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CCL2 secreted from cancer-associated mesothelial cells promotes peritoneal metastasis of ovarian cancer cells through the P38-MAPK pathway

Abstract

Epithelial ovarian cancer (EOC) is considered to secrete various factors in order to promote peritoneal dissemination through cell-to-cell interaction between cancer and mesothelial cells. We previously revealed that TGF-β secreted from EOC induces normal human peritoneal mesothelial cells (HPMCs) to differentiate into cancer-associated mesothelial cells (CAMCs). However, the relationship between tumor cells and CAMCs in EOC is still unclear. We hypothesized that CAMCs also secrete chemokines that attract cancer cells and induce peritoneal dissemination of EOC. We examined chemokines secreted from HPMCs and CAMCs by human chemokine array, and revealed that conditioned medium of CAMCs (CAMCs-CM) included many types of chemokines. The signals of CCL2 were the highest compared with other chemokines. The secretion and relative expression of CCL2 were significantly higher in CAMCs. Recombinant CCL2 promoted trans-mesothelial migration of HPMCs and the migration and invasion by EOC cells. In addition, CCL2 secreted from CAMCs promoted invasion of EOC cells. Furthermore, the neutralizing antibody of CCL2 reduced invasion by EOC. Clinical outcomes of patients whose tissue expressed higher CCR2 were significantly poorer than in patients whose tissue expression was lower. CCL2 activated the phosphorylation of p38 mitogen-activated protein kinase (MAPK). In addition, CAMCs-CM activated the p38 MAPK pathway. Phosphorylation of p38 MAPK reduced with the presence of neutralizing antibody of CCL2. In conclusion, these data indicate CCL2 in CAMCs-CM promoted the malignant potential of EOC. CCL2 plays a crucial role in the tumor microenvironment of EOC.

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Acknowledgements

We sincerely thank Mr. Y. Koya and Y. Yamakita (Bell Research Center, Department of Obstetrics and Gynecology Collaborative Research, Nagoya University Graduate School of Medicine) for various forms of technical support.

Funding

This study was supported by JSPS (Japan Society for the Promotion of Science) KAKENHI Grants-in-Aid for Scientific Research: Grant Numbers 17H04338 and 16K15704.

Author information

Correspondence to Hiroaki Kajiyama.

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All authors declare that they have no conflict of interest.

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Electronic supplementary material

Table S1 Clinical characteristics of patient ascites (DOCX 15 kb)

Table S2 Clinical characteristics of patients on survival analyses (DOCX 18 kb)

Fig. S1 The expressions of CCR2 in EOC cell lines. The protein levels of SKOV-3, A-2780, and OVCAR-3 were high. However, that of ES-2 was low (JPG 76 kb)

Fig. S2 Proliferation assay to assess the effect of CCL2 for the cell proliferation. There was no correlation between CCL2 and the cell proliferation (JPG 224 kb)

Fig. S3 Survival analyses on stratification by each histological type of EOC. A: overall survival (serous carcinoma), B: overall survival (clear-cell carcinoma), C: progression-free survival (serous carcinoma), D: progression-free survival (clear-cell carcinoma) (JPG 430 kb)

Fig. S4 Schematic summary of the current study. Enhanced production of CCL2 from CAMCs is crucial for the development of peritoneal carcinomatosis of EOC through the adhesion-, migration-, and invasion-promoting effects of tumor cells. These findings indicate the scheme of the auto-stimulatory triangle axis among cancer, ascites, and the mesothelium (JPG 528 kb)

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Yasui, H., Kajiyama, H., Tamauchi, S. et al. CCL2 secreted from cancer-associated mesothelial cells promotes peritoneal metastasis of ovarian cancer cells through the P38-MAPK pathway. Clin Exp Metastasis 37, 145–158 (2020). https://doi.org/10.1007/s10585-019-09993-y

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Keywords

  • Ovarian carcinoma
  • Peritoneal dissemination
  • Mesothelial cells
  • CCL2
  • CCR2
  • TGF-β