METHODS: LINC00022 expression in CRC tissues was analyzed by using the TNMplot software. LINC00022 expression in CRC cells was measured by quantitative real-time PCR. The effects of LINC00022 on the malignant behaviors of CRC cells were detected by a series of in vitro and in vivo experiments. Dual-luciferase assays were used to verify the targeting relationship between LINC00022 and miR-375-3p and between miR-375-3p and Forkhead box F1 (FOXF1), followed by the rescue experiment.
RESULTS: LINC00022 was highly expressed in CRC tissues compared with paired para-carcinoma tissues (n = 41). CRC cells with LINC00022 knockdown exhibited decreased cell proliferation, migration, and invasion abilities but increased apoptosis accompanied by decreased protein levels of c-Myc, cyclin D1, cleaved caspase 3, cleaved poly(ADP-ribose) polymerase, matrix metalloproteinase (MMP) 2, and MMP9. Additionally, LINC00022 downregulation in CRC cells suppressed the tube formation of human umbilical vein endothelial cells (HUVECs) as evidenced by decreased vascular endothelial growth factor A levels in LINC00022-silenced cells. The inhibitory effect of LINC00022 knockdown on tumor growth was also observed in an in vivo model. Conversely, LINC00022 overexpression showed that opposite effect. We further demonsrtaed that LINC00022 could upregulate FOXF1 expression through sponging miR-375-3p. Moreover, miR-375-3p knockdown reversed the effects of LINC00022 down-regulation.
CONCLUSIONS: LINC00022 may up-regulate FOXF1 expression via competitively binding miR-375-3p, thereby promoting the development of CRC.
方法:用TNMplot软件分析CRC组织中LINC00022的表达。通过定量实时PCR测量CRC细胞中的LINC00022表达。通过一系列的体内外实验检测到LINC00022对CRC细胞恶性行为的影响。使用双荧光素酶实验来验证LINC00022与miR-375-3p之间以及miR-375-3p与叉头盒F1(FOXF1)之间的靶向关系。随后是救援实验。
结果:LINC00022在CRC组织中比成对的癌旁组织高表达(n=41)。具有LINC00022敲低的CRC细胞表现出降低的细胞增殖,迁移,和侵袭能力,但伴随着c-Myc蛋白水平降低的细胞凋亡增加,细胞周期蛋白D1,裂解的半胱天冬酶3,裂解的聚(ADP-核糖)聚合酶,基质金属蛋白酶(MMP)2和MMP9。此外,CRC细胞中的LINC00022下调抑制人脐静脉内皮细胞(HUVEC)的管形成,如LINC00022沉默细胞中血管内皮生长因子A水平降低所证明。在体内模型中还观察到LINC00022敲低对肿瘤生长的抑制作用。相反,LINC00022过表达显示相反的效果。我们进一步证实LINC00022可以通过海绵化miR-375-3p上调FOXF1表达。此外,miR-375-3p敲低逆转了LINC00022下调的作用。
结论:LINC00022可能通过竞争性结合miR-375-3p上调FOXF1表达,从而促进CRC的发展。