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Fibrosis of Mesothelial Cell-Induced Peritoneal Implantation of Ovarian Cancer Cells

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Fibrosis of Mesothelial Cell-Induced Peritoneal Implantation of Ovarian Cancer Cells

Jinou Wang et al. Cancer Manag Res.

Abstract

Background: Peritoneal metastasis frequently occurs in patients with advanced ovarian cancer and is the main basis for a poor prognosis. The mechanism underlying preferential ovarian cancer spread to the peritoneum is not well understood.

Methods: Herein, we investigated the significance and mechanism underlying fibrosis of mesothelial cells promoting peritoneal implantation of ovarian cancer. We have assessed the mesothelial cell fibroblast transformation process in peritoneal tissues of omentum and fibrotic mesothelial cell release of chemokines to promote dissemination by scanning electron microscopy, ELISA, Western blot, and Transwell chamber assay.

Results: We showed that the fibrosis of mesothelial cells exists in the peritoneum of ovarian cancer patients with peritoneal metastasis. Fibrosis of the mesothelial cells was induced by TGF-β1, which upregulates the CXCL12-CXCR4 and CXCL16-CXCR6 axes of mesothelial cells.

Conclusion: CXCL12-CXCR4 and CXCL16-CXCR6 may be important signaling pathways closely involved in peritoneal metastasis of ovarian cancer that require further investigation. The findings may lead to developing alternative strategies aimed at preventing and treating the metastasis of ovarian cancer.

Keywords: fibrosis; mesothelial cell; ovarian cancer