circ-SFMBT2通过靶向miR-7-5p/ADAM10轴对非小细胞肺癌细胞生物学行为的影响

ffect of circ-SFMBT2 on the biological behavior of non-small cell lung cancer cells by targeting the miR-7-5p/ADAM10 axis

目的探讨circ-SFMBT2对非小细胞肺癌(non-small cell lung cancer,NSCLC)细胞生物学行为的影响以及对miR-7-5p/ADAM10分子轴的调控作用。方法﹑采用qRT-PCR法与Western印迹法分别检测NSCLC与癌旁组织中circ-SFMBT2、miR-7-5p、ADAM10的表达量;用Pearson法分析circ-SFMBT2与miR-7-5p,以及miR-7-5p与ADAM10的相关性;体外培养人支气管上皮样细胞(human bronchial epithelial-like cells,HBE)与肺癌细胞系H1650、H460、A549、H1299。用CCK-8与EdU实验检测细胞的增殖能力。用平板克隆形成实验检测细胞的克隆形成能力。用流式细胞术检测细胞凋亡率。用Transwell小室实验检测细胞侵袭。用双荧光素酶报告实验检测circ-SFMBT2与miR-7-5p、以及miR-7-5p与ADAM10的靶向关系。用裸鼠移植瘤实验检测敲低circ-SFMBT2对移植瘤生长的影响。用免疫组化实验检测移植瘤组织中ADAM10与Ki67蛋白阳性率。结果circ-SFMBT2与ADAM10在NSCLC组织及细胞系中表达升高,而miR-7-5p的表达降低,circ-SFMBT2与miR-7-5p的表达呈负相关,而miR-7-5p与ADAM10的表达呈负相关。沉默circ-SFMBT2及miR-7-5p过表达可抑制细胞增殖、克隆形成及侵袭,还可促进其凋亡。circ-SFMBT2可靶向调控miR-7-5p,而ADAM10是miR-7-5p的靶基因。沉默circ-SFMBT2与抑制miR-7-5p联合作用,以及miR-7-5p过表达与ADAM10过表达联合作用均可促进细胞增殖、克隆形成及侵袭,并抑制其凋亡。沉默circ-SFMBT2可抑制移植瘤的生长。结论︰沉默circ-SFMBT2可通过调控miR-7-5p/ADAM10分子轴而减弱NSCLC细胞增殖、克隆形成,侵袭能力并诱导其凋亡。

Objective To explore the effect of circ-SFMBT2 on the biological behavior of non-smallcell lung cancer (NSCLC) cells and its regulatory role on the miR-7-5p/ADAM10 axis,MethodsqRT-PCRand Western blotting were used to determine the expression of circ-SFMBT2,miR-7-5p,and ADAM10 inNSCLC tissues and adjacent tissues. Pearson analysis was used to analyze the correlation between circ-SFMBT2 and miR-7-5p, and between miR-7-5p and ADAM10. In vitro cultured human bronchial epithelial-like ceils(HBE) and lung cancer cell lines H1650,H460,A549,H1299.CCK-8 and EdU methods wereused to assess the ability of cell proliferation.Plate experiment was used to detect the clone formationability. Flow cytometry was used to detect the apoptosis rate. Transwell experiment was used to detect cellinvasion ability.Dual luciferase reporter experiment detects the targeting relationship between circ-SFMBT2and miR-7-5p,and between miR-7-5p and ADAM10.Transplanted tumor experiment in nude mice assessedthe effect of knocking down circ-SFMBT2 on the growth of transplanted tumor. Immunohistochemicalexperiments were performed to detect the positive rates of ADAM10 and Ki67 proteins in transplanted tumortissues. Results The expression levels of circ-SFMBT2 and ADAM10 were increased in NSCLC tissues andcel lines,while decreased the expression of miR-7-5p. circ-SFMBT2 was negatively correlated with miR-7-5p,while miR-7-5p was negatively correlated with ADAM10、Silencing the overexpression of circ-SFMBT2and miR-7-5p could inhibit cell proliferation,clone formation and invasion,and also promote apoptosis. circ-SFMBT2 could target miR-7-5p,and ADAM10 was the target gene of miR-7-5p.The combined effect ofsilencing circ-SFMBT2 and inhibition of miR-7-5p,as well as miR-7-5p overexpression and ADAM10overexpression could promote cell proliferation,clone formation and invasion,and also suppress cellapoptosis,Silencing circ-SFMBT2 could inhibit the growth of transplanted tumors. Conclusion Silencingcirc-SFMBT2 can suppress the proliferation,clone formation,invasion ability and induce apoptosis ofNSCLC cells by regulating the miR-7-5p/ADAM10 axis.