【作者】 虞佳；

【导师】 文格波；

【作者基本信息】 南华大学 ， 基础医学， 2022， 博士

【摘要】 背景与目的:乳腺癌(Breast cancer,BC)已经成为全球发病率最高的恶性肿瘤,在女性癌症死亡原因中位列第一。尽管近年来在乳腺癌的诊断和治疗方面取得了很大的进步,但由于乳腺癌具有早期隐蔽性、高度异质性且易复发的特点,其进展期结果不尽人意。因此,积极探索乳腺癌发生发展的分子机制,寻找与乳腺癌诊断和预后相关的生物标记物至关重要。长链非编码RNA(Long noncoding RNA,lncRNA)是一类长度超过200 nt的非编码RNA。目前,众多研究已经证实lncRNA参与乳腺癌的发生发展。lncRNA可以作为微小RNA(micro RNA,miRNA)的竞争内源性RNA(competing endogenous RNA,ceRNA),阻断miRNA与其靶mRNA的结合从而调控下游基因表达,进而影响乳腺癌的发生发展。然而,现在人们了解到的lncRNA只是冰山一角,绝大部分lncRNA在乳腺癌中的作用仍然未知。高通量测序技术和公共数据库的开发为研究乳腺癌驱动基因和相关信号通路提供了极大的便利。本研究从美国癌症基因图谱数据库(The Cancer Genome Atlas,TCGA)筛选到在乳腺癌中表达显著上调且与预后相关的LINC01234,进一步在临床组织样本中验证其表达水平,分析与临床病理特征的相关性,并在体外探索其对乳腺癌细胞增殖、凋亡、侵袭和迁移等生物学功能的影响及其可能的分子机制,为乳腺癌的预后和靶向治疗提供新的实验依据。第一章 基于TCGA数据库筛选与乳腺癌预后相关的lncRNA并验证其表达方法:(1)使用TCGA biolinks R包下载TCGA-BRCA lncRNA readscount数据;利用limma包进行数据标准化;使用Deseq2 R包对lncRNA表达数据进行差异表达分析,使用ggplot2 R包绘制lncRNA差异表达的火山图,使用pheatmap包绘制病人lncRNA差异表达的热图;使用R软件cor.test函数计算lncRNA在肿瘤组和正常组的表达相关性;采用survival包和survminer包绘制KM曲线分析lncRNA表达对生存预后的影响。(2)收集86对乳腺癌患者的癌组织和癌旁正常组织,采用qRT-PCR检测LINC01234在乳腺癌组织的表达水平,卡方检验分析LINC01234表达水平与乳腺癌患者临床病理指标的相关性,qRT-PCR检测LINC01234在人正常乳腺上皮细胞和乳腺癌细胞系中的表达。结果:(1)差异分析结果显示共有3034个差异表达的lncRNAs,其中有2151个上调表达和883个下调表达,与乳腺癌的相关系数r=0.953。生存分析结果显示276个lncRNA的表达与乳腺癌患者的生存预后显著相关,其中LINC01234高表达的乳腺癌患者生存时间显著低于LINC01234低表达的乳腺癌患者(P=0.0113)。(2)与癌旁正常组织相比,LINC01234在乳腺癌组织中的表达显著上调(P<0.001)。LINC01234表达水平与患者肿瘤大小显著相关(P=0.005),与年龄、性别、分期、淋巴结转移、ER、PR及HER2的表达无显著相关性(P>0.05)。与人正常乳腺上皮细胞(MCF-10A)相比,LINC01234在人乳腺癌细胞系(SKBR-3、MCF-7、HCC1806、HS-578T、MDA-MB-468、T-47D和MDA-MB-231)中均显著高表达(P<0.01)。第二章 LINC01234对乳腺癌细胞增殖、侵袭和迁移的影响方法:培养人乳腺癌细胞MCF-7和MDA-MB-468,瞬时转染LINC01234过表达质粒和小干扰RNA,CCK-8实验、克隆形成实验、流式细胞术、Transwell实验和划痕实验检测LINC01234对MCF-7和MDA-MB-468细胞增殖、周期、凋亡、侵袭和迁移的影响。结果:在MCF-7和MDA-MB-468中,分别转染pc DNA3.1-LINC01234和siRNA-LINC01234,与对照组相比,pc DNA3.1-LINC01234组的细胞增殖、侵袭和迁移能力均显著增强,siRNA-LINC01234组的细胞增殖、侵袭和迁移能力均显著减弱(P<0.01)。pc DNA3.1-LINC01234组的G0/G1期细胞数量显著减少而处于S期的细胞增加,siRNA-LINC01234组的G0/G1期的细胞数量显著增加而处于S期的细胞减少(P<0.05)。pc DNA3.1-LINC01234组的细胞凋亡率显著减少,siRNA-LINC01234组的细胞凋亡率显著增加(P<0.01)。第三章 LINC01234调控乳腺癌细胞生物学功能的分子机制方法:(1)荧光原位杂交(FISH)检测LINC01234在MCF-7和MDA-MB-468中的细胞定位;miRcode、Star Base和lnc Basev2.0在线预测LINC01234的靶miRNA。(2)qRT-PCR检测86对乳腺癌组织样本和乳腺癌细胞系中miR-525-5p的表达;Pearson相关性分析LINC01234与miR-525-5p表达相关性;双荧光素酶报告实验、RNA pull-down和AGO2-RIP实验验证LINC01234与miR-525-5p靶向关系。(3)CCK-8实验、克隆形成实验、流式细胞术、Transwell实验和划痕实验检测miR-525-5p对MCF-7和MDA-MB-468细胞增殖、周期、凋亡、侵袭和迁移的影响;互补实验分析LINC01234与miR-525-5p调控关系。(4)RNA22、Target Scan、miRmap、micro T以及Pic Tar在线预测miR-525-5p的靶基因;免疫组化检测乳腺癌组织样本中CSDE1蛋白的表达;qRT-PCR和Western blot分别检测乳腺癌细胞系中CSDE1mRNA和蛋白的表达水平;双荧光素酶报告实验、RNA pull-down和AGO2-RIP实验验证miR-525-5p与CSDE1 mRNA靶向关系。(5)CCK-8实验、克隆形成实验、流式细胞术、Transwell实验和划痕实验检测CSDE1对MCF-7和MDA-MB-468细胞增殖、周期、凋亡、侵袭和迁移的影响;互补实验分析miR-525-5p与CSDE1 mRNA调控关系。(6)功能互补实验分析LINC01234、miR-525-5p和CSDE1mRNA调控关系;KEGG信号通路富集分析miR-525-5p下游靶基因参与的信号通路;Western blot检测各处理组中PI3K/AKT信号通路相关蛋白的表达水平;CCK-8和流式细胞术检测LINC01234联合PI3K抑制剂对细胞增殖和凋亡的影响。