Over-expression of Circular RNA-103809 Regulating Oxidative Stress and Immunity of Breast Cancer Cells

Abstract

Abstract: Objective To explore the regulation mechanism of over-expression of circular RNA-103809(circRNA-103809) on oxidative stress and immunity of breast cancer cells. Methods The pcDNA empty plasmid and circRNA-103809 over-expression plasmid were transfected into breast cancer cells MCF-7, which were included into negative control group and circ103809 group, respectively, and the blank control group was set up. The expression of circ103809 in each group was detected by real-time fluorescence quantitative polymerase chain reaction(RT-PCR). The cell growth was detected by clone formation assay. The oxidative stress markers [superoxide dismutase(SOD), malondialdehyde(MDA), glutathione(GSH)] were detected by biochemical kits. The changes of mitochondrial membrane potential(MMP) were detected by flow cytometry. The expressions of proliferation-related proteins(Ki67, p21), proliferating cell nuclear antigen (PCNA), B lymphoma-2 gene(Bcl-2), Bcl-2 associated X protein(Bax) and cell proliferation gene c-Myc were detected by Western blot. The levels of inflammatory factors (IL-6, IL-10) and inducible nitric oxide synthase(iNOS) were detected by enzyme-linked immunosorbent assay(ELISA) and qRT-PCR, and p65 content was detected by immunofluorescence. Results Compared with blank control group and negative control group, green fluorescence signals were enhanced in circ103809 group, relative expression levels of circ103809 and p21 protein, levels of GSH and MDA, Bax/Bcl-2 and IL-10 were significantly increased. Formation rate of cell colony, relative expression levels of Ki67, PCNA and c-Myc proteins, SOD, IL-6, iNOS and positive rate of nuclear p65 were significantly deceased(P<0.05). Conclusion Over-expression of circRNA-103809 can inhibit the proliferation of breast cancer cells MCF-7, which may be related to promoting oxidative stress level and alleviating inflammation response.

Key words: circular RNA-103809, breast cancer, oxidative stress, immune regulation, inflammatory response

摘要： 目的探究过表达环状RNA-103809（circRNA-103809）对乳腺癌细胞氧化应激和免疫的调节。方法将pcDNA空质粒、circRNA-103809过表达质粒转染乳腺癌细胞MCF-7,分别作阴性对照组和circ103809组,另设置空白对照组。qRT-PCR检测各组细胞中circ103809的表达,克隆形成实验检测细胞生长,生化试剂盒检测氧化应激标记物超氧化物歧化酶（SOD）、丙二醛（MDA）和谷胱甘肽（GSH）,流式细胞术检测线粒体膜电位的变化,Western blot检测细胞中增殖相关蛋白Ki67、p21、增殖细胞核抗原（PCNA）、B淋巴细胞瘤-2基因（Bcl-2）、Bcl-2相关X蛋白（Bax）、细胞增殖基因c-Myc的表达,酶联免疫吸附法（ELISA）和qRT-PCR检测炎症因子白细胞介素（IL-6、IL-10）和诱导型一氧化氮合酶（iNOS）,免疫荧光检测p65的含量。结果与空白对照组和阴性对照组相比,circ103809组细胞中绿色荧光信号增强,circ103809的相对表达量、p21蛋白相对表达量、GSH和MDA含量、Bax/Bcl-2水平、IL-10表达明显升高,细胞集落形成率、Ki67、PCNA、c-Myc蛋白相对表达量、SOD含量、IL-6、iNOS表达及p65入核阳性率明显降低（P<0.05）。结论过表达circRNA-103809可抑制乳腺癌细胞MCF-7的增殖,可能与促进氧化应激水平、减轻炎症反应有关。

关键词: 环状RNA-103809, 乳腺癌, 氧化应激, 免疫调节, 炎症反应

CLC Number:

R737.9

LIU A-min, ZHAO Wen-bin, CHEN Bao-ping. Over-expression of Circular RNA-103809 Regulating Oxidative Stress and Immunity of Breast Cancer Cells[J]. Journal of Chengde Medical University, 2022, 39(6): 455-460.

刘阿敏, 赵文斌, 陈保平. 过表达环状RNA-103809调节乳腺癌细胞氧化应激和免疫[J]. 承德医学院学报, 2022, 39(6): 455-460.

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