长链非编码 RNA DUXAP8 的下调通过 TRIM44 介导的 AKT/mTOR 通路调控 miR-498 抑制非小细胞肺癌的增殖、转移和 EMT。
Down-regulation of long non-coding RNA DUXAP8 suppresses proliferation, metastasis and EMT by modulating miR-498 through TRIM44-mediated AKT/mTOR pathway in non-small-cell lung cancer.
机构信息
Department of Respiratory and Critical Care Medicine, Lin Yi People's Hospital, Lin Yi, Shandong, China.
出版信息
Eur Rev Med Pharmacol Sci. 2020 Mar;24(6):3152-3165. doi: 10.26355/eurrev_202003_20682.
OBJECTIVE
The long non-coding RNA double homeobox A pseudogene 8 (DUXAP8) was reported to be involved in the initiation and development of multiple cancers. However, the detailed biological role of DUXAP8 in non-small-cell lung cancer (NSCLC) remains unclear. Herein, we aimed to explore the biological function and molecular mechanism of DUXAP8 in NSCLC.
PATIENTS AND METHODS
The levels of DUXAP8, microRNA-498 (miR-498) and tripartite motif-44 (TRIM44) were detected by Quantitative Real-time polymerase chain reaction (qRT-PCR). The cell proliferation, migration and invasion were detected by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and transwell assays. Protein expression levels were detected by Western blot. The target relationships among DUXAP8, miR-498 and TRIM44 were predicted by starBase2.0 and confirmed using luciferase reporter and RNA pull-down assays. To detect the role of DUXAP8 in vivo, tumor xenografts were created.
RESULTS
DUXAP8 and TRIM44 were upregulated in NSCLC tissues and cell lines, while miR-498 was downregulated. Functionally, knockdown of DUXAP8 could repress proliferation, migration, invasion, Epithelial-Mesenchymal Transition (EMT) and phosphorylation of AKT/mTOR in NSCLC cells. This inhibition could be restored by inhibiting miR-498 or overexpressing TRIM44. Furthermore, we also observed a positive correlation between DUXAP8 and TRIM44 expression, while the expressions of miR-498 and DUXAP8, as well as miR-498 and TRIM44, were negatively correlated in NSCLC tissues. Importantly, DUXAP8 could regulate the expression of TRIM44 via miR-498. Moreover, knockdown of DUXAP8 notably decreased the xenograft tumor volume, weight and number of metastatic nodules in vivo.
CONCLUSIONS
Our results identified that LncRNA DUXAP8 could regulate cell proliferation, metastasis and EMT in NSCLC cells by inhibiting miR-498 through the activation of TRIM44-mediated AKT/mTOR pathway.
目的
长链非编码 RNA 双同源盒 A 假基因 8（DUXAP8）被报道参与多种癌症的发生和发展。然而，DUXAP8 在非小细胞肺癌（NSCLC）中的详细生物学作用仍不清楚。本研究旨在探讨 DUXAP8 在 NSCLC 中的生物学功能和分子机制。
方法
采用实时定量聚合酶链反应（qRT-PCR）检测 DUXAP8、微小 RNA-498（miR-498）和三肽基 44（TRIM44）的水平。通过 3-（4,5-二甲基噻唑-2-基）-2,5-二苯基四唑溴盐（MTT）和 Transwell 检测细胞增殖、迁移和侵袭。通过 Western blot 检测蛋白表达水平。通过 starBase2.0 预测 DUXAP8、miR-498 和 TRIM44 之间的靶关系，并通过荧光素酶报告和 RNA 下拉实验进行验证。为了检测 DUXAP8 在体内的作用，创建了肿瘤异种移植模型。
结果
在 NSCLC 组织和细胞系中，DUXAP8 和 TRIM44 上调，而 miR-498 下调。功能上，敲低 DUXAP8 可抑制 NSCLC 细胞的增殖、迁移、侵袭、上皮间质转化（EMT）和 AKT/mTOR 的磷酸化。这种抑制作用可以通过抑制 miR-498 或过表达 TRIM44 来恢复。此外，我们还观察到 NSCLC 组织中 DUXAP8 和 TRIM44 表达之间呈正相关，而 miR-498 和 DUXAP8 以及 miR-498 和 TRIM44 表达之间呈负相关。重要的是，DUXAP8 可以通过 miR-498 调节 TRIM44 的表达。此外，体内敲低 DUXAP8 显著减少了异种移植肿瘤的体积、重量和转移结节数。
结论
本研究结果表明，LncRNA DUXAP8 通过激活 TRIM44 介导的 AKT/mTOR 通路抑制 miR-498 的表达，从而调节 NSCLC 细胞的增殖、转移和 EMT。
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