结果:(1)FISH定位分析发现LINC01234主要定位于MCF-7和MDA-MB-468细胞质中。生物信息学预测LINC01234与miR-525-5p具有靶向结合位点。(2)与癌旁正常组织相比,miR-525-5p在乳腺癌组织中显著低表达(P<0.001),miR-525-5p表达水平与LINC01234表达水平呈负相关(r=-0.2620)。与MCF-10A相比,miR-525-5p在乳腺癌细胞系(SKBR-3、MCF-7、HS-578T、MDA-MB-468、T-47D和MDA-MB-231)中均显著低表达(P<0.05)。在MCF-7和MDA-MB-468细胞中成功过表达和敲低LINC01234后,与对照组相比,LINC01234过表达组中miR-525-5p显著低表达,LINC01234敲低组中miR-525-5p显著高表达(P<0.01)。双荧光素酶报告实验结果显示只有LINC01234-WT和miR-525-5p共转染组的荧光素酶活性下降(P<0.05);RNA pull-down结果显示只有生物素标记的miR-525-5p正义链组能够检测到LINC01234的目的片段;AGO2-RIP实验结果显示在共沉淀的复合物中能检测到LINC01234与miR-525-5p。(3)在MCF-7和MDA-MB-468中,分别转染miR-525-5p mimics和inhibitor,与对照组比较,miR-525-5p mimics组细胞增殖、侵袭和迁移能力减弱,miR-525-5p inhibitor组细胞增殖、侵袭和迁移能力增强(P<0.05);miR-525-5p mimics组G0/G1期的细胞数量显著增加而处于S期的细胞减少,miR-525-5p inhibitor组G0/G1期的细胞数量显著减少而处于S期的细胞增加(P<0.05);miR-525-5p mimics组细胞凋亡率升高,miR-525-5p inhibitor组细胞凋亡率下降(P<0.01)。互补实验发现miR-525-5p能削弱LINC01234对MCF-7和MDA-MB-468细胞增殖、周期、侵袭和迁移的促进作用以及细胞凋亡的抑制作用。(4)生物信息学预测miR-525-5p的靶基因,提示miR-525-5p与CSDE1 mRNA具有靶向结合位点。免疫组化结果显示与癌旁正常组织相比,CSDE1蛋白在癌组织中高表达。与MCF-10A相比,CSDE1mRNA和蛋白水平均在乳腺癌细胞系(SKBR-3、MCF-7、HCC1806、HS-578T、MDA-MB-468、T-47D和MDA-MB-231)中显著高表达(P<0.05)。在MCF-7和MDA-MB-468中成功过表达和敲低miR-525-5p后,与对照组相比,miR-525-5p mimics组中CSDE1 mRNA和蛋白水平均显著低表达,miR-525-5p inhibitor组中CSDE1 mRNA和蛋白水平均显著高表达(P<0.01)。双荧光素酶报告实验结果显示只有CSDE1-3’UTR-WT和miR-525-5p共转染组的荧光素酶活性下降(P<0.01);RNA pull-down结果显示只有生物素标记的miR-525-5p正义链组能够检测到CSDE1 mRNA的目的片段;AGO2-RIP实验结果显示共沉淀的复合物中能检测到CSDE1 mRNA与miR-525-5p。(5)在MCF-7和MDA-MB-468中,成功转染pc DNA-CSDE1和siRNA-CSDE1,与对照组相比,pc DNA-CSDE1组的细胞增殖、侵袭和迁移能力增强,siRNA-CSDE1组的细胞增殖、侵袭和迁移能力减弱(P<0.05);pc DNA-CSDE1组G0/G1期的细胞数量显著减少而处于S期的细胞增加,siRNA-CSDE1组G0/G1期的细胞数量显著增加而处于S期的细胞减少(P<0.05);pc DNA-CSDE1组的细胞凋亡率下降,siRNA-CSDE1组的细胞凋亡率显著上升(P<0.01)。互补实验发现在CSDE1能削弱miR-525-5p对MCF-7和MDA-MB-468细胞增殖、周期、侵袭和迁移的抑制作用以及细胞凋亡的促进作用。(6)在MCF-7和MDA-MB-468中成功过表达和敲低LINC01234,与对照组相比,LINC01234过表达组中CSDE1 mRNA和蛋白水平均显著高表达,LINC01234敲低组中CSDE1 mRNA和蛋白水平均显著低表达(P<0.01)。在挽救实验中,与对照组相比,miR-525-5p mimics组中的CSDE1 mRNA和蛋白水平显著低表达,而在共转染miR-525-5p mimics和pc DNA-LINC01234后,CSDE1 mRNA和蛋白表达水平得以回升(P<0.01)。CCK-8和流式细胞术结果显示,与对照组相比,miR-525-5p mimics组细胞增殖活力下降,细胞凋亡增加,而在共转染miR-525-5p mimics和pc DNA-LINC01234后,这种现象得以逆转(P<0.01)。KEGG信号通路分析结果显示,miR-525-5p下游靶mRNA与PI3K/AKT通路有关。在MCF-7和MDA-MB-468中转染siRNA-LINC01234和pc DNA-LINC01234,Westen blot结果显示,与对照组相比,pc DNA-LINC01234组中PTEN的表达显著降低,PI3K-p85(Y607)和p-AKT的表达显著增加,siRNA-LINC01234组中PTEN的表达显著增加,PI3K-p85(Y607)和p-AKT表达显著降低(P<0.01);在过表达LINC01234的MCF-7和MDA-MB-468中,共转染miR-525-5p mimics或siRNA-CSDE1或采用PI3K抑制剂LY294002处理细胞后,与LINC01234过表达组比较,PI3K-p85(Y607)和p-AKT的表达水平显著降低(P<0.05)。CCK-8和流式细胞术结果显示,PI3K抑制剂LY294002能够抑制LINC01234诱导的细胞增殖,逆转LINC01234介导的凋亡抑制(P<0.01)。结论:(1)LINC01234在乳腺癌组织和乳腺癌细胞系中高表达,其表达水平与肿瘤大小相关,生物信息学分析提示LINC01234表达水平与乳腺癌预后相关。(2)LINC01234能抑制乳腺癌细胞凋亡,促进乳腺癌细胞增殖、周期进程、侵袭和迁移。(3)LINC01234作为ceRNA靶向结合miR-525-5p介导CSDE1上调,抑制PTEN表达,激活PI3K/AKT信号通路促进乳腺癌发生发展。更多还原

