LncRNA ZEB1-AS1调节miR-429/PDX-1轴对结肠癌细胞增殖、凋亡和上皮间质转化的影响

Influences of LncRNA ZEB1-AS1 on proliferation,apoptosis and epithelial-mesenchymal transition of colon cancer cells by regulating the miR-429/PDX-1 axis

目的:探究长链非编码RNA E盒锌指蛋白1反转录本1(LncRNA ZEB1-AS1)调节miR-429/胰腺十二指肠同源盒基因-1(PDX-1)轴对结肠癌细胞增殖、凋亡以及上皮间质转化(EMT)的影响。方法:q RT-PCR法测定细胞LncRNA ZEB1-AS1、miR-429、PDX-1 m RNA表达水平;将SW480细胞分为对照组、si-NC组、si-LncRNA ZEB1-AS1组、si-LncRNA ZEB1-AS1+inhibitor NC组、si-LncRNA ZEB1-AS1+miR-429 inhibitor组、si-LncRNA ZEB1-AS1+miR-429 inhibitor+sh NC组、si-LncRNA ZEB1-AS1+miR-429 inhibitor+sh PDX-1组,检测SW480细胞增殖、凋亡、EMT相关蛋白、紧密连接蛋白以及PDX-1表达;双荧光素酶报告基因实验验证LncRNA ZEB1-AS1、PDX-1与miR-429关系。结果:与人正常结肠上皮细胞相比,NCM-460、HCT116、SW620、CT26、SW480中LncRNA ZEB1-AS1、PDX-1 m RNA表达升高,miR-429表达降低,其中SW480细胞升高或降低最为显著(P<0.05);与对照组相比,si-LncRNA ZEB1-AS1组SW480细胞增殖抑制率、凋亡率及E-cad、ZO-1、Claudin-1表达升高,N-cad、Vimentin表达降低(P<0.05);miR-429 inhibitor下调可逆转LncRNA ZEB1-AS1沉默对SW480细胞的作用;si-LncRNA ZEB1-AS1与miR-429 inhibitor基础上沉默PDX-1可恢复LncRNA ZEB1-AS1沉默对SW480细胞的作用;miR-429与LncRNA ZEB1-AS1以及PDX-1均有靶向关系。结论:沉默ZEB1-AS1可能通过促进miR-429表达,抑制PDX-1表达,进而抑制SW480细胞增殖与EMT,并促进细胞凋亡。

Objective:To explore the influences of long non-coding RNA zinc finger E-box binding homeobox 1 antisense 1(LncRNA ZEB1-AS1)on the proliferation,apoptosis and epithelial-mesenchymal transition(EMT)of colon cancer cells through regulation of the miR-429/pancreatoduodenal homeobox-1(PDX-1)axis.Methods:The expression levels of LncRNA ZEB1-AS1,miR-429,and PDX-1 mRNA in cells were determined by qRT-PCR;SW480 cells were divided into control group,si-NC group,si-LncRNA ZEB1-AS1 group,and si-LncRNA ZEB1-AS1+inhibitor NC,si-LncRNA ZEB1-AS1+miR-429 inhibitor group,si-LncRNA ZEB1-AS1+miR-429 inhibitor+sh NC,si-LncRNA ZEB1-AS1+miR-429 inhibitor+sh PDX-1 group,detection The proliferation of SW480 cells,the expression of apoptosis EMT-related proteins,zonula occludes protein and PDX-1 were measured;the dual-luciferase reporter gene experiment verified the relationship between LncRNA ZEB1-AS1,PDX-1 and miR-429.Results:Compared with human normal colon epithelial cells,the mRNA expressions of LncRNA ZEB1-AS1 and PDX-1 in NCM-460,HCT116,SW620,CT26 and SW480 were increased,and the expression of miR-429 was decreased,among which the increase or decrease in SW480 cells was the most significant(P<0.05),compared with the control group,the SW480 cell proliferation inhibition rate,apoptosis rate,E-cad,ZO-1,and Claudin-1 were increased,and the expressions of N-cad and Vimentin were decreased in the si-LncRNA ZEB1-AS1 group(P<0.05),miR-429 inhibitor down-regulation can reverse the effect of LncRNA ZEB1-AS1 silencing on SW480 cells,Silencing PDX-1 on the basis of si-LncRNA ZEB1-AS1 and miR-429 inhibitor can restore the effect of LncRNA ZEB1-AS1 silencing on SW480 cells,miR-429 has a targeting relationship with LncRNA ZEB1-AS1 and PDX-1,miR-429 is associated with LncRNA ZEB1-AS1 and PDX-1 has a targeting relationship.Conclusion:Silencing ZEB1-AS1 may promote the expression of miR-429 and inhibit the expression of PDX-1,thereby inhibiting the proliferation and EMT of SW480 cells,and promoting cell apoptosis.