Epithelial-stromal communication via CXCL1-CXCR2 interaction stimulates growth of ovarian cancer cells through p38 activation

Abstract

Purpose

Paracrine interactions with the stromal environment, including fibroblasts, may be important in the pathogenesis of ovarian cancer. Here, we evaluated the effect of conditioned media derived from ovarian fibroblasts (fibroblast-CMs) and their major cytokines on the growth of ovarian cancer cells, as well as the involvement of mitogen-activated protein kinases (MAPKs) and AKT in mediating this effect.

Methods

Ovarian cancer cells were cultured in serum-free media (SF), or conditioned media of fibroblasts derived from normal ovary (CM1) and ovarian tumor tissue (CM2). Cell proliferation was measured by MTT assay. Phosphorylation of MAPKs and AKT was evaluated by Western blotting. Specific inhibitors of MAPKs and AKT were used to evaluate their respective involvement in mediating increased cell growth. Cytokine levels in fibroblast-CMs were measured using Luminex assays. Immunohistochemical staining was conducted for CXCL1, CXCR2 and phosphorylated p38 in primary ovarian tumors.

Results

CM1 and CM2 significantly increased the growth of ovarian cancer cells relative to SF. In OVCAR3 and OVCAR4 cells, p38 phosphorylation was strongly induced by fibroblast-CMs, and pre-treatment with a p38 inhibitor prevented the growth increase induced by fibroblast-CMs. Fibroblasts secreted high levels of IL-6, IL-8, MCP1 and CXCL1. Treatment with only CXCL1 (1 μg/ml) increased cell growth and p38 phosphorylation. Treatment with a CXCR2 inhibitor effectively prevented p38 activation and cell growth induced by fibroblast-CMs. High expression of both CXCL1 and CXCR2 correlated with high expression of phosphorylated p38 in primary ovarian tumors.

Conclusions

From our data, we conclude that CXCL1 is a key factor derived from ovarian fibroblasts that is responsible for increased ovarian cancer cell growth in part through p38 activation. Phosphorylated p38 can be used as a biomarker to predict CXCL1-CXCR2 interaction in vivo.

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Acknowledgments

This study was supported in part by a grant from the National Research Foundation of Korea (NRF), funded by the Ministry of Science, ICT & Future Planning (2014R1A1A3050916) to YK, the NIGMS (P20 GM130423) and the KU Cancer Center’s Cancer Center Support Grant (P30 CA168524) to AKG. AKG is the Chancellors Distinguished Chair in Biomedical Sciences Endowed Professor.

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Correspondence to Youngjoo Kwon.

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Table S1

Levels of cytokines secreted by ovarian fibroblasts, ovarian cancer cells, and their co-cultures. Fibroblasts were seeded in the lower wells and cancer cells in the upper wells of a 24-well Transwell culture system. Values represent means of duplicate experiments when only ovarian cancer cell lines were used (no fibroblasts); all others are values from single experiments. (DOCX 44 kb)

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Park, GY., Pathak, H.B., Godwin, A.K. et al. Epithelial-stromal communication via CXCL1-CXCR2 interaction stimulates growth of ovarian cancer cells through p38 activation. Cell Oncol. 44, 77–92 (2021). https://doi.org/10.1007/s13402-020-00554-0

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Keywords

  • Ovarian cancer
  • Tumor microenvironment
  • CXCL1
  • CXCR2
  • Cell proliferation
  • Ovarian fibroblast
  • p38 activation