miR-4306通过下调SATB2表达抑制骨肉瘤细胞增殖、迁移与侵袭

目的: 研究miR-4306在骨肉瘤细胞中的表达及其对骨肉瘤细胞生物学行为的影响。方法: 选取人成骨细胞hFOB 1.19和骨肉瘤细胞Saos-2、MNNG/HOS CI #5，采用qRT-PCR检测miR-4306表达水平。将miR-4306 mimics和阴性对照分别转染至骨肉瘤细胞Saos-2和MNNG/HOS CI #5，采用CCK-8法和克隆形成实验检测细胞增殖，Transwell实验检测细胞迁移及侵袭，蛋白质免疫印迹检测细胞内上皮间质转化（epithelial mesenchymal transition, EMT）标志蛋白表达水平。通过在线靶基因预测网站Targetscan、Starbase预测miR-4306靶基因可能为富含AT序列特异性结合蛋白2（special AT rich sequence binding protein 2, SATB2），荧光素酶报告系统鉴定靶向关系。将miR 4306 mimics、LV SATB2共转染至骨肉瘤细胞中，检测细胞增殖、迁移、侵袭以及EMT标志蛋白表达水平变化。结果: 与人成骨细胞hFOB 1.19比较，骨肉瘤细胞Saos 2、MNNG/HOS CI #5中miR-4306表达明显降低（P均＜0.01）。转染miR-4306 mimics后，骨肉瘤细胞增殖、迁移、侵袭能力下降，N-钙黏蛋白、血管内皮生长因子表达水平显著降低，E 钙黏蛋白表达水平显著升高（P＜0.05或＜0.01）。SATB2是miR-4306下游的直接靶标；与miR-4306 mimics+LV-NC组比较，miR-4306 mimics+LV-SATB2组部分逆转miR-4306对骨肉瘤细胞增殖、侵袭、迁移和EMT的影响。结论: miR-4306在骨肉瘤细胞中呈低表达，其可通过下调SATB2表达抑制骨肉瘤细胞增殖、迁移、侵袭和EMT。

miR 4306 inhibits the proliferation,migration and invasion of osteosarcoma cells by down-regulating the expression of special AT-rich sequence-binding protein 2

Objective To study the expression of miR 4306 in osteosarcoma cells and its effect on the biological behavior of osteosarcoma cells. Methods Human osteoblast hFOB 1.19 and osteosarcoma cells Saos 2, MNNG/HOS CI #5 were selected, and qRT PCR was used to detect the expression level of miR 4306. miR 4306 mimics and negative control were transfected into osteosarcoma cells Saos 2 and MNNG/HOS CI #5, respectively; and cell proliferation was detected by CCK 8 assay and colony formation assay, cell migration and invasion were detected by Transwell assay, and Western blotting was used to detect the expression levels of epithelial mesenchymal transition （EMT） marker proteins. The online target gene prediction websites Targetscan and Starbase predicted that the target gene of miR 4306 might be special AT rich sequence binding protein 2 （SATB2）, and the luciferase reporter system was used to identify the target relationship. miR 4306 mimics and LV SATB2 were co transfected into osteosarcoma cells, and the changes in cell proliferation, migration, invasion and EMT marker protein expression levels were detected. Results Compared with human osteoblast hFOB 1-19, the expression of miR-4306 in osteosarcoma cells Saos-2 and MNNG/HOS CI #5 was significantly decreased （both P＜0.01）. After transfection with miR-4306 mimics, the proliferation, migration and invasion abilities of osteosarcoma cells were significantly decreased, the expression levels of N cadherin and vascular endothelial growth factor were significantly decreased, and the expression level of E cadherin was greatly increased （P<0.05 or<0.01）. SATB2 is a direct downstream target of miR-4306; compared with the miR-4306 mimics+LV-NC group, the miR-4306 mimics+LV-SATB2 group partially reversed the effects of miR-4306 on osteosarcoma cell proliferation, invasion, migration and EMT. Conclusion miR-4306 is lowly expressed in osteosarcoma cells, and it could inhibit osteosarcoma cell proliferation, migration, invasion and EMT by down regulating the expression of SATB2.