靶向下调 CXCR7 表达对低氧环境下肾癌细胞活力、凋亡和侵袭的影响。

Effect of MicroRNA-218 on the viability, apoptosis and invasion of renal cell carcinoma cells under hypoxia by targeted downregulation of CXCR7 expression.

机构信息

Department of General Surgery, The Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi 330006, China; Department of General Surgery, Jiangxi Provincial Children's Hospital, Nanchang, Jiangxi 330006, China.

Department of Urology, The Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi 330006, China.

出版信息

Biomed Pharmacother. 2016 May;80:213-219. doi: 10.1016/j.biopha.2016.03.011. Epub 2016 Mar 28.

DOI:10.1016/j.biopha.2016.03.011

PMID:27133059

Abstract

OBJECTIVE

To investigate the effect of microRNA-218 on the viability, apoptosis and invasion of renal cell carcinoma cells under hypoxia by targeted regulation of expression of chemokine receptor 7 (CXCR7).

METHODS

The expression of miR-218 in renal cell carcinoma cell lines under normal and hypoxia conditions, as well in normal renal tubular epithelial cells (HK2) was measured using RT-PCR. MiR-218 mimic and NC were transfected into renal cell carcinoma cell line ACHN using Lipofectamine™ 2000. The expression of miR-218 was analyzed using RT-PCR. The viability, apoptosis, migration and invasion of the transfected cells were assayed using the MTT assay, flow cytometry and transwell assays. The expression of CXCR7 was assayed using RT-PCR and Western blot. Luciferase reporter was used to verify the downstream target of miR-218.

RESULTS

The expression of miR-218 was lower than in renal cell carcinoma cell lines ACHN, 769-p and Caki-1 that in HK-2. The expression of miR-218 in the renal carcinoma cell lines was lower under hypoxia than under normal oxygen conditions. The expression of miR-218 in ACHN cells under normal and hypoxic conditions was significantly increased after transfection with miR-218 mimic. Compared with NC transfected cells under normal oxygen condition, the mimic-transfected cells had reduced viability, migration ability and invasion ability, and increased apoptosis, and mimic transfected-cells under hypoxia had significantly reduced viability, migration ability and invasion ability, and increased apoptosis. Overexpression of miR-218 mimic resulted in significant reduction in the expression of CXCR7 at protein and mRNA levels under normal and hypoxic conditions. Luciferase reporter assay confirmed that CXCR7 is the target protein of miR-218.

CONCLUSION

Up-regulation of miR-218 expression in renal cell carcinoma under hypoxia can result in significant and targeted down-regulation of CXCR7 expression, which could reduce cell viability, migration and invasion ability and induce apoptosis in the cancer cells.

摘要

目的

通过靶向调控趋化因子受体 7（CXCR7）的表达，研究微小 RNA-218 对缺氧状态下肾癌细胞活力、凋亡和侵袭的影响。

方法

采用 RT-PCR 检测正常和缺氧条件下肾癌细胞系及正常肾小管上皮细胞（HK2）中 miR-218 的表达。采用 Lipofectamine™ 2000 将 miR-218 模拟物和对照物（NC）转染入肾癌细胞系 ACHN。采用 RT-PCR 分析 miR-218 的表达。采用 MTT 法、流式细胞术和 Transwell 法检测转染细胞的活力、凋亡、迁移和侵袭能力。采用 RT-PCR 和 Western blot 检测 CXCR7 的表达。采用荧光素酶报告基因验证 miR-218 的下游靶基因。

结果

miR-218 的表达在 HK-2 中低于肾癌细胞系 ACHN、769-p 和 Caki-1。肾癌细胞系在缺氧条件下的 miR-218 表达低于正常氧条件。在正常氧条件下，与 NC 转染细胞相比，miR-218 模拟物转染细胞的活力、迁移能力和侵袭能力降低，凋亡增加；在缺氧条件下，与 NC 转染细胞相比，miR-218 模拟物转染细胞的活力、迁移能力和侵袭能力显著降低，凋亡增加。在正常和缺氧条件下，过表达 miR-218 模拟物可导致 CXCR7 蛋白和 mRNA 表达显著降低。荧光素酶报告基因证实 CXCR7 是 miR-218 的靶蛋白。