Wang Wei, Ma Jun, Lu Jianjun, Fang Danqing, Xiong Xinming, Yang Xin, Xie Tingting
a Department of Thoracic Surgery, the Second Affiliated Hospital of Guangzhou Medical University , Guang Zhou , China.
b Department of Thoracic Surgery, the First Affiliated Hospital of Sun Yat-Sen University , Guangzhou , China.
Cancer Biol Ther. 2018;19(12):1117-1127. doi: 10.1080/15384047.2018.1480889. Epub 2018 Sep 25.
: The study aimed to investigate candidate circular RNAs (circRNAs) in regulating the pathogenic process of esophageal carcinoma. : Specimens were collected from the patients with esophageal carcinoma. Total RNA was purified and treated with RNase R followed by RNA-seq in the purpose of screening the circRNAs in significant differentially expression. The expression level of the screened circRNAs were further validated using RT-PCR. The circular structure of the circRNA was validated with divergent and convergent primers. Overexpression vector was prepared in the purpose of raising the expression level of circ0043898 in the ECA-109 and Kyse-520 cells. The cell colony assay and MTS assay were conducted to determine the capacity of cell proliferation. Chamber assays were applied to determine the capacity of cell migration and invasion while flowcytometry was applied to determine the cell cycle and cell apoptosis. In vivo animal assay was conducted by injecting the cells to the chest of the mice. RNA-seq was performed followed by GO and KEGG study to further verify the regulation mechanism of circ0043898. : circ0043898 was validated that down-regulated expressed in the specimens from the patients with esophageal carcinoma. The cell assays proved that overexpression of circ0043898 can obviously inhibit the cell proliferation, cell migration and invasion and induce cell apoptosis and death in the cancerous cells. The in vivo animal study also suggested that the circ0043898 performed inhibitory functions on oncogenesis. The RNA-seq presented the potential regulation mechanism of circ0043898. Histone H3 and BMI1 were presented significantly differential expression in both ECA-109 and Kyse-520 cells, indicating they might be the targets of circ0043898. : circ0043898 is presented as tumor inhibitor and could be a candidate biomarker in the therapeutic target and diagnosis of esophageal carcinoma.
该研究旨在探究候选环状RNA(circRNA)在食管癌致病过程中的调控作用。收集食管癌患者的标本。纯化总RNA并用RNase R处理,随后进行RNA测序,以筛选出具有显著差异表达的circRNA。使用RT-PCR进一步验证筛选出的circRNA的表达水平。用发散引物和收敛引物验证circRNA的环状结构。制备过表达载体以提高ECA-109和Kyse-520细胞中circ0043898的表达水平。进行细胞集落测定和MTS测定以确定细胞增殖能力。采用小室测定法确定细胞迁移和侵袭能力,同时采用流式细胞术确定细胞周期和细胞凋亡。通过将细胞注射到小鼠胸部进行体内动物实验。进行RNA测序,随后进行基因本体(GO)和京都基因与基因组百科全书(KEGG)研究,以进一步验证circ0043898的调控机制。circ0043898在食管癌患者标本中被证实表达下调。细胞实验证明,circ0043898的过表达可明显抑制癌细胞的增殖、迁移和侵袭,并诱导细胞凋亡和死亡。体内动物研究也表明,circ0043898对肿瘤发生具有抑制作用。RNA测序揭示了circ0043898的潜在调控机制。组蛋白H3和BMI1在ECA-109和Kyse-520细胞中均呈现显著差异表达,表明它们可能是circ0043898的靶点。circ0043898表现为肿瘤抑制因子,可能成为食管癌治疗靶点和诊断中的候选生物标志物。