【Abstract】 Background and objectiveBreast cancer(BC)has been become the most common malignant tumor in the world and the leading cause of cancer death in women.Although great progress has been made in the diagnosis and treatment of BC in recent years,the results of advanced BC are still unsatisfactory due to the early concealment,high heterogeneity and easy recurrence of BC.Therefore,it is very important to explore the molecular mechanism of BC,and to find biomarkers related to the diagnosis and prognosis of BC.Long noncoding RNAs(lncRNAs)are a class of noncoding RNA with a length of more than 200 nt.At present,numerous studies have confirmed that lncRNAs are involved in the occurrence and development of BC.LncRNAs can served as the competing endogenous RNA(ceRNA)of micro RNA(miRNA)to block the binding of miRNA and its target mRNA,thereby regulating the expression of downstream genes and thus affecting the progression of BC.However,the current understanding of lncRNAs are only the tip of the iceberg,and the role of most lncRNAs in BC are still unclear.Recently,high-throughput sequencing technology and the development of public databases have greatly facilitated the study of BC driver genes and relevant signaling pathways.In this study,long non-coding RNA LINC01234,which was differentially upregulated in BC and related to prognosis,was screened from The Cancer Genome Atlas(TCGA)database.The expression level of LINC01234 was verified in clinical tissue specimens.The correlation was analyzed between the expression level of LINC01234 and clinicopathological features.To provide a new experimental evidence for prognosis and targeted therapy of BC,in vitro experiments were conducted to verify the biological functions and mechanism of LINC01234 on the cell proliferation,cell cycle,apoptosis,invasion and migration of BC cells.Chapter 1 Screening of lncRNAs associated with breast cancer prognosis based on TCGA database and verifying its expressionMethods(1)TCGA-BRCA lncRNA readscount data were downloaded by TCGA biolinks R package,which was normalized with the limma package.Deseq2 R package was used to analyze the differentially expressed lncRNA(DElncRNA),ggplot2 R package was used to draw the volcano map of DElncRNA,and pheatmap package was used to draw the heatmap of DElncRNA in patients.Cor.test function of R software was used to calculate the correlation of DElncRNA expression between the tumor group and the normal group.The survival package and survminer package were used to draw KM curves to analyze the effect of lncRNA expression on survival prognosis.