微小RNA-98靶向N-Ras调控涎腺腺样囊性癌侵袭和迁移

MicroRNA -98 regulates invasion and migration of salivary adenoid cystic carcinoma by targeting N -Ras

目的观察微小RNA（miRNA，miR）-98对涎腺腺样囊性癌（SACC）细胞增殖、迁移和侵袭能力的影响。方法运用实时定量反转录聚合酶链反应（RT-qPCR）检测miR-98在SACC新鲜组织及细胞株中的表达。利用生物信息软件预测出miR-98的靶基因为N-Ras。免疫组织化学法检测N-Ras在SACC组织标本中的表达并分析其临床意义。上调/下调SACC细胞株miR-98的表达后，应用噻唑盐（MTT）法、平板克隆形成实验、划痕实验、Transwell侵袭实验检测miR-98对SACC细胞增殖、迁移及侵袭影响。并采用双荧光素酶报告基因系统、RT-qPCR和Western blot检测miR-98与N-Ras的靶向关系。结果miR-98在SACC组织中及ACC-M细胞中呈低表达，N-Ras在SACC组织中呈高表达，其表达与肿瘤T分期和临床分期相关。转染miR-98 mimics后，ACC-M细胞的增殖能力降低，发生侵袭的细胞数由（158.0±5.5）个减少至（92.0±4.7）个（t＝15.800，P＝0.000），细胞24 h迁移距离由（0.42±6.30） mm减少至（0.25±6.00） mm（t＝4.630，P＝0.001）；而当miR-98被抑制后，SACC-83细胞的增殖能力增强，发生侵袭的细胞数[（123.0±10.6）]个比对照组[（88.0±4.0）个]增多（t＝5.254，P＝0.006），细胞24 h迁移距离由（0.15±7.50） mm增加（0.38±8.80） mm（t＝9.109，P＝0.000）。此外双荧光素酶报告基因系统、RT-qPCR和Western blot均证实miR-98能够调控N-Ras的表达。结论miR-98可以作用于N-Ras，进一步调控SACC细胞的侵袭和迁移能力。

ObjectiveTo investigate the effect of microRNA （miRNA, miR）-98 on suppression of proliferation, migration and invasion in salivary adenoid cystic carcinoma （SACC）.MethodsThe expression of miR-98 was compared between SACC tissues and the adjacent tissues of 3 patients, as well as between SACC cell lines by real-time reverse transcriptase-polymerase chain reaction （RT-qPCR）. Bioinfornatics analysis predicted miR-98 targeting site in gene N-Ras. N-Ras expression was detected in SACC tissues using immunohistochemistry. And the associations between N-Ras expression and clinicopathological features were analyzed. The effects of miR-98 on the proliferation, migration and invasion of SACC cell lines were also determined. Finally, the target relationship between miR-98 and N-Ras was identified by Dual-Luciferase Assay system, RT-qPCR and Western blotting.Results The expression of miR-98 in SACC tissues and ACC-M cells were both significantly lower than those in adjacent tissues and SACC-83 cells. N-Ras was overexpressed in SACC tissues and its overexpression was associated with the clinical stage and tumor size. After transfection of miR-98 mimics to ACC-M cells, the ability of cell proliferation reduced; invasion cell number reduced from 158.0±5.5 of control group to 92.0±4.7 of experimental group （t=15.800, P=0.000）; and the migration ability reduced from （0.42±6.30）mm to （0.25±6.00） mm （t=4.630, P=0.001）. While the downregulation of miR-98 increased the ability of proliferation. Invasion cell number increased from 88.0±4.0 to 123.0±10.6 （t=5.254, P=0.006） and migration distance increased from （0.15±7.50） mm to （0.38±8.80） mm after transfected with miR-98 inhibitor （t=9.109, P=0.000）. At last, the target relationship between miR-98 and N-Ras was indentified by Dual-Luciferase Assay System.Conclusion MiR-98 can regulate invasion and migration of SACC via targeting N-Ras.