miR-181a-5p通过HOXB4抑制骨肉瘤细胞HOS增殖和迁移
摘要点击次数: 44   全文下载次数: 17   投稿时间:2021-08-20    
作者Author单位AddressE-Mail
李嘉熙 LI Jia-xi 西安交通大学第二附属医院骨科, 陕西 西安 710004 Department of Orthopaedics, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710004, Shaanxi, China  
贺西京 HE Xi-jing 西安交通大学第二附属医院骨科, 陕西 西安 710004 Department of Orthopaedics, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710004, Shaanxi, China  
李飞 LI Fei 西安交通大学第二附属医院骨科, 陕西 西安 710004 Department of Orthopaedics, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710004, Shaanxi, China  
雷雨田 LEI Yu-tian 西安交通大学第二附属医院骨科, 陕西 西安 710004 Department of Orthopaedics, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710004, Shaanxi, China  
杨于冰 YANG Yu-bing 西安交通大学第二附属医院骨科, 陕西 西安 710004 Department of Orthopaedics, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710004, Shaanxi, China  
李京 LI Jing 西安交通大学第二附属医院骨科, 陕西 西安 710004 Department of Orthopaedics, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710004, Shaanxi, China  
宗高阳 ZONG Gao-yang 西安交通大学第二附属医院骨科, 陕西 西安 710004 Department of Orthopaedics, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710004, Shaanxi, China  
赵敏超 ZHAO Min-chao 西安交通大学第二附属医院骨科, 陕西 西安 710004 Department of Orthopaedics, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710004, Shaanxi, China  
常素娥 CHANG Su-e 西安交通大学第二附属医院骨科, 陕西 西安 710004 Department of Orthopaedics, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710004, Shaanxi, China suechang@xjtu.edu.cn 
期刊信息:《中国骨伤》2022年,第35卷,第11期,第1097-1103页
DOI:10.12200/j.issn.1003-0034.2022.11.017
基金项目:国家自然科学基金(编号:81601081);陕西省自然科学基础研究计划青年项目(编号:2021JQ-413)
中文摘要:

目的: 研究miR-181a-5p对HOS骨肉瘤细胞增殖、周期和迁移的影响及其机制。

方法: 采用实时定量PCR检测hFOB1.19成骨细胞和HOS、U2OS、MG63骨肉瘤细胞系中miR-181a-5p及HOXB4的表达情况。利用Lipofectamine 2000将miR-181a-5p mimics和miR-181a-5p inhibitor分别转染至人骨肉瘤HOS细胞中(分别为过表达组和抑制剂组),并设置miR 阴性对照组;CCK-8法检测各组细胞的增殖能力变化,流式细胞术检测各组细胞的细胞周期变化,划痕愈合实验以及Transwell迁移实验检测各组细胞的迁移能力变化。Targetscan网站预测miR-181a-5p的靶向基因,并通过双荧光素酶报告基因系统及Western blot验证靶向关系。

结果: 与成骨细胞hFOB1.19相比,miR-181a-5p在骨肉瘤细胞HOS、U2OS和MG63中低表达(P<0.05),而HOXB4在骨肉瘤中高表达(P<0.05)。与阴性对照组相比,过表达miR-181a-5p抑制骨肉瘤HOS细胞的增殖和迁移能力,并且处于细胞周期S期的细胞减少(P<0.05)。敲低miR-181a-5p促进骨肉瘤HOS细胞的增殖和迁移能力,并且处于S期细胞增加(P<0.05)。生物信息学预测及双荧光素酶报告基因系统验证HOXB4为miR-181a-5p的下游靶基因(P<0.05)。Western blot显示,过表达miR-181a-5p的HOS细胞中,HOXB4蛋白表达低于阳性对照组(P<0.05),而敲低miR-181a-5p的HOS细胞中HOXB4蛋白表达高于对照组(P<0.05)。

结论: miR-181a-5p在骨肉瘤细胞中低表达,过表达miR-181a-5p能够抑制骨肉瘤细胞HOS增殖、周期和迁移能力,该作用可能通过靶向HOXB4发挥作用。
【关键词】骨肉瘤  细胞增殖  细胞周期  细胞迁移
 
miRNA-181a-5p inhibits proliferation and migration of osteosarcoma cell line HOS by targeting HOXB4
ABSTRACT  

Objective: To study the effects and mechanisms of miR-181a-5p on the proliferation,cycle and migration of HOS osteosarcoma cells.

Methods: Real-time quantitative PCR was used to detect the expression of miR-181a-5p and HOXB4 in osteoblast hFOB1.19 cell line and osteosarcoma cell lines (HOS,U2OS,MG63). miR-181a-5p mimics and miR-181a-5p inhibitors were respectively transfected into HOS cells by Lipofectamine 2000,and miR NC group was set as control group. CCK-8 method was used to detect the change in cell proliferation. Flow cytometry was used to detect the changes in cell cycles. Wound healing experiments and Transwell migration experiments were used to detect the changes in cell migration ability. The target gene of miR-181a-5p was predicted by Targetscan website and validated by Dual-luciferase reporter gene system and Western blot.

Results: Compared with osteoblast hFOB1.19,miR-181a-5p was low expressed in osteosarcoma cells HOS,U2OS,and MG63(P<0.05),while HOXB4 was high expressed in osteosarcoma cells HOS,U2OS,and MG63(P<0.05). Compared with the miR NC group,over expression of miR-181a-5p inhibited the proliferation and migration of osteosarcoma HOS cells,and the number of cells in S phase decreased(P<0.05). However,knockdown miR-181a-5p promoted the proliferation and migration of osteosarcoma HOS cells,the cells in S phase increased(P<0.05). Bioinformatics prediction and Dual-luciferase reporter gene system validate HOXB4 as a downstream target gene of miR-181a-5p(P<0.05). Western blot showed that miR-181a-5p over expression or knockdown significantly down-regulated or up-regulated HOXB4 expressions in the HOS cells respectively(P<0.05).

Conclusion: miR-181a-5p is down expressed in osteosarcoma cells,and over-expression miR-181a-5p inhibits the proliferation,cell cycle and migration ability of osteosarcoma cells by targeting HOXB4.
KEY WORDS  Osteosarcoma  Cell proliferation  Cell cycle  Cell migration
 
引用本文,请按以下格式著录参考文献:
中文格式:李嘉熙,贺西京,李飞,雷雨田,杨于冰,李京,宗高阳,赵敏超,常素娥.miR-181a-5p通过HOXB4抑制骨肉瘤细胞HOS增殖和迁移[J].中国骨伤,2022,35(11):1097~1103
英文格式:LI Jia-xi,HE Xi-jing,LI Fei,LEI Yu-tian,YANG Yu-bing,LI Jing,ZONG Gao-yang,ZHAO Min-chao,CHANG Su-e.miRNA-181a-5p inhibits proliferation and migration of osteosarcoma cell line HOS by targeting HOXB4[J].zhongguo gu shang / China J Orthop Trauma ,2022,35(11):1097~1103
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