(2)Eighty six pairs of BC tissue specimens and adjacent normal tissue specimens were collected.The expression level of LINC01234 in BC tissue specimens and cultural cells was detected by qRT-PCR.The correlation between LINC01234 expression level and clinicopathological indicators of BC patients was analyzed by chi-square test.Results(1)The results of differential analysis showed that 3034 lncRNAs were differentially expressed,of which 2151 were up-regulated and 883 were down-regulated.The correlation coefficient between lncRNAs and BC was 0.953.Survival analysis showed that the expression of 276 lncRNAs significantly affected the survival prognosis of BC patients.Survival time of BC patients with high LINC01234 expression was significantly lower than that of BC patients with low LINC01234expression(P = 0.0113).(2)Compared with patinets’ adjacent normal tissues,cancer tissues had significantly higher expression of LINC01234(P < 0.001).The expression level of LINC01234 was significantly correlated with tumor size(P = 0.005),but not with age,gender,stage,lymph node metastasis and the levels of ER,PR and HER2(P > 0.05).Compared with the normal breast cell line(MCF-10A),LINC01234 was significantly higher expressed in multiple BC cell lines(SKBR-3、MCF-7、HCC1806、HS-578T、MDA-MB-468、T-47 D and MDA-MB-231)(P < 0.01).Chapter 2: The effects of LINC01234 on proliferation,invasion and migration of breast cancer cellsMethodsBC cells(MCF-7 and MDA-MB-468)were cultured and transiently transfected with LINC01234 overexpressing plasmid and small interfering RNA.CCK-8 assay,clone formation assay,flow cytometry,transwell assay and scratch wound assay were used to detect the effects of LINC01234 on the proliferation,cell cycle,apoptosis,invasion and migration of MCF-7 and MDA-MB-468 cell lines.ResultsIn MCF-7 and MDA-MB-468 cell lines,the cell proliferation,invasion and migration abilities were significantly enhanced in LINC01234 overexpression group,while weakened in LINC01234 knockdown group(P < 0.01).The number of G0/G1 phase cells were significantly decreased,but S phase cells increased in LINC01234 overexpression group.On the contrary,the number of G0/G1 phase cells were significantly increased,but S phase cells decreased in LINC01234 knockdown group(P < 0.05).The apoptosis rate of cells significantly decreased in LINC01234 overexpression group,while increased in LINC01234 knockdown group(P < 0.01).Chapter 3: Molecular mechanism of LINC01234 regulating the biological function of breast cancer cellsMethods(1)Fluorescence in situ hybridization(FISH)was used to detect the localization of LINC01234 in MCF-7 and MDA-MB-468 cell lines.MiRcode,Star Base and lnc Basev2.0 database were applied to predict the targeting miRNAs of LINC01234.