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唐乃高,李鹏程,翁海勇,董慧,郑根建.甘草素抑制口腔鳞状细胞癌细胞增殖的机制研究[J].湖南中医药大学学报英文版,2022,42(11):1863-1869.[Click to copy
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| 甘草素抑制口腔鳞状细胞癌细胞增殖的机制研究 |
| 唐乃高,李鹏程,翁海勇,董慧,郑根建 |
| (海南医学院第一附属医院口腔科, 海南 海口 570100) |
| 摘要: |
| 目的 研究甘草素通过微RNA-486-3p（microRNA-486-3p, miR-486-3p）靶向盘蛋白结构域受体-1（discoidin domain receptor 1, DDR1）抑制口腔鳞状细胞癌及其作用机制。方法 采用不同浓度甘草素处理人口腔鳞状细胞癌SCC-15细胞48 h，MTT法检测甘草素对细胞生存率的影响。将细胞分为对照组、甘草素组、miR抑制剂组、病毒对照组、抑制慢病毒组和过表达慢病毒组，RT-PCR检测miR-486-3p和DDR1表达；5-乙炔基-2′脱氧尿嘧啶核苷（5-ethynyl-2′-deoxyuridine, EdU）染色检测细胞增殖情况；Western blot检测cleaved Caspase-3和Caspase-3表达。将细胞分为转染对照组和转染组，使用包含野生型或突变型DDR1 3'-UTR的双荧光素酶报告基因检测系统检测miR-486-3p和DDR1的结合情况。结果 与0 μmol/L甘草素比较，100、120、150 μmol/L甘草素能明显抑制SCC-15细胞的生存率（P＜0.05）。与对照组比较，甘草素组miR-486-3p的相对表达量、cleaved Caspase-3/Caspase-3相对比率均明显升高（P＜0.05），DDR1的相对表达量、EdU阳性细胞相对比值均明显降低（P＜0.05）。与甘草素组比较，miR抑制剂组miR-486-3p的相对表达量、cleaved Caspase-3/Caspase-3相对比率均明显降低（P＜0.05），DDR1的相对表达量、EdU阳性细胞相对比值均明显升高（P＜0.05）。与病毒对照组比较，抑制慢病毒组EdU阳性细胞相对比率明显降低（P＜0.05），cleaved Caspase-3/Caspase-3相对比率明显升高（P＜0.05）；过表达慢病毒组EdU阳性细胞相对比率明显升高（P＜0.05），cleaved Caspase-3/Caspase-3相对比率明显降低（P＜0.05）。miR-486-3p与DDR1存在结合位点。与转染对照组比较，转染组相对荧光素酶活力明显降低（P＜0.05），野生型DDR1 3'-UTR相对荧光素酶活力明显降低（P＜0.05）。结论 甘草素通过miR-486-3p靶向DDR1抑制SCC-15细胞增殖，促进细胞凋亡，进而发挥抗肿瘤作用。 |
| 关键词: 口腔癌 鳞状细胞癌 甘草素 microRNA-486-3p 盘蛋白结构域受体-1 增殖 凋亡 |
| DOI：10.3969/j.issn.1674-070X.2022.11.015 |
| Received:August 19, 2022 |
| 基金项目:国家自然科学基金地区科学基金项目（81760196）。 |
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| Mechanism of liquiritigenin inhibiting proliferation of oral squamous cell carcinoma |
| TANG Naigao,LI Pengcheng,WENG Haiyong,DONG Hui,ZHENG Genjian |
| (Department of Stomatology, The First Hospital of Hainan Medical College, Haikou, Hainan 570100, China) |
| Abstract: |
| Objective To study mechanism of liquiritigenin inhibiting oral squamous cell carcinoma based on targeting discoidin domain receptor 1 (DDR1) through microRNA-486-3p (miR-486-3p). Methods We adopted different concentrations of liquiritigenin to treat human oral squamous cell carcinoma (SCC-15) cells for 48 h, detected the effects of liquiritigenin on cell survival rates by MTT assay. Cells were divided into control group, liquiritigenin group, miR-486-3p inhibitor group, virus control group, inhibiting slow virus group and over-expression lentivirus group. The expression of miR-486-3p and DDR1 was detected by RT-PCR. Cell proliferation was detected by 5-ethynyl-2'-deoxyuridine (EdU) staining. The expression of cleaved Caspase-3 and Caspase-3 was detected by Western blot. Cells were divided into transfection control group and transfection group. The combined condition of miR-486-3p and DDR1 was detected by double luciferase reporter gene detection system containing wildtype or mutant DDR1 3'-UTR. Results Compared with 0 μmol/L liquiritigenin, MTT assay showed that 100, 120, 150 μmol/L liquiritigenin could significantly inhibit the survival rates of SCC-15 cells (P＜0.05). Compared with control group, RT-PCR results showed that liquiritigenin significantly increased the relative expression of miR-486-3p in SCC-15 cells. The relative rates of cleaved Caspase-3/Caspase-3 (P＜0.05), the relative expression of DDR1 and the relative ratio of EdU positive neurons all significantly decreased (P＜0.05). Compared with liquiritigenin group, the assay results showed that the relative expression of miR-486-3p in SCC-15 cells and the relative rates of cleaved Caspase-3/Caspase-3 significantly decreased, and the relative expression of DDR1 and the relative ratio of EdU positive neurons significantly increased (P＜0.05). Compared with virus control group, the relative rates of EdU positive neurons of the inhibiting slow virus group significantly decreased (P＜0.05), and the relative rates of cleaved Caspase-3/Caspase-3 significantly increased (P＜0.05). The relative rates of EdU positive cells in the over-expression lentivirus group significantly increased (P＜0.05), and the relative rates of cleaved Caspase-3/Caspase-3 significantly decreased (P＜0.05). There existed combination points sitting between miR-486-3p and DDR1. Compared with transfection control group, the relative luciferase activity of transfection group significantly decreased (P＜0.05), and the relative luciferase activity of wildtype DDR13'-UTR significantly decreased (P＜0.05). Conclusion Liquiritigenin can inhibit the proliferation of SCC-15 cells and promote the apoptosis process of SCC-15 by targeting DDR1 through miR-486-3p, which plays a role to suppress cancer. |
| Key words: oral carcinoma squamous cell carcinoma liquiritigenin microRNA-486-3p discoidin domain receptor 1 proliferation apoptosis |
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