Yang Xiaohong, Luo En, Liu Xin, Han Bin, Yu Xiaoping, Peng Xiaoli
Department of Public Health, Chengdu Medical College, Chengdu, China.
Department of General Surgery, The Fifth People's Hospital of Chengdu, Chengdu, China.
BMC Cancer. 2016 Jul 7;16:423. doi: 10.1186/s12885-016-2465-0.
The long non-coding RNA (lncRNA) HOX transcript antisense RNA (HOTAIR) plays a crucial role in cancer progression, which is regulated by the interferon regulatory factor-1 (IRF1) and up-streaming Akt activation. The present study evaluated the chemopreventive effects of delphinidin-3-glucoside (Dp), a major anthocyanin present in pigmented fruits and vegetables, on breast carcinogenesis, and investigate the role of the Akt/HOTAIR signaling pathway.
Human breast epithelial cells MCF10A were treated with carcinogens (NNK and B[a]P) or co-treated with carcinogens plus Dp for 30 days. Then, the cancer-associated properties of the treated cells were evaluated to assess the carcinogenesis and the effects of Dp. HOTAIR levels were detected by qRT-PCR. The proteins expression was measured by western blots, immunofluorescence and immunohistochemistry. Xenografted tumors were made by implanting breast cancer cells MDA-MB-231-Luc-GFP in athymic mice. ChIP-qPCR analysis was used to detect the IRF1 binding to the HOTAIR promoter.
Carcinogens treatment induces apparent carcinogenic transformation in MCF10A cells including reduced dependence on growth factors, anchorage-independent cell growth and aberrant wound-healing ability, which is effectively suppressed by Dp co-treatment. The level of HOTAIR significantly increases in a time-dependent manner during chronic breast carcinogenesis. Dp treatment down-regulates HOTAIR expression in breast carcinogenesis and breast cancer cells. Furthermore, Dp administration inhibits the growth of xenografted breast tumors in athymic mice, and decreases HOTAIR in vivo. Further studies showed that Dp represses Akt activation, promotes IRF1 expression and increases IRF1 binding to the HOTAIR promoter. Silence of IRF1 expression via transfecting cells with IRF1 siRNAs significantly reduced the effects of Dp on HOTAIR, resulting in decreased cytotoxic effects of Dp on breast cancer cells.
These data suggest the effective chemopreventive effect of Dp on breast carcinogenesis, in which down-regulation of HOTAIR plays a critical role.
长链非编码RNA（lncRNA）HOX转录本反义RNA（HOTAIR）在癌症进展中起关键作用，其受干扰素调节因子-1（IRF1）和上游Akt激活调控。本研究评估了矢车菊素-3-葡萄糖苷（Dp）（一种存在于有色水果和蔬菜中的主要花青素）对乳腺癌发生的化学预防作用，并探究Akt/HOTAIR信号通路的作用。
人乳腺上皮细胞MCF10A用致癌物（NNK和苯并[a]芘）处理，或与致癌物加Dp共同处理30天。然后，评估处理后细胞的癌症相关特性，以评估致癌作用及Dp的影响。通过qRT-PCR检测HOTAIR水平。用蛋白质印迹、免疫荧光和免疫组织化学法检测蛋白质表达。通过将乳腺癌细胞MDA-MB-231-Luc-GFP植入无胸腺小鼠体内制备异种移植瘤。采用染色质免疫沉淀-定量聚合酶链反应（ChIP-qPCR）分析检测IRF1与HOTAIR启动子的结合。
致癌物处理诱导MCF10A细胞发生明显的致癌转化，包括对生长因子的依赖性降低、不依赖贴壁细胞生长和异常的伤口愈合能力，而Dp共同处理可有效抑制这些转化。在慢性乳腺癌发生过程中，HOTAIR水平以时间依赖性方式显著升高。Dp处理可下调乳腺癌发生过程及乳腺癌细胞中HOTAIR的表达。此外，给予Dp可抑制无胸腺小鼠异种移植乳腺肿瘤的生长，并在体内降低HOTAIR水平。进一步研究表明，Dp可抑制Akt激活，促进IRF1表达，并增加IRF1与HOTAIR启动子的结合。通过用IRF1小干扰RNA转染细胞使IRF1表达沉默，可显著降低Dp对HOTAIR的影响，导致Dp对乳腺癌细胞的细胞毒性作用降低。
这些数据表明Dp对乳腺癌发生具有有效的化学预防作用，其中HOTAIR的下调起关键作用。