(2)The expression of miR-525-5p in eighty six pairs BC tissue specimens and BC cells were detected by qRT-PCR.The correlation between LINC01234 and miR-525-5p was analyzed with pearson correlation coefficient.Dual luciferase reporter assay,RNA pull-down assay and AGO2-RIP assay were used to comfirm the binding relationship between LINC01234 and miR-525-5p.(3)CCK-8 assay,clone formation assay,flow cytometry,transwell assay and scratch wound assay were used to detect the effects of miR-525-5p on the proliferation,cell cycle,apoptosis,invasion and migration of MCF-7 and MDA-MB-468.The regulatory relationship between LINC01234 and miR-525-5p was analyzed by complementation assay.(4)RNA22,Target Scan,miRmap,micro T,and Pic Tar database were applied to predict the target genes of miR-525-5p.The expression levels of CSDE1 protein were detected by immunohistochemistry(IHC).The expression levels of CSDE1 mRNA and its protein in BC cells were detected by qRT-PCR and Western blot respectively.Double luciferase reporter assay,RNA pull-down assay and AGO2-RIP assay were used to verify the targeting relationship between miR-525-5p and CSDE1 mRNA.(5)CCK-8 assay,clone formation assay,flow cytometry,transwell assay and scratch wound assay were used to detect the effects of CSDE1 on the proliferation,cell cycle,apoptosis,invasion and migration of MCF-7 and MDA-MB-468.The regulatory relationship between miR-525-5p and CSDE1 mRNA was analyzed by complementation assay.(6)Functional complementarity experiments were performed to verify the regulatory mechanism among LINC01234,miR-525-5p and CSDE1 mRNA.The downstream target genes of miR-525-5p involved in cell signaling pathways were analyzed by using the KEGG signaling pathway enrichment.Western blot was used to detect the relevant protein expression levels of PI3K/AKT signaling pathway in control and experimental groups.CCK-8 assay and flow cytometry were used to detect the effects of LINC01234 supplemented with PI3 K inhibitor on cell proliferation and apoptosis.Results(1)FISH localization analysis showed that LINC01234 was mainly localized in the cytoplasm of MCF-7 and MDA-MB-468 cell lines.Bioinformatics predicted that LINC01234 had binding sites with miR-525-5p.(2)Compared with patinets’ adjacent normal tissues,cancer tissues had significantly lower expression of miR-525-5p(P < 0.001).It was negative correlation between the expression level of miR-525-5p and LINC01234(r =-0.2620).Compared with MCF-10 A cell line,miR-525-5p was significantly down-regulated in BC cell lines(SKBR-3,MCF-7HS-578 T,MDA-MB-468,T-47 D and MDA-MB-231)(P < 0.05).When the overexpression or knockdown of LINC01234 was acheieved in MCF-7 and MDA-MB-468 cell lines,the expression of miR-525-5p was significantly decreased in LINC01234 overexpression group,and the expression of miR-525-5p was significantly increased in LINC01234 knockdown group(P < 0.01).The results of dual luciferase reporter assay showed that only LINC01234-WT and miR-525-5p co-transfected group had decreased luciferase activity(P < 0.05).RNA pull-down results showed that only the biotin-labeled miR-525-5p sense strand group could detect the target fragment of LINC01234.The AGO2-RIP assay showed that LINC01234 and miR-525-5p were detected in the co-precipitated complex.