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Purification of recombinant human chemokine CCL2 in E. coli and its function in ovarian cancer.

ABSTRACT: Chemokine (CC-motif) ligand 2 (CCL2) is an inflammatory cytokine that regulates the infiltration and migration of monocytes. It is highly expressed by both tumor and stromal cells and has been associated with tumorigenesis. However, the effect of the exogenous administration of CCL2 on ovarian cancer remains largely unknown. In this report, we attempted to establish an expression system in Escherichia coli to produce recombinant hCCL2. The recombinant plasmid containing the hCCL2 cDNA was prepared using the prokaryotic-expression plasmid pGEX-5X-3 and transformed into E. coli BL21. GST-hCCL2 was successfully induced by 0.1 mmol/L IPTG at 20 °C for 6 h, and the recombinant protein was purified using affinity chromatography. The purified protein was identified by SDS-PAGE and Western Blot. In vitro experiments revealed that rhCCL2 promoted the proliferation of ovarian cancer cells and increased the levels of phosphorylation of MEK and ERK1/2, and the levels of JUN, RELB and NF-κB2 mRNA. Furthermore, inhibition of ERK signaling by treatment with PD98059 decreased ovarian cancer cell proliferation and levels of JUN, RELB, and NF-κB2 mRNA, indicating that exogenous rhCCL2 increased the proliferation of ovarian cancer cells, partially by activating the MAPK/ERK pathway, and by targeting JUN, RELB, and NF-κB2. Our study uncovered a promoting role of exogenous CCL2 on ovarian cancer cell proliferation through the MAPK/ERK signaling pathway, which may facilitate the discovery of more potential roles of CCL2 in ovarian cancer.

Supplementary information

The online version contains supplementary material available at 10.1007/s13205-020-02571-0.

SUBMITTER: Liu W

PROVIDER: S-EPMC7778648 | biostudies-literature |

REPOSITORIES: biostudies-literature

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