Shi Hui, Pu Jin, Zhou Xiao-Li, Ning Yun-Ye, Bai Chong
1 Department of Respiratory and Critical Care Medicine, Changhai Hospital, Second Military Medical University, Shanghai, P.R. China.
2 Department of Special Clinic, Changhai Hospital, Second Military Medical University, Shanghai, P.R. China.
Tumour Biol. 2017 May;39(5):1010428317697568. doi: 10.1177/1010428317697568.
This study aimed to investigate the effects of long non-coding RNA ROR (regulator of reprogramming) on cisplatin (DDP) resistance in patients with non-small-cell lung cancer by regulating PI3K/Akt/mTOR signaling pathway. Human cisplatin-resistant A549/DDP cell lines were selected and divided into control group, negative control group, si-ROR group, ROR over-expression group, Wortmannin group, and ROR over-expression + Wortmannin group. MTT assay was used to determine the optimum inhibitory concentration of DDP. Quantitative real-time polymerase chain reaction and western blotting were applied to detect expressions of long non-coding RNA ROR, PI3K, Akt, and mTOR. Colony-forming assay, scratch test, Transwell assay, and flow cytometry were conducted to detect cell proliferation, migration, invasion, and apoptosis, respectively. Tumor-formation assay was performed to detect the growth of transplanted tumors. Long non-coding RNA ROR expression was high in human A549/DDP cell lines. Compared with the control and negative control groups, the mRNA and protein expressions of PI3K, Akt, mTOR, and bcl-2 decreased, whereas the mRNA and protein expression of bax and the sensitivity of cells to DDP significantly increased. Cell proliferation, migration, and invasion abilities decreased in the si-ROR and Wortmannin groups. In comparison with control and negative control groups, the mRNA and protein expressions of PI3K, Akt, mTOR, and bcl-2 increased, whereas the mRNA and protein expressions of bax decreased, the sensitivity of cells to DDP significantly increased, and cell proliferation, migration, and invasion abilities decreased in the ROR over-expression group. For nude mice in tumor-formation assay, compared with control and negative control groups, the tumor weight was found to be lighter (1.03 ± 0.15) g, the protein expressions of PI3K, Akt, mTOR, and bcl-2 decreased, and the protein expression of bax increased in the si-ROR group. Long non-coding RNA ROR may affect the sensitivity of lung adenocarcinoma cells to DDP by targeting PI3K/Akt/mTOR signaling pathway.
本研究旨在通过调节PI3K/Akt/mTOR信号通路，探讨长链非编码RNA ROR（重编程调节因子）对非小细胞肺癌患者顺铂（DDP）耐药性的影响。选取人顺铂耐药A549/DDP细胞系，分为对照组、阴性对照组、si-ROR组、ROR过表达组、渥曼青霉素组和ROR过表达+渥曼青霉素组。采用MTT法测定DDP的最佳抑制浓度。应用定量实时聚合酶链反应和蛋白质印迹法检测长链非编码RNA ROR、PI3K、Akt和mTOR的表达。分别进行集落形成试验、划痕试验、Transwell试验和流式细胞术检测细胞增殖、迁移、侵袭和凋亡情况。进行肿瘤形成试验检测移植瘤的生长情况。长链非编码RNA ROR在人A549/DDP细胞系中表达较高。与对照组和阴性对照组相比，PI3K、Akt、mTOR和bcl-2的mRNA和蛋白表达降低，而bax的mRNA和蛋白表达以及细胞对DDP的敏感性显著增加。si-ROR组和渥曼青霉素组细胞增殖、迁移和侵袭能力降低。与对照组和阴性对照组相比，ROR过表达组PI3K、Akt、mTOR和bcl-2的mRNA和蛋白表达增加，而bax的mRNA和蛋白表达降低，细胞对DDP的敏感性显著增加，细胞增殖、迁移和侵袭能力降低。在肿瘤形成试验的裸鼠中，与对照组和阴性对照组相比，si-ROR组肿瘤重量较轻（1.03±0.15）g，PI3K、Akt、mTOR和bcl-2的蛋白表达降低，bax的蛋白表达增加。长链非编码RNA ROR可能通过靶向PI3K/Akt/mTOR信号通路影响肺腺癌细胞对DDP的敏感性。
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