(3)MiR-525-5p mimics and inhibitor were transfected into MCF-7 and MDA-MB-468 respectively.Compared with each control group,the cell proliferation,invasion and migration abilities were decreased in miR-525-5p mimics group,while increased in inhibitor group(P < 0.05).The number of G0/G1 phase cells were significantly increased,but S phase cells decreased in miR-525-5p mimics group.On the contrary,the number of G0/G1 phase cells were significantly decreased,but S phase cells increased in the miR-525-5p inhibitor group(P < 0.05).The apoptosis rate was increased in miR-525-5p mimics group,while decreased in miR-525-5p inhibitor group(P < 0.01).Complementation assay showed that miR-525-5p could attenuate the promoting effects of LINC01234 on the proliferation,cell cycle,invasion and migration of MCF-7 and MDA-MB-468 cell lines,as well as the inhibitory effects of cell apoptosis.(4)Bioinformatics predicted the target genes of miR-525-5p,which reminded that miR-525-5p had targeted binding sites with CSDE1 mRNA.The expression of CSDE1 was up-regulated in BC tissues compared with adjacent tissues.Compared with MCF-10 A cell line,CSDE1 mRNA and its protein levels were significantly up-regulated in BC cell lines(SKBR-3,MCF-7,HCC1806,HS-578 T,MDA-MB-468,T-47 D and MDA-MB-231)(P < 0.05).When the expression of miR-525-5p was up-regulated or down-regulated in MCF-7 and MDA-MB-468 cell lines,compared with each control group,the expression of CSDE1 mRNA and its protein was significantly decreased in the miR-525-5p overexpression group,and significantly increased in the miR-525-5p knockdown group(P < 0.01).The results of dual luciferase reporter assay showed that the luciferase activity of CSDE1-3’UTR-WT and miR-525-5p co-transfected group was decreased(P < 0.01).RNA pull-down results showed that only biotin-labeled miR-525-5p sense strand group was able to detect the target fragment of CSDE1 mRNA.AGO2-RIP assay showed that CSDE1 mRNA and miR-525-5p could be detected in the co-precipitated complex.(5)pc DNA-CSDE1 and siRNA-CSDE1 were successfully transfected into MCF-7 and MDA-MB-468 cell lines.The abilities of cell proliferation,invasion and migration were enhanced in pc DNA-CSDE1 group,while weakened in siRNA-CSDE1 group(P < 0.05).The number of G0/G1 phase cells were significantly decreased,but S phase cells were increased in pc DNA-CSDE1 group,while the number of G0/G1 phase cells were significantly increased,but S phase cells were decreased in siRNA-CSDE1 group(P < 0.05).The apoptosis rate was decreased in pc DNA-CSDE1 group,while significantly increased in siRNA-CSDE1group(P < 0.01).Complementation assay showed that CSDE1 could attenuate the inhibitory effects of miR-525-5p on the cell proliferation,cell cycle,invasion and migration of MCF-7 and MDA-MB-468 cell lines,as well as the promoting effects of miR-525-5p on cell apoptosis.