LncRNA SNHG6调节miR-101-3p/SEPT2轴对乳腺癌细胞恶性生物学行为的影响

Influence of LncRNA SNHG1 on malignant biological behavior of breast cancer cells by regulating miR-101-3p/SEPT2 axis

目的探究长链非编码RNA小核仁RNA宿主基因6(LncRNA SNHG6)对乳腺癌(BC)细胞恶性生物学行为的影响及其分子机制。方法qRT-PCR和Western blot检测人正常乳腺上皮细胞以及不同人BC细胞中SNHG6、miR-101-3p、SEPT2的mRNA以及SEPT2蛋白表达。选择MDA-MB-231细胞进行转染实验,分为对照组、si-NC组、si-SNHG6组、si-SNHG6+miR inhibitor-NC组、si-SNHG6+miR-101-3p inhibitor组。转染培养48 h后,qRT-PCR和Western blot检测MDA-MB-231细胞中SNHG6、miR-101-3p、SEPT2的mRNA以及SEPT2蛋白表达;CCK-8试剂盒检测细胞存活率;克隆形成实验检测细胞克隆数量;流式细胞术检测细胞凋亡率;Transwell小室检测细胞迁移和侵袭能力;双荧光素酶活性实验分别验证miR-101-3p和SNHG6、SEPT2的靶向关系。结果BC细胞系中MDA-MB-231细胞中SNHG6、SEPT2mRNA及蛋白水平高于其他细胞,miR-101-3p表达水平低于其他细胞(P<0.05)。转染si-SNHG6降低MDA-MB-231细胞中SNHG6、SEPT2的表达,提高miR-101-3p的表达,同时降低细胞的存活率、克隆数量、细胞迁移和侵袭数量,升高细胞的凋亡率(P<0.05)。双荧光素酶活性实验验证了SNHG6、SEPT2均可与miR-101-3p结合。结论敲低SNHG6能够上调miR-101-3p表达,下调SEPT2的表达,抑制BC细胞活性、迁移和侵袭,促进BC细胞凋亡。

Objective To investigate the influence of long non-coding RNA small nucleolar RNA host gene 6(LncRNA SNHG6)on the malignant biological behavior of breast cancer(BC)cells and its molecular mechanism.Methods QRT-PCR and Western blot were applied to detect the mRNA expression of SNHG6,miR-101-3p and SEPT2,and the protein expression of SEPT2 in human normal breast epithelial cells and different human BC cells.MDA-MB-231 cells were selected for transfection experiment,and they were grouped into control group,si-NC group,si-SNHG6 group,si-SNHG6+miR inhibitor-NC group,and si-SNHG6+miR-101-3p inhibitor group.After 48 h of transfection,qRT-PCR and Western blot were applied to detect the mRNA expressions of SNHG6,miR-101-3p and SEPT2,and protein expressions of SEPT2 in MDA-MB-231 cells.CCK-8 kit was used to detect cell viability.Clone formation assay was used to detect the number of cell clones.Flow cytometry was applied to detect the rate of apoptosis.Transwell chamber was used to detect cell migration and invasion,and dual luciferase activity assay was applied to verify the targeting relationship between miR-101-3p,SNHG6 and SEPT2.Results The mRNA and protein levels of SNHG6 and SEPT2 in MDA-MB-231 cells of BC cell line were higher than those of other cells,and the expression level of miR-101-3p was lower than that of other cells(P<0.05).Transfection of si-SNHG6 decreased the expression of SNHG6 and SEPT2 in MDA-MB-231 cells,increased the expression of miR-101-3p,decreased cell viability,the number of clones,the number of cell migration and invasion,and increased cell apoptosis rate(P<0.05).The dual luciferase activity assay verified that both SNHG6 and SEPT2 could bind to miR-101-3p.Conclusion Knockdown of SNHG6 can up-regulate the expression of miR-101-3p,down-regulate the expression of SEPT2,inhibit the activity,migration and invasion of BC cells,and promote the apoptosis of BC cells.