Wang Qiushi, Yan Chunhua, Zhang Pengfei, Li Guanghua, Zhu Ruidong, Wang Hanbing, Wu Libo, Xu Guangquan
The Second Department of General Thoracic Surgery, The Second Affiliated Hospital of Harbin Medical University, Harbin, China.
Department of Respiratory, Longgang District People's Hospital of Shenzhen, Shenzhen, China.
Front Oncol. 2021 Nov 11;11:757541. doi: 10.3389/fonc.2021.757541. eCollection 2021.
This study aimed to explore the circular RNA (circRNA/circ) profile engaged in non-small cell lung cancer (NSCLC) development and metastasis and to investigate potentially key carcinogenic circRNAs related to NSCLC.
CircRNA profiles between 10 NSCLC tissues and 10 adjacent tissues and between five NSCLC tissues with lymph node metastasis (LNM) and five NSCLC tissues without LNM were detected by Arraystar Human circRNA Array followed by bioinformatics. Circ_0008594 knockdown, circ_0004293 overexpression, and circ_0003832 overexpression plasmids were transfected into H23 and H460 cells to sort potential oncogenic circRNA. Then circ_0008594 overexpression and knockdown plasmids were transfected, followed by that circ_0008594 knockdown plus miR-760 knockdown plasmids were transfected into these cells. Cell proliferation, apoptosis, invasion, stemness, and pathways were detected. In addition, xenograft mice models were constructed injecting H23 cells with circ_0008594 overexpression or knockdown to validate the findings.
A total of 455 dysregulated circRNAs in NSCLC tissues adjacent tissues and 353 dysregulated circRNAs in NSCLC tissues with LNM those without LNM were discovered. cross-analysis, 19 accordant circRNAs were uncovered, among which three candidate circRNAs (circ_0008594, circ_0004293, circ_0003832) were chosen for functional experiments, during which it was observed that circ_0008549 affected H23 and H460 cell proliferation and apoptosis more obviously than circ_0004293 and circ_0003832. Subsequent experiments showed that circ_0008594 promoted H23 and H460 cell proliferation and invasion but affected stemness less and negatively regulated miR-760 direct binding. Furthermore, miR-760 attenuated the effect of circ_0008549 on regulating H23 and H460 cell functions and the PI3K/AKT and MEK/ERK pathways. experiments further confirmed that circ_0008549 increased tumor volume, epithelial-mesenchymal transition, and the PI3K/AKT and MEK/ERK pathways while reducing tumor apoptosis and miR-760 NSCLC xenograft models.
Our study identifies several valuable circRNAs related to NSCLC development and LNM. Furthermore, as a key functional circRNA, circ_0008594 was observed to promote NSCLC progression by regulating the miR-760-mediated PI3K/AKT and MEK/ERK pathways.
本研究旨在探索参与非小细胞肺癌（NSCLC）发生和转移的环状RNA（circRNA）图谱，并研究与NSCLC相关的潜在关键致癌circRNA。
采用Arraystar人类circRNA芯片检测10例NSCLC组织与10例癌旁组织之间以及5例有淋巴结转移（LNM）的NSCLC组织与5例无LNM的NSCLC组织之间的circRNA图谱，随后进行生物信息学分析。将circ_0008594敲低、circ_0004293过表达和circ_0003832过表达质粒转染至H23和H460细胞中，以筛选潜在的致癌circRNA。然后转染circ_0008594过表达和敲低质粒，接着将circ_0008594敲低加miR-760敲低质粒转染至这些细胞中。检测细胞增殖、凋亡、侵袭、干性及相关通路。此外，构建异种移植小鼠模型，注射过表达或敲低circ_0008594的H23细胞以验证研究结果。
在NSCLC组织与癌旁组织中发现共455个差异表达的circRNA，在有LNM的NSCLC组织与无LNM的NSCLC组织中发现353个差异表达的circRNA。通过交叉分析，发现19个一致的circRNA，其中选择3个候选circRNA（circ_0008594、circ_0004293、circ_0003832）进行功能实验，在此过程中观察到circ_0008549对H23和H460细胞增殖和凋亡的影响比circ_0004293和circ_0003832更明显。后续实验表明，circ_0008594促进H23和H460细胞增殖和侵袭，但对干性影响较小，并负向调节miR-760的直接结合。此外，miR-760减弱了circ_0008549对H23和H460细胞功能及PI3K/AKT和MEK/ERK通路的调节作用。体内实验进一步证实，circ_0008549增加了NSCLC异种移植模型的肿瘤体积、上皮-间质转化以及PI3K/AKT和MEK/ERK通路的活性，同时减少肿瘤凋亡和miR-760表达。
我们的研究鉴定了几种与NSCLC发生和LNM相关的有价值的circRNA。此外，作为一种关键的功能性circRNA，circ_0008594被观察到通过调节miR-760介导的PI3K/AKT和MEK/ERK通路促进NSCLC进展。
