Background: MicroRNAs (miRNAs) are non-coding small molecule RNAs that are widely found in eukaryotic organisms, although some miRNAs have been found in tumors, the expression and effects of miR-665 on small cell lung cancer (SCLC) are unclear. The aim of this study was to analyze the effects of miR-665 on proliferation, cycle, invasion and migration of SCLC cells, and to explore the role of miR-665 in SCLC and its working mechanism.
Methods: The expression of miR-665 in SCLC tissues and adjacent normal tissues was detected by qRT-PCR. TargetScan predicted potential target genes for miR-665 and validated with dual luciferase reporter assays, qRT-PCR and Western blot. CCK8 assay, flow cytometry, Transwell and wound healing assay to detect the effects of miR-665 and LLGL1 on proliferation, invasion, migration and S-phase fraction of SCLC cell line NCI-H446, NCI-H1688. A nude mouse xenograft model of SCLC was constructed and the effect of miR-665 on tumor growth in mice was observed.
Results: The expression of miR-665 in SCLC tissues was significantly higher than that in non-tumor normal tissues. MiR-665 could target 3'-UTR of LLGL1 and inhibit its expression. Compared with non-tumor normal tissues, the expression of LLGL1 was significantly lower in SCLC tissues. Inhibition of miR-665 expression could inhibit proliferation, S-phase fraction, invasion and migration ability of SCLC NCL-H446 cells, and interference LLGL1 expression could reverse this inhibition effect. Up-regulation of miR-665 expression could promoted proliferation, S-phase fraction, invasion and migration ability of SCLC NCI-H1688 cells, but this promotion effect was also reversed by overexpression of LLGL1. In a nude mouse xenograft model of SCLC, inhibition of miR-665 expression could up-regulate LLGL1 protein expression and inhibit tumor growth, while up-regulation of miR-665 expression could produce opposite results.
Conclusions: The expression of miR-665 is closely related to SCLC. miR-665 can promote the biological behavior of SCLC cells by inhibiting the expression of target gene LLGL1, and miR-665 play a role in tumor-promoting genes in SCLC.
【中文题目：miR-665通过靶向调控LLGL1促进小细胞肺癌生物学行为的研究】 【中文摘要：背景与目的 MicroRNAs（miRNAs）是一种广泛存在于真核生物体中的非编码小分子RNA，尽管一些miRNA在肿瘤中作用已被发现，但是miR-665对小细胞肺癌的中的表达及影响还尚不清楚。本研究旨在分析miR-665对肺癌细胞增殖、周期、侵袭和迁移的影响，探讨miR-665在小细胞肺癌中发挥的作用及其工作机制。方法 qRT-PCR检测miR-665在肺癌组织和癌旁正常组织中的表达水平；TargetScan预测miR-665的潜在靶基因并用双荧光素酶报告基因实验、qRT-PCR和Western blot进行验证；免疫组化、qRT-PCR和Western blot检测LLGL1在肺癌组织和癌旁正常组织中的表达水平；CCK8法、流式细胞法、Transwell和细胞划痕实验检测miR-665和LLGL1对肺癌细胞NCI-H446、NCI-H1688增殖、侵袭、迁移以及S期细胞比值的影响；构建肺癌裸鼠移植瘤模型并观察miR-665对小鼠肿瘤生长的影响。结果 miR-665在肺癌组织中的表达水平明显高于癌旁正常组织；miR-665能靶向作用于LLGL1的3’-UTR并抑制其表达；相比于癌旁正常组织，LLGL1在肺癌组织中的表达水平明显降低；抑制miR-665的表达可以抑制肺癌NCI-H446细胞的增殖、S期细胞比值、侵袭和迁移能力，而干扰LLGL1能逆转这种抑制效果；上调miR-665则促进肺癌NCI-H1688的增殖、S期细胞比值、侵袭和迁移能力，但这种促进效果同样被LLGL1的过表达逆转；在肺癌裸鼠移植瘤模型中，抑制miR-665能上调LLGL1蛋白的表达并抑制肿瘤的生长，而上调miR-665的表达则可以产生相反的结果。结论 miR-665表达水平的变化与肺癌密切相关，miR-665可以通过抑制其靶基因LLGL1的表达促进肺癌细胞的生物学行为，在小细胞肺癌中发挥促癌基因的作用。】 【中文关键词：小细胞肺癌；miR-665；LLGL1】.
Keywords: LLGL1; Small cell lung cancer; miR-665.