miR-449通过抑制CXCL5表达调控胃癌进程机制研究

Study on the mechanism of miR-449 regulating gastric cancer progression by inhibiting the expression of CXCL5

目的探讨miR-449在胃癌中的作用情况及其深入机制,为胃癌的治疗提供可靠的数据支撑。方法基于微阵列分析筛选异常表达的miRNA。通过qRT-PCR观察到组织标本和细胞中miR-449的表达,而通过qRT-PCR和Western blotting证实了胃癌组织中CXCL5的表达。通过TargetScan预测了miR-449在CXCL53′UTR上的结合位点。miR-449和CXCL5之间的关系通过双荧光素酶报告基因测定来确定。miR-449/CXCL5对胃癌细胞活力、迁移性、侵袭性和凋亡能力的影响通过MTT测定和克隆形成、划痕测定、Transwell测定和流式细胞术测定进行验证。构建异种移植小鼠模型以证明miR-449/CXCL5对体内肿瘤生长的影响。结果与癌旁组织及正常细胞相比,胃癌组织和癌细胞中miR-449表达下调(P<0.05);miR-449的过表达抑制了胃癌细胞的活力、迁移和细胞周期,同时显著促进了细胞凋亡且差异均有统计学意义(P<0.05);CXCL5表达的上调促进了细胞活力、迁移和侵袭性并抑制了胃癌细胞凋亡且差异均有统计学意义(P<0.05);miR-449靶向并下调CXCL5表达,从而抑制胃癌细胞的增殖和转移并刺激细胞凋亡(P<0.05);体内实验证明,与NC组比较,miR-449能够通过CXCL5抑制肿瘤生长且差异有统计学意义(P<0.05)。结论miR-449具有肿瘤抑制剂的作用,通过调控CXCL5的表达抑制胃癌细胞活力和转移,促进细胞凋亡。

Objective To explore the role of miR-449 in gastric cancer and its in-depth mechanism,so as to provide reliable data support for the treatment of gastric cancer.Methods Screening of abnormally expressed miRNAs based on microarray analysis.The expression of miR-449 in tissue specimens and cells was observed by qRT-PCR,and the expression of CXCL5 in gastric cancer tissue was confirmed by qRT-PCR and Western blotting.The binding site of miR-449 on CXCL53′UTR was predicted by TargetScan.The relationship between miR-449 and CXCL5 was determined by dual luciferase reporter gene assay.The effect of miR-449/CXCL5 on the viability,migration,invasiveness and apoptosis of gastric cancer cells was verified by MTT assay and clone formation,scratch assay,Transwell assay and flow cytometry assay.A xenograft mouse model was constructed to demonstrate the effect of miR-449/CXCL5 on tumor growth in vivo.Results Compared with adjacent tissues and normal cells,the expression of miR-449 in gastric cancer tissues and cells was down-regulated(P<0.05);the overexpression of miR-449 inhibited the vitality,migration and cell cycle of gastric cancer cells,while significantly promoting.The difference in cell apoptosis was statistically significant(P<0.05);the up-regulation of CXCL5 expression promoted cell viability,migration and invasiveness,and inhibited cell apoptosis in gastric cancer,and the difference was statistically significant(P<0.05);miR-449 targeted and down-regulated the expression of CXCL5,thereby inhibiting the proliferation and metastasis of gastric cancer cells and stimulating apoptosis(P<0.05);in vivo experiments showed that miR-449 could inhibit tumor growth through CXCL5 and the difference was statistically significant compared with the NC group(P<0.05).Conclusion miR-449 has a tumor suppressor effect.It can inhibit the viability and metastasis of gastric cancer cells by regulating the expression of CXCL5,and promote cell apoptosis.