摘要
目的建立结直肠癌KRAS突变细胞DKO-1的miR-139-5p功能获得模型,探讨miR-139-5p对KRAS突变结直肠癌细胞恶性表型的影响及可能的分子机制。方法利用real-time PCR检测miR-139-5p在KRAS突变同源细胞系中表达;通过瞬时转染建立miR-139-5p功能获得细胞模型;通过绘制MTT生长曲线和集落形成实验检测miR-139-5p对DKO-1细胞生长增殖的影响;通过细胞划痕实验和Transwell侵袭实验检测miR-139-5p对DKO-1细胞生长迁移侵袭的影响;通过生物信息学分析miR-139-5p的下游靶基因,利用双荧光酶报告基因实验验证miR-139-5p与靶基因的直接结合情况。结果 miR-139-5p在KRAS突变结直肠癌细胞中表达降低(P<0.01);过表达miR-139-5p能够抑制DKO-1细胞的生长能力(P<0.01)、集落形成能力(P<0.01)、迁移和侵袭能力(P<0.01);miR-139-5p能够直接结合FOS基因的3'-UTR区(P<0.01)。结论 miR-139-5p能够抑制DKO-1细胞的增殖、迁移和侵袭能力,其机制可能与抑制FOS基因表达有关。
Objective To explore the effect of miR-139-5p on the malignant phenotype of KRAS mutant colon cancer cells and its po- tential molecular mechanisms. Methods Real-time PCR was performed to detect the expression of miR-139-5p in KRAS isogenic cell lines. Gain-of-function cell model was established by transient transfection with miR-139-5p mimics. MTT growth curve and colony for- mation assay were conducted to determine the effect of miR-139-5p on growth and proliferation of DKO-1 cells. Would healing and Tran- swell invasion assays were used to investigate the effect of miR-139-Sp on migration and invasion of DKO-1 cells. Bioinformatic analysis was employed to predict downstream target genes of miR-139-5p. Dual luciferase reporter assay was used to verify direct interaction between miR-139-5p and its target gene. Results The miR-139-5p was down-regulated in KRAS mutant colon cancer cells （P 〈 0.01 ）. The miR-139-5p overexpression significantly inhibited the proliferation, migration and invasion of DKO-1 cells （P 〈 0. 01 ）. The miR-139-5p was directly bound to the 3＇ - UTR of the FOS gene. Conclusion The miR-139-5p could inhibit the proliferation, migra- tion and invasion of DKO-1 cells via inhibiting expression of FOS gene.
出处
《山西医科大学学报》
CAS
2017年第4期322-328,共7页
Journal of Shanxi Medical University
基金
国家自然科学基金资助项目(81602641
81572929)
