AMD3100 联合雷公藤红素抑制人 U2OS 骨肉瘤细胞的增殖、侵袭和转移并诱导其凋亡。

AMD3100 combined with triptolide inhibit proliferation, invasion and metastasis and induce apoptosis of human U2OS osteosarcoma cells.

机构信息

Department of Pediatrics, Hangzhou First People's Hospital, Nanjing Medical University, Hangzhou 310006, Zhejiang, China.

Department of Clinical Laboratory, Hangzhou First People's Hospital, Nanjing Medical University, Hangzhou 310006, Zhejiang, China.

出版信息

Biomed Pharmacother. 2017 Feb;86:677-685. doi: 10.1016/j.biopha.2016.12.055. Epub 2016 Dec 27.

DOI:10.1016/j.biopha.2016.12.055

PMID:28038429

Abstract

BACKGROUND

Osteosarcoma (OS) mainly occurs in children and adolescents, and has a high propensity for lung metastasis. Little is known about the role of SDF-1/CXCR4 axis in OS progression. AMD3100 is a specific CXCR4 antagonist. Triptolide can induce apoptosis and proliferation inhibition in various cancer cell lines.

OBJECTIVE

This work aimed to investigate the effects of AMD3100 plus triptolide on the proliferation, apoptosis, invasion and metastasis of OS cells.

METHODS

The expression levels of SDF-1 and CXCR4 in five OS cell lines was analyzed by qRT-PCR, western blotting and ELISA assays. The effect of AMD3100 and triptolide on the proliferation, apoptosis and invasion of U2OS cells was evaluated by CCK-8, flow cytometry and transwell assay, respectively. Orthotopic intra-tibial growth and lung metastasis mouse model of OS were employed to evaluate the inhibition effect of AMD3100 and triptolide on primary OS growth and lung metastasis.

RESULTS

CXCR4 protein expression was detected in HOS-8603, MG-63, U2OS and 143B but not Saos2 cells, and all these cell lines expressed SDF-1. AMD3100 plus triptolide induced proliferation inhibition and apoptosis of U2OS cells, which was attributed to the downregulation of c-Myc, survivin, cyclin D1 and increased cleaved caspase-3 and PARP. AMD3100 and triptolide also suppressed SDF-1 induced invasion of CXCR4+ U2OS cells, which was validated by decreased expression of MMP-2 and 9, VEGF, m-Calpain and β-catenin. Moreover, the phosphorylation levels of Erk1/2, Akt and STAT3, as well as the nuclear translocation and phosphorylation of NF-κB p65 in U2OS cells were also reduced by AMD3100 and triptolide. In vivo, AMD3100 and triptolide significantly reduced primary tumor growth and lung metastasis of U2OS cells.

CONCLUSIONS

AMD3100 combined with triptolide can reduce proliferation and metastasis, and induce apoptosis of U2OS cells, which may be related to the Erk1/2, Akt, STAT3 and NF-κB pathways.

摘要

背景

骨肉瘤（OS）主要发生在儿童和青少年中，具有很高的肺转移倾向。目前对于 SDF-1/CXCR4 轴在 OS 进展中的作用知之甚少。AMD3100 是一种特异性 CXCR4 拮抗剂。雷公藤内酯醇可诱导多种癌细胞系凋亡和增殖抑制。

目的

本研究旨在探讨 AMD3100 联合雷公藤内酯醇对 OS 细胞增殖、凋亡、侵袭和转移的影响。

方法

通过 qRT-PCR、Western blot 和 ELISA 检测 5 种 OS 细胞系中 SDF-1 和 CXCR4 的表达水平。通过 CCK-8、流式细胞术和 Transwell 检测 AMD3100 和雷公藤内酯醇对 U2OS 细胞增殖、凋亡和侵袭的影响。采用原位胫骨内生长和肺转移 OS 小鼠模型评价 AMD3100 和雷公藤内酯醇对原发性 OS 生长和肺转移的抑制作用。

结果

在 HOS-8603、MG-63、U2OS 和 143B 细胞中检测到 CXCR4 蛋白表达，但在 Saos2 细胞中未检测到，所有这些细胞系均表达 SDF-1。AMD3100 联合雷公藤内酯醇诱导 U2OS 细胞增殖抑制和凋亡，这归因于下调 c-Myc、survivin、cyclin D1 和增加 cleaved caspase-3 和 PARP。AMD3100 和雷公藤内酯醇还抑制了 SDF-1 诱导的 CXCR4+U2OS 细胞侵袭，这通过 MMP-2 和 9、VEGF、m-Calpain 和 β-catenin 的表达下调得到验证。此外，AMD3100 和雷公藤内酯醇还降低了 U2OS 细胞中 Erk1/2、Akt 和 STAT3 的磷酸化水平，以及 NF-κB p65 的核转位和磷酸化。在体内，AMD3100 和雷公藤内酯醇显著降低了 U2OS 细胞的原发肿瘤生长和肺转移。