BACKGROUND:Brain metastasis (BM) is associated with poor prognosis in patients with non-small cell lung cancer (NSCLC). Recent studies demonstrated that microRNA-330-3p (miR-330-3p) was involved in NSCLC brain metastasis (BM). However, the exact parts played by miR-330-3p in BM of NSCLC remain unknown. Discovery and development of biomarkers and elucidation of the mechanism underlying BM in NSCLC is critical for effective prophylactic interventions. Here, we evaluated the expression and biological effects of miR-330-3p in NSCLC cells and explored the underlying mechanism of miR-330-3p in promoting cell migration and invasion in NSCLC. METHODS:Stable over-expression and knockdown of miR-330-3p in NSCLC cells was constructed with lentivirus. Expression levels of miR-330-3p in NSCLC cells were quantified by quantitive real-time PCR (qRT-PCR). The effects of miR-330-3p on NSCLC cells were investigated using assays of cell viability, migration, invasion, cell cycle, apoptosis, western blotting, immunohistochemical, and immunofluorescence staining. A xenograft nude mouse model and in situ brain metastasis model were used to observe tumor growth and brain metastasis. The potential target of miR-330-3p in NSCLC cells was explored using the luciferase reporter assay, qRT-PCR, and western blotting. The miR-330-3p targets were identified using bioinformatics analysis and verified by luciferase reporter assay. The correlation between GRIA3 and DNA methyltransferase (DNMT) 1 and DNMT3A was tested by RT-PCR, western blotting, and co-immunoprecipitation (IP). RESULTS:miR-330-3p was significantly up-regulated in NSCLC cell lines. MTT assay, transwell migration, and invasion assays showed that miR-330-3p promoted the growth, migration, and invasion of NSCLC cells in vitro and induced tumor growth and metastasis in vivo. Luciferase reporter assays showed that GRIA3 was a target of miR-330-3p. qRT-PCR and western blotting exhibited that miR-330-3p promoted the growth, invasion, and migration of NSCLC cells by activating mitogen-activated protein kinase (MAPK)/extracellular-regulated protein kinases (ERK) signaling pathway. Furthermore, miR-330-3p up-regulated the total DNA methylation in NSCLC cells, and co-IP-demonstrated GRIA3 was directly related with DNMT1 and DNMT3A. CONCLUSIONS:miR-330-3p promoted the progression of NSCLC and might be a potential target for the further research of NSCLC brain metastasis.

译文

背景:脑转移(BM)与非小细胞肺癌(NSCLC)患者预后差有关。最近的研究表明,microRNA-330-3p(miR-330-3p)参与了NSCLC脑转移(BM)。但是,miR-330-3p在NSCLC的BM中所起的确切作用仍然未知。发现和开发生物标志物以及阐明NSCLC中BM的潜在机制对于有效的预防性干预至关重要。在这里,我们评估了miR-330-3p在NSCLC细胞中的表达和生物学效应,并探讨了miR-330-3p在促进NSCLC细胞迁移和侵袭中的潜在机制。
方法:用慢病毒构建稳定表达miR-330-3p的NSCLC细胞。通过定量实时PCR(qRT-PCR)定量检测NSCLC细胞中miR-330-3p的表达水平。使用细胞活力,迁移,侵袭,细胞周期,凋亡,蛋白质印迹,免疫组织化学和免疫荧光染色的方法研究了miR-330-3p对NSCLC细胞的影响。用异种移植裸鼠模型和原位脑转移模型观察肿瘤的生长和脑转移。使用荧光素酶报告基因检测,qRT-PCR和Western印迹探索了NSCLC细胞中miR-330-3p的潜在靶标。使用生物信息学分析鉴定了miR-330-3p靶标,并通过萤光素酶报告基因检测法对其进行了验证。通过RT-PCR,蛋白质印迹和免疫共沉淀(IP)测试了GRIA3与DNA甲基转移酶(DNMT)1和DNMT3A之间的相关性。
结果:miR-330-3p在NSCLC细胞系中显着上调。 MTT测定,穿孔迁移和侵袭测定显示,miR-330-3p在体外促进NSCLC细胞的生长,迁移和侵袭,并在体内诱导肿瘤生长和转移。萤光素酶报告基因检测表明GRIA3是miR-330-3p的靶标。 qRT-PCR和western blotting显示,miR-330-3p通过激活有丝分裂原激活的蛋白激酶(MAPK)/细胞外调节的蛋白激酶(ERK)信号通路,促进了NSCLC细胞的生长,侵袭和迁移。此外,miR-330-3p上调了NSCLC细胞中的总DNA甲基化,并且共同IP展示的GRIA3与DNMT1和DNMT3A直接相关。
结论:miR-330-3p促进了非小细胞肺癌的发展,可能是进一步研究非小细胞肺癌脑转移的潜在靶标。

+1
+2
100研值 100研值 ¥99课程
检索文献一次
下载文献一次

去下载>

成功解锁2个技能,为你点赞

《SCI写作十大必备语法》
解决你的SCI语法难题!

技能熟练度+1

视频课《玩转文献检索》
让你成为检索达人!

恭喜完成新手挑战

手机微信扫一扫,添加好友领取

免费领《Endnote文献管理工具+教程》

微信扫码, 免费领取

手机登录

获取验证码
登录