(6)When the overexpression or knockdown of LINC01234 was acheieved in MCF-7 and MDA-MB-468 cell lines,compared with each control group,the expression of CSDE1 mRNA and its protein was significantly increased in LINC01234 overexpression group,and significantly decreased in LINC01234 knockdown group(P < 0.01).In the rescue experiment,the expression of CSDE1 mRNA and its protein in miR-525-5p mimics group was significantly lower than the control group,and the expression of CSDE1 was recovered after co-transfecting with miR-525-5p mimics and pc DNA-LINC01234(P < 0.01).CCK-8 assay and flow cytometry results showed that compared with the control group,the cell proliferation activity of miR-525-5p mimics group was decreased,and the cell apoptosis was increased,which was reversed after co-transfecting of miR-525-5p mimics and pc DNA-LINC01234 into cells(P < 0.01).KEGG showed that the target mRNAs of miR-525-5p were related with the PI3K/AKT pathway.After transfecting si NRALINC01234 or pc DNA-LINC01234 in MCF-7 and MDA-MB-468 cell lines,compared with each control group,the expression of PTEN was significantly decreased,but PI3K-p85(Y607)and p-AKT increased in pc DNA-LINC01234 group.On the contrary,the expression of PTEN was significantly increased,but PI3K-p85(Y607)and p-AKT decreased in si NRA-LINC01234 group(P < 0.01).Co-transfecting miR-525-5p mimics or siRNA-CSDE1 or treatment of PI3 K inhibitor,LY294002,in overexpressing LINC01234 MCF-7 and MDA-MB-468 cell lines,the expression levels of PI3K-p85(Y607)and p-AKT were significantly decreased compared with the control group(P < 0.05).CCK-8 assay and flow cytometry results showed that PI3 K inhibitor LY294002 could inhibit LINC01234-induced cell proliferation and reverse the inhibition of LINC01234-mediated apoptosis(P < 0.01).Conclusion(1)LINC01234 is highly expressed in BC tissues and cell lines.The expression level of LINC01234 is correlated with tumor size in BC patients.Bioinformatics analysis indicates that the expression of LINC01234 is associated with poor prognosis.(2)LINC01234 inhibits the apoptosis and promotes the proliferation,cell cycle progression,invasion and migration of BC cell lines.(3)LINC01234 is a ceRNA targeting miR-525-5p to mediate the upregulation of CSDE1,resulting in inhibiting the expression of PTEN and activating PI3K/AKT signaling pathway to promote BC progression.更多还原