Background: Cyclic RNA (circRNA) is a new type of non-coding RNA (ncRNA) which is different from traditional linear RNA. More and more studies suggest that circRNA can be used as a biological marker of many malignant tumors and becomes a potential target for treatment. Therefore, searching for new molecular targets of lung adenocarcinoma from the circRNA will help to reveal the new mechanism of the occurrence and development of lung adenocarcinoma, and provide new ideas for clinical diagnosis and treatment. In this study, the biological function of circ_0007766, a highly expressed circRNA found in a screen of lung adenocarcinoma tissue, was verified and analyzed in vitro, so as to preliminarily explore the mechanism of circ_0007766 in promoting the proliferation of lung adenocarcinoma.
Methods: The expression level of circ_0007766 in lung adenocarcinoma cells was detected by qPCR. Then siRNA was used to knock down the expression of circ_0007766. The effects of knockdown of circ_0007766 on proliferation, cell cycle and apoptosis of lung adenocarcinoma cells were detected by CCK8, scratch test, PI staining and Annexin V/PI double staining. In addition, the biological mechanism of circ_0007766 in lung adenocarcinoma was preliminarily studied by qPCR and Western blots.
Results: The expression of circ_0007766 in lung adenocarcinoma cell lines was detected by qPCR. The expression of circ_0007766 was interfered in SPCA-1 cells. The proliferation and migration abilities of cells were inhibited. The cell cycle was arrested in G0/G1 phase, but the apoptosis was not affected. The deletion of circ_0007766 did not affect the expression of ERBB2, but influenced the mRNA and protein expression of Cyclin D1/Cyclin E1/CDK4.
Conclusions: In vitro functional studies have shown that circ_0007766 may promote the proliferation and migration of lung adenocarcinoma cells. Further molecular mechanism studies have found that circ_0007766 can up-regulate the expression of Cyclin D1/Cyclin E1/CDK4, which are the key proteins of cell cycle, and thus promote the malignant proliferation of lung adenocarcinoma. From the perspective of circRNA, this study will provide new clues for the pathogenesis, development and prognosis of lung adenocarcinoma, and provide new target for clinical treatment.
【中文题目：环状RNA分子circ_0007766通过上调细胞周期相关蛋白Cyclin D1/CyclinE1/CDK4的表达促进肺腺癌细胞增殖】 【中文摘要：背景与目的 环状RNA（circular RNA, circRNA）是有别于传统线性RNA的一类新型非编码RNA（non-coding RNA, ncRNA），越来越多的研究提示circRNA可作为诸多恶性肿瘤的生物学标志物，并成为治疗的潜在靶标。因此从circRNA角度挖掘肺腺癌新型分子靶标，将有助于揭示肺腺癌发生发展新机制，并为临床诊疗提供新思路。本研究针对前期筛选出的一个肺腺癌组织高表达circRNA即circ_0007766进行体外生物学功能验证和分析，从而对circ_0007766促进肺腺癌增殖的相关机制进行初步探索。方法 首先通过qPCR检测考察肺腺癌细胞中circ_0007766的表达水平，然后采用siRNA干扰circ_0007766的表达，通过CCK8法、划痕修复、PI单染和AnnexinV/PI双染方法检测circ_0007766沉默表达对肺腺癌细胞的增殖、周期和凋亡作用的影响。此外，我们还采用qPCR和western blots方法初步研究了circ_0007766在肺腺癌细胞中的生物学作用机制。结果 qPCR检测表明肺腺癌细胞株中表达有circ_0007766，在SPCA-1 细胞中干扰circ_0007766表达，发现细胞增殖和迁移能力受到了抑制，细胞周期发生了G0/G1期阻滞，而细胞凋亡则不受影响。circ_0007766的缺失并不影响其亲本基因ERBB2的mRNA表达，而对eIF4A3调控的细胞周期相关基因Cyclin D1/Cyclin E1/CDK4的mRNA和蛋白水平均有影响。结论 本研究通过体外功能学研究，发现circ_0007766可能对肺腺癌细胞增殖、迁移具有促进的作用，进一步的分子机制研究发现circ_0007766可上调细胞周期关键蛋白Cyclin D1/Cyclin E1/CDK4的表达，进而促进肺腺癌恶性增殖。本研究以circRNA为视角，将对肺腺癌发生发展机理及预后判断提供新线索，为临床治疗应用提供新靶标。 】 【中文关键词：肺腺癌；环状RNA/circRNA；circ_0007766】.
Keywords: Lung adenocarcinoma; circRNA; circ_0007766.