Guizhou Provincial People Hospital
目的：分析长链非编码RNA SNHG16（Long non-coding RNA small nucleolar RNA host gene16，lncRNA SNHG16）通过调控微小RNA-570（miR-570）对肝癌细胞索拉非尼耐药的机制研究。方法：采用实时荧光RT-PCR检测人体正常肝脏组织、肝癌细胞组织中HepG2、HepG2-R细胞的lncRNA SNHG16、miR-570表达，用MTT法、流式细胞仪、Transwell试验检测细胞增殖、凋亡及侵袭，Western Blot法测定细胞CyclinD1、P21MMP-9、MMP-2表达变化。结果：与正常的肝细胞相比，肝癌细胞中lncRNA SNHG16表达升高，miR-570表达下降（P<0.05）。与HepG2-P组相比，HepG2-R组LncRNA SNHG16、IC50值表达上升，miR-570、HepG2-R细胞在索拉非尼浓度为1、2、4、8、16μmol/L中的抑制率水平下降（P<0.05）。HepG2-R pcDNA SNHG16作为过表达组，其lncRNA SNHG16表达明显升高（P<0.05），与HepG2-R pcDNA组对比，HepG2-R pcDNA SNHG16组的迁移的细胞个数、CyclinD1、P21、MMP-9、MMP-2表达减低，抑制率、凋亡率、P21表达上升（P<0.05）。与HepG2-R anti-miR-NC组相比，HepG2-R anti-miR-570组miR-570水平降低（P<0.05），与HepG2-R anti-miR-NC组比较，HepG2-R anti-miR-570组CyclinD1、MMP-9、MMP-2水平降低，抑制率、凋亡率、P21水平升高（P<0.05）。双荧光素酶报告实验显示，与miR-NC组相比，miR-570使WT-SNHG16荧光素酶活性减低（P<0.05），而对MUT-SNHG16荧光素酶活性影响较小（P>0.05）。过表达lncRNA SNHG16可使HepG2-R细胞中miR-570表达下降（P<0.05），与HepG2-R pcDNA SNHG16 miR-NC组相比，HepG2-R pcDNA SNHG16 miR-570组迁移的细胞个数、CyclinD1、MMP-9、MMP-2水平升高，抑制率、凋亡率、P21水平降低（P<0.05）。结论：lncRNA SNHG16通过调控HepG2-R肝癌细胞的耐药性，其机制与lncRNA SNHG16靶向调控miR-570有关，为临床治疗肝癌细胞提供新的靶点。
Objective: To explore the mechanism of long non-coding RNA SNHG16 (Long non-coding RNA small nucleolar RNA host gene16, lncRNA SNHG16) to anti-liver cancer cell resistance by regulating minimal RNA-570 (miR-570). Methods: The expression of lncRNA SNHG16 and miR-570 of HepG 2 and HepG 2-R cells in human normal liver and hepatoma cells was measured by real-time fluorescence RT-PCR, cell proliferation, apoptosis and invasion were measured by MTT assay, FACS and Transwell test, and cell expression changes of CyclinD1, P21MMP-9 and MMP-2 were determined by Western Blot assay. Results: Compared with normal hepatocytes, lncRNA SNHG16 expression was increased and miR-570 expression was decreased in hepatoma cells (P<0.05). Compared with HepG2-P group, LncRNA SNHG16 and IC50 values were increased in HepG2-R group, and the inhibition rates of miR-570 and HepG2-R cells were decreased at Sorafenib concentrations of 1, 2, 4, 8, 16μmol/L (P<0.05). HepG2-R pcDNA SNHG16 as overexpression group, lncRNA SNHG16 expression was significantly increased (P<0.05), and compared with HepG2-R pcDNA group, In HepG2-R pcDNA SNHG16 group, the number of migrated cells, the expressions of CyclinD1, P21, MMP-9 and MMP-2 were decreased, while the expression of inhibition rate, apoptosis rate and P21 were increased (P<0.05). Compared with HepG2-R anti-miR-NC group, miR-570 level in HepG2-R anti-miR-570 group was decreased (P<0.05), and compared with HepG2-R anti-miR-NC group, HepG2-R anti-miR-570 group decreased the levels of CyclinD1, MMP-9 and MMP-2, and increased the levels of inhibition rate, apoptosis rate and P21 (P<0.05). Dual luciferase reporting experiment showed that compared with miR-NC group, miR-570 reduced the luciferase activity of WT-SNHG16 (P<0.05), but had little effect on the luciferase activity of T-SnHG16 (P>0.05). Overexpression of lncRNA SNHG16 decreased the expression of miR-570 in HepG2-R cells (P<0.05). Compared with HepG2-R pcDNA SNHG16 miR-NC group, In HepG2-R pcDNA SNHG16 miR-570 group, the number of migrated cells, the levels of CyclinD1, MMP-9 and MMP-2 were increased, while the levels of inhibition rate, apoptosis rate and P21 were decreased (P<0.05). Conclusion: By regulating the drug resistance of HepG2-R hepatoma cells, the mechanism is related to the regulation of miR-570 by lncRNA SNHG16, and it can be used as a new target for drug-resistant HCC cell therapy.