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Expression and purification of recombinant human CCL5 and its biological characterization

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Abstract

C-C motif chemokine ligand 5 (CCL5) is crucial in the tumor microenvironment. It has been previously reported to act as a key role in tumor invasion and metastasis. However, the function of exogenous CCL5 in ovarian cancer has not been well-characterized. The present study attempted to express and purify recombinant CCL5 protein and investigate the exogenous CCL5 in ovarian cancer cell proliferation. The human CCL5 was amplified and inserted into the pET-30a vectors for prokaryotic expression in Escherichia coli BL21. Soluble His-CCL5 was successfully expressed with 0.1 mmol/L of isopropyl-β-D-1-tiogalactopiranoside at 25 ℃ and purified by affinity chromatography. Additionally, methyl thiazolyl tetrazolium (MTT) assay demonstrated that CCL5 promotes ovarian cancer cell proliferation; increases the phosphorylation levels of extracellular-signal-regulated kinase and mitogen-activated protein kinase/ERK kinase, and increases the mRNA levels of Jun, NF-κB2, Nras, Relb, and Traf2. Furthermore, treatment with the MEK inhibitor reduced the Jun, NF-κB2, and Traf2 mRNA levels, indicating that exogenous CCL5 increased ovarian cancer cell proliferation, through MEK/ERK pathway activation, and Jun, NF-κB2, and Traf2 expression. The present study provided primary data for further studies to discover more CCL5 functions in ovarian cancer.

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Acknowledgements

This research was supported by grants from the Scientific and Technological Research Project of Henan Province (No. 212102310898), the Key scientific research project of Henan Province (No. 22A180017), and the research start-up fund to topnotch talents of Henan Agricultural University (No. 30500424).

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Authors and Affiliations

  1. Henan Provincial People’s Hospital/People’s Hospital of Zhengzhou University, 7 Weiwu Road, Jinshui District, 450000, Zhengzhou, China

    Zhenling Ma & Li Xiang

  2. College of Life Sciences, Henan Agricultural University, 450002, Zhengzhou, China

    Zhenling Ma, Jiajia Zhang, Lei Wang, Yiying Liu, Yunpeng Wang, Wei Liu, Guozhen Xing, Kun Cheng & Wenming Zheng

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  1. Zhenling Ma
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  2. Jiajia Zhang
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  3. Lei Wang
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  4. Yiying Liu
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  5. Yunpeng Wang
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  6. Wei Liu
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  7. Guozhen Xing
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  8. Kun Cheng
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  9. Wenming Zheng
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  10. Li Xiang
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Contributions

Z.M. and L.X. designed the study. Z.M., J.Z. and L.W. performed the experiments. Y.L., G.X. and K.C. performed systematic research of the literature. Z.M. wrote the paper. W.Z. and L.X. revised it.

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Correspondence to Li Xiang.

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Ma, Z., Zhang, J., Wang, L. et al. Expression and purification of recombinant human CCL5 and its biological characterization. Protein J 41, 337–344 (2022). https://doi.org/10.1007/s10930-022-10047-8

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  • DOI: https://doi.org/10.1007/s10930-022-10047-8

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