miR-488靶向NRSN2介导NF-κB信号通路在骨肉瘤增殖和侵袭中的作用

MicroRNA-488 regulates the proliferation and invasion of osteosarcoma by targeting Neurensin-2 and thus mediates the nuclear factor-kappaB signaling pathway

目的探究miR-488靶向NRSN2介导的NF-κB信号通路在骨肉瘤增殖和侵袭中的作用及相关机制。方法收集32例骨肉瘤患者肿瘤组织和癌旁组织并检测NRSN2和miR-488表达水平。miRNA-seq检测NRSN2高表达和NRSN2低表达肿瘤组织差异miRNA,Spearman相关系数分析miR-488与NRSN2相关性。在MG63骨髓瘤细胞中,通过荧光素酶报告基因实验检测miR-488与NRSN2靶向性。将si-NC、si-miR-488转染至MG63骨髓瘤细胞,分别加入NF-κB抑制剂(IKK-16),分为si-NC组、si-miR-488组和si-miR-488+IKK-16组。qRT-PCR检测细胞miR-488和NRSN2表达;Western blot检测细胞NRSN2和NF-κB(p65)蛋白表达;CCK8检测细胞增殖水平;Transwell检测细胞侵袭能力。结果NRSN2高表达肿瘤组织中有276个miRNA表达升高,294个miRNA表达降低(P<0.05)。与癌旁组织比较,肿瘤组织miR-488表达降低(P<0.01),NRSN2表达升高(P<0.01),miR-488表达与NRSN2表达呈负相关(P<0.05)。荧光素酶报告基因实验结果显示miR-488靶向NRSN2。与si-NC组比较,si-miR-488组miR-488表达降低(P<0.05),NRSN2和NF-κB(p65)表达升高(P<0.05),细胞增殖水平和侵袭能力增加(P<0.05)。与si-miR-488组比较,si-miR-488+IKK-16组NF-κB(p65)表达降低(P<0.05),细胞增殖水平和侵袭能力降低(P<0.05)。结论在骨肉瘤中,miR-488能够靶向NRSN2抑制NF-κB信号通路,从而降低肿瘤细胞增殖和侵袭的能力。

Objective To investigate the role of microRNA-488(miR-488)in regulating the proliferation and invasion of osteosarcoma by targeting Neurensin-2(NRSN2)and thus mediates the nuclear factor-kappaB(NF-κB)signaling pathway.Methods Tumor tissues and adjacent tissues of 32 osteosarcoma patients were collected to detect expression levels of NRSN2 and miR-488.Differentially expressed miRNAs in osteosarcoma tissues expressing high and low levels of NRSN2 were screened by miRNA-seq.The correlation between miR-488 and NRSN2 was analyzed by Spearman correlation coefficient.The targeting of miR-488 to NRSN2 was detected by dual-luciferase reporter assay in MG63 cells.MG63 cells were transfected with si-NC and si-miR-488,followed by the treatment of NF-κB inhibitor(IKK-16).The mRNA levels of miR-488 and NRSN2 in MG63 cells were measured by quantitative real-time polymerase chain reaction(qRT-PCR).The protein expressions of NRSN2 and NF-κB(p65)were detected by Western blot.Cell proliferation and invasion were detected by Cell Counting Kit-8(CCK-8)assay and Transwell assay,respectively.Results In osteocarcinoma tissues expressing high levels of NRSN2,there were 276 upregulated miRNAs and 294 downregulated miRNAs.Compared with those of adjacent tissues,miR-488 was downregulated and NRSN2 was upregulated in osteocarcinoma tissues,showing a negative correlation(P<0.05).Dual-luciferase reporter assay showed that miR-488 targeted NRSN2.Transfection of si-miR-488 significantly downregulated miR-488,upregulated NRSN2 and NF-κB(P65),and increased proliferative and invasive abilities(P<0.05),which were reversed by the treatment if IKK-16(P<0.05).Conclusion In osteosarcoma,miR-488 can target NRSN2 to inhibit NF-κB signaling pathway,then reducing the proliferation and invasion ability of tumor cells.