shRNA干扰长链非编码RNA UCA1对结肠癌细胞体外生长、运动能力及移植肿瘤形成的影响

Effects of shRNA Interfering with Long Non-Coding RNA UCA1 on in vitro Growth and Motility of Colon Cancer Cells,and Formation of Transplanted Tumor

目的研究shRNA干扰LncRNA尿路上皮癌抗原1(UCA1)对结肠癌细胞的体外生长、运动能力及移植瘤形成的影响.方法qRT-PCR检测UCA1、miR-185-5p表达情况;双荧光素酶报告实验检测靶向关系;Brdu染色检测细胞生长,Tranwell试验和细胞免疫荧光检测细胞运动;Western印迹检测Ki67、VEGF、MAPKAPK2、p-MAPKAPK2、HSP27、p-HSP27表达;皮下注射SW480细胞构建移植瘤模型,检测肿瘤重量、存活率、UCA1、miR-185相对表达量、Ki67、VEGF阳性表达率.结果UCA1在结肠癌细胞中高表达,miR-185-5p低表达(P<0.05);UCA1表靶向下调miR-185-5p;sh-UCA1能够显著抑制SW480细胞生长、运动、p38 MAPK通路蛋白表达(均P<0.05);与control组裸鼠比较,sh-UCA1组裸鼠瘤体重量、存活率、Ki67和VEGF蛋白阳性表达均显著降低(均P<0.05).结论UCA1可能通过靶向下调miR-185-5p激活p38 MAPK信号通路促进结肠癌SW480细胞系的细胞生长、运动能力,并促进体内肿瘤形成.

To explore the effects of shRNA interfering with LneRNA urothelial careinoma antigen 1(UCA1)on in vitro growth and motility of colon cancer cells,and formation of transplanted tumor.Methods The expression of UCA1 and miR-185-5p was detected by qRT-PCR.And their targeted relationship was detected by dual luciferase reporter assay.The cells growth was detected by Brdu staining,and cells movement by Tranwell assay and cellular immunofluorescence.The expression of Ki67,VEGF,MAPKAPK2,p-MAPKAPK2,HSP27 and p-HSP27 was detected by Western blotting.The subcutaneous injection of SW480 cells was conducted to construct the transplanted tumor model.The tumor weight,survival rate,relative expression levels of UCA1 and miR-185,and positive expression rates of Ki67 and VECF were detected.Results UCA1 was highly expressed,while miR-185-5p was lowly expressed in colon cancer cells(P<0.05).UCAI could down-regulate miR-185-5p.sh-UCA1 could significeantly inhibit the growth and movement of SW480 cells,and the expression of p38 MAPK pathway protein(P<0.05).Compared with control group,the tumor weight,survival rate,positive expression of Ki67 and VEGF proteins in sh-UCA1 group were significantly decreased(P<0.05).Conclusion UCAI may promote the growth and motility of colon cancer SW480 cell line by down-regulating miR-185-5p and activating p38 MAPK signaling pathway,and promote tumor formation in vivo.