LINC00294通过AKT/mTOR/HIF-1a通路促进乳腺癌细胞的增殖、侵袭与迁移

LINC00294 promotes proliferation,invasion,and migration of breast cancer cells via AKT/mTOR/HIF-1αpathway

目的探讨长链非编码RNA(long non-coding RNA,LncRNA)LINC00294在乳腺癌组织和细胞中的表达变化,及其对乳腺癌细胞生物学特性的影响和可能的机制。方法利用GEO数据库资源(GSE45827)中的41例乳腺癌组织和11例癌旁组织转录组测序数据分析差异基因,采用qRT-PCR实验检测20对来自本院2021-2022年确诊的乳腺癌患者的癌组织和癌旁组织,以及正常人乳腺上皮细胞MCF-10A和人乳腺癌细胞MCF-7、SK-BR-3和MDA-MB-231中LINC00294的表达;干扰LINC00294后采用CCK-8检测细胞增殖变化;采用流式细胞术、Western blot检测细胞凋亡和相关蛋白表达变化;采用划痕实验和Transwell检测细胞迁移和侵袭;裸鼠原位成瘤实验检测干扰LINC00294对体内肿瘤生长的影响;采用Western blot检测干扰LINC00294后AKT/mTOR通路相关蛋白p-mTOR、mTOR、p-AKT、AKT及其下游靶分子缺氧诱导因子-1(hypoxia inducible factor-1α,HIF-1α)的表达情况。结果通过GEO数据库筛选出差异表达的基因LINC00294;LINC00294在乳腺癌组织和乳腺癌细胞MCF-7、SK-BR-3和MDA-MB-231中表达显著增高(P<0.05);干扰LINC00294后MCF-7细胞增殖能力下降、凋亡相关蛋白表达增加、侵袭和迁移能力下降(P<0.05)。干扰HIF-1α可以抑制LINC00294的表达(P<0.05);干扰LINC00294后AKT/mTOR信号通路中p-AKT和p-mTOR蛋白及其下游靶分子HIF-1α的表达减少(P<0.05)。结论乳腺癌中高表达的LINC00294可能通过AKT/mTOR/HIF-1α信号通路影响乳腺癌的增殖、侵袭和迁移等,LINC00294可能与乳腺癌的发展密切相关。

Objective To explore the changes in the expression of long non-coding RNA(LncRNA)LINC00294 in breast cancer tissues and cells,and investigate its effects and the possible mechanisms on the biological properties of breast cancer cells.Methods The differentially expressed genes(DEGs)were analyzed using transcriptome sequencing data from 41 breast cancer tissues and 11 para-cancerous tissues from the Gene Expression Omnibus(GEO)database(GSE45827).Then qRT-PCR assay was used to measure the expression of LINC00294 in 20 pairs of breast cancer and para-cancerous tissues from the breast cancer patients diagnosed in our department,as well as in the normal human breast epithelial MCF-10A cells and the human breast cancer cell lines MCF-7,K-BR-3 and MDA-MB-231.After MCF-7 cells were transfected with lentiviral vector of si-LINC to interfere LINC00294 expression,CCK-8 assay was used to detect cell proliferation,flow cytometry and Western blotting were employed to measure cell apoptosis and expression of related proteins,cell scratch assay and Transwell assay were applied to observe cell migration and invasion.Then,the effect of interference with LINC00294 on tumor growth in nude mice model of in situ subcutaneous tumorigenesis was observed.The expression of AKT/mTOR pathway-related proteins p-mTOR,mTOR,p-AKT,AKT,and its downstream target molecule HIF-1αwas detected by Western blotting.Results LncRNA LINC00294 was screened out as a DEG in breast cancer tissues from the GEO database.It was also highly expressed in breast cancer tissues and MCF-7,SK-BR-3 and MDA-MB-231 cells(P<0.05).Interference of LINC00294 resulted in reduced proliferation,enhanced expression of apoptosis-related proteins,and decreased invasion and migration abilities in MCF-7 cells(P<0.05).The expression of LINC00294 was inhibited by HIF-1αinterference(P<0.05),and LINC00294 interference decreased the expression of p-AKT,p-mTOR,mTOR and its downstream target molecule HIF-1αin the AKT/mTOR signaling pathway(P<0.05).Conclusion High expression of LINC00294 in breast cancer may affect the proliferation,invasion,and migration of breast cancer through the AKT/mTOR/HIF-1αsignaling pathway,and LINC00294 may be closely associated with the pathogenesis of breast cancer.