微小RNA-548c-3p通过靶向调控PCNA基因表达对喉癌细胞增殖及凋亡影响的研究  

Effect of miR-548c-3p on proliferation and apoptosis of laryngeal carcinoma cells via targeting PCNA gene

作　　者：朱姝 王平 李丹 ZHU Shu;WANG Ping;LI Dan(Department of Otolaryngology,Chengdu Medical College second affiliated Hospital Nuclear Industry 416 Hospital,Chengdu,Sichuan,610000,China;Department of Otolaryngology,Hubei Medical College affiliated Dongfeng Hospital,Shiyan,Hubei,442000,China)

机构地区：[1]成都医学院第二附属医院核工业四一六医院耳鼻咽喉科,四川成都610000 [2]湖北医药学院附属东风医院耳鼻咽喉科,湖北十堰442000

出　　处：《中国耳鼻咽喉头颈外科》2021年第7期409-414,共6页Chinese Archives of Otolaryngology-Head and Neck Surgery

摘　　要：目的探讨微小RNA-548c-3p(miR-548c-3p)对喉癌细胞增殖及凋亡的影响及其作用机制。方法用miR-548c-3p mimic、增殖细胞核抗原(PCNA)-siRNA及其阴性对照分别或同时转染喉癌TU686和M4e细胞,用qRT-PCR与Western blot法检测miR-548c-3p和PCNA的表达;MTT法和流式细胞术分别检测转染细胞的增殖和凋亡;双荧光素酶报告基因实验观察miR-548c-3p和PCNA的靶向关系。Western blot法检测PCNA、细胞周期蛋白1(CyclinD1)、B淋巴细胞瘤-2(Bcl-2)、B淋巴细胞瘤-2相关蛋白(Bax)的表达。结果 miR-548c-3p的表达量在癌旁组织(0.74±0.10)、喉癌组织(1.94±0.16),TU686细胞(0.35±0.07)和M4e细胞(0.41±0.03)中均呈低表达,而PCNA mRNA和蛋白均呈高表达,差异有统计学意义(P均<0.05)。证实PCNA上存在miR-548c-3p的结合位点。上调miR-548c-3p可抑制细胞增殖及CyclinD1、Bcl-2的表达,并可促进细胞凋亡及Bax的表达,差异有统计学意义(P均<0.05);上调miR-548c-3与抑制PCNA对细胞增殖及凋亡的作用相似。pcDNA-PCNA显著减弱miR-548c-3p mimic对细胞增殖和凋亡的作用。结论 miR-548c-3p通过抑制PCNA表达而降低喉癌细胞增殖能力并诱导细胞凋亡。OBJECTIVE To investigate the effect of microRNA-548c-3p(miR-548c-3p) on proliferation and apoptosis of laryngeal carcinoma cells and analyze its mechanism.METHODS The laryngeal carcinoma TU686 and M4e cells were transfected with miR-548c-3p mimic,PCNA-siRNA and their negative controls,respectively.The expression of miR-548c-3p and PCNA was detected by qRT-PCR and Western blot.MTT assay and flow cytometry were used to detect proliferation and apoptosis of transfected cells,respectively.Dual luciferase reporter gene experiments observed the targeting relationship between miR-548c-3p and PCNA.Western blot was used to detect the expression of PCNA,CyclinD1,Bcl-2 and Bax.RESULTS The expression level of miR-548c-3p was decreased in paracancerous tissue(0.74±0.10),Laryngeal cancer tissue(1.00±0.14),TU686 cell(0.35±0.07),and M4e cell(0.41±0.03).PCNA protein and mRNA was highly expressed,the difference was statistically significant(all P<0.05).At the same time,it was confirmed that there was a binding site of miR-548c-3p on PCNA.Up-regulation of miR-548c-3p could inhibit cell proliferation and could inhibit the protein level of CyclinD1,Bcl-2,and could promote cell apoptosis and the protein level of Bax,the difference was statistically significant(all P<0.05).Up-regulation of miR-548c-3 was similar to the effect of inhibiting PCNA on cell proliferation and apoptosis.pcDNA-PCNA could attenuate the effects of miR-548c-3p mimic on cell proliferation and apoptosis.CONCLUSION miR-548c-3p reduces the proliferation of laryngeal carcinoma cells and induces apoptosis by inhibiting PCNA expression.

关 键 词：喉肿瘤 细胞增殖 细胞凋亡 微RNAS 细胞周期蛋白D 

分 类 号：R739.65[医药卫生—肿瘤]