Bach1在肿瘤细胞的侵袭表型发展、细胞分裂和凋亡中发挥作用。本研究的目的是探讨特异性巴赫1小干扰RNA（siRNAs）对MDA-MB-468细胞（乳腺癌细胞）增殖、迁移、侵袭、凋亡诱导、细胞周期阻滞以及上皮-间质转化（EMT）相关微小RNA（miRNA）的影响。
使用转染试剂进行siRNA转染。分别通过定量逆转录聚合酶链反应（qRT-PCR）和蛋白质免疫印迹分析测定Bach1信使核糖核酸（mRNA）和蛋白质的表达水平。使用MTT法测定细胞存活率，用Tunel法检测细胞凋亡，用划痕试验检测细胞迁移，通过碘化丙啶（PI）DNA染色法并利用流式细胞术对MDA-MB-468细胞进行细胞周期分析。通过qRT-PCR测定基质金属蛋白酶-9（MMP-9）和趋化因子受体4（CXCR4）的表达水平。
转染siRNA后48小时，以剂量依赖方式显著抑制了巴赫1基因的表达（p<0.0001）。令人惊讶的是，用巴赫1 siRNA处理使细胞周期停滞在S期（p<0.0001）。此外，siRNA转染对乳腺腺癌细胞有影响，抑制迁移（p<0.0001）、增殖（p<0.0001）、细胞周期阻滞（p=0.03）并诱导凋亡（p<0.0001），还降低了miR-21的表达（P=0.0014）。
我们的结果表明，巴赫1可被视为乳腺癌治疗中的一种有效佐剂。
