LncRNA LINC00174调控miR-153-5p对乳腺癌细胞MDA-MB-468增殖凋亡及迁移的影响

Effect of LncRNA LINC00174 on proliferation, apoptosis, and migration of breast cancer cell MDA-MB-468 by regulating miR-153-5p

目的分析长链非编码RNA(LncRNA)LINC00174调控miR-153-5p对乳腺癌细胞MDA-MB-468增殖、凋亡及迁移的影响。方法收集2018年6月-2019年12月在宁波大学附属人民医院接受手术治疗的35例乳腺癌患者的乳腺癌组织和癌旁组织,分为si-NC组、si-LINC00174组、si-LINC00174+anti-miR-NC组及si-LINC00174+anti-miR-153-5p组。采用qRT-PCR检测LINC00174、miR-153-5p的表达水平,采用MTT检测细胞增殖率,采用平板克隆形成实验检测克隆形成能力,采用划痕实验检测细胞迁移率,采用流式细胞术检测细胞凋亡率,采用双荧光素酶报告基因检测LINC00174与miR-153-5p的靶向关系。结果癌旁组织中LINC00174和miR-153-5p的表达水平分别为(1.03±0.11)和(0.96±0.15),乳腺癌组织中LINC00174和miR-153-5p的表达水平分别为(5.54±0.44)和(0.21±0.05),差异均有统计学意义(t=58.829,P=0.000;t=28.062,P=0.000)。si-NC组的LINC00174表达水平、miR-153-5p表达水平、抑制率、划痕愈合率及克隆形成数分别为(1.00±0.00)、(1.00±0.00)、(0.00±0.00)%、(65.78±2.92)%及(96.00±3.74)个,si-LINC00174组的LINC00174表达水平、miR-153-5p表达水平、抑制率、划痕愈合率及克隆形成数分别为(0.15±0.01)、(5.28±0.09)、(58.60±3.48)%、(24.71±1.46)%及(47.67±1.25)个,差异均有统计学意义(t=147.224,P=0.000;t=82.369,P=0.000;t=19.166,P=0.000;t=48.723,P=0.000;t=21.228,P=0.000)。si-NC组细胞凋亡率为(6.50±0.35)%,Bax蛋白水平为(0.17±0.02),Bcl-2蛋白水平为(0.60±0.06);si-LINC00174组细胞凋亡率为(24.40±0.70)%,Bax蛋白水平为(0.72±0.06),Bcl-2蛋白水平为(0.11±0.01)。与si-NC组比较,si-LINC00174组细胞凋亡率升高,Bcl-2蛋白水平降低,Bax蛋白水平升高,差异均有统计学意义(t=52.562,15.062,13.953,均P<0.05)。与si-LINC00174+anti-miR-NC组比较,si-LINC00174+anti-miR-153-5p组细胞增殖抑制率降低,划痕愈合率升高,克隆形成数增多,差异均有统计学意义(均P<0.05)。与si-LINC00174+anti-miR-NC组比较,si-LINC00174+anti-miR-153-5p组细胞凋亡率降低,Bax蛋白水平降低,Bcl-2蛋白水平升高,差异均有统计学意义(均P<0.05)。结论乳腺癌组织中LINC00174的表达水平升高,miR-153-5p的表达水平降低,乳腺癌细胞增殖、迁移及克隆形成可通过干扰LINC00174的表达而有效抑制,进而促进细胞凋亡。

Objective To analyze the effect of long non-coding RNA(LncRNA)LINC00174 on proliferation,apoptosis,and migration of breast cancer cell MDA-MB-468 by regulating miR-153-5 p.Methods From June 2018 to December 2019,the breast cancer tissue samples and tissue samples adjacent to breast cancer of 36 patients with breast cancer treated by surgery in Affiliated People’s Hospital of NingboUniversity were collected and divided into si-NC group,si-LINC00174 group,si-LINC00174+anti-miR-NC group,and si-LINC00174+anti-miR-153-5 p group.qRT-PCR was used to detect the expression levels of LINC00174 and miR-153-5 p,cell proliferation rate was detected by MTT method,clone forming ability was detected by plate clone formation experiment,cell migration rate was detected by scratch assay,cell apoptosis rate was detected by flow cytometry,the targeted relationship between LINC00174 and miR-153-5 p was detected by double fluorescent enzyme gene experiment.Results The expression levels of LINC00174 and miR-153-5 p in the tissue samples adjacent to breast cancer were(1.03±0.11)and(0.96±0.15),respectively,the expression levels of LINC00174 and miR-153-5 p in breast cancer tissue samples were(5.54±0.44)and(0.21±0.05),respectively,there were statistically significant differences(t=58.829,P=0.000;t=28.062,P=0.000).The expression levels of LINC00174 and miR-153-5 p,the inhibition rate,the scratch healing rate,and clone formation number in si-NC group were(1.00±0.00),(1.00±0.00),(0.00±0.00)%,(65.78±2.92)%,and(96.00±3.74),respectively,the expression levels of LINC00174 and miR-153-5 p,the inhibition rate,the scratch healing rate,and clone formation number in si-LINC00174 group were(0.15±0.01),(5.28±0.09),(58.60±3.48)%,(24.71±1.46)%,and(47.67±1.25),respectively,there were statistically significant differences between the two groups(t=147.224,P=0.000;t=82.369,P=0.000;t=19.166,P=0.000;t=48.723,P=0.000;t=21.228,P=0.000).In si-NC group,cell apoptosis rate was(6.50±0.35)%,Bax protein level was(0.17±0.02),Bcl-2 protein level was(0.60±0.06);in si-LINC00174 group,cell apoptosis rate was(24.40±0.70)%,Bax protein level was(0.72±0.06),Bcl-2 protein level was(0.11±0.01).Compared with si-NC group,cell apoptosis rate in si-LINC00174 group increased,Bcl-2 protein level decreased,Bax protein level increased,there were statistically significant differences between the two groups(P<0.05).Compared with si-LINC00174+anti-miR-NC group,the inhibition rate of cell proliferation decreased,scratch healing rate increased,clone formation number increased,there were statistically significant differences between the two groups(P<0.05).Compared with si-LINC00174+anti-miR-NC group,cell apoptosis rate in si-LINC00174+anti-miR-153-5 p group decreased,Bax protein level decreased,Bcl-2 protein level increased,there were statistically significant differences(P<0.05).Conclusion The expression level of LINC00174 in breast cancer tissue increases,the expression level of miR-153-5 p decreases,the proliferation,migration,and clone formation of breast cancer cells can be inhibited by interfering LINC00174 expression,then cell apoptosis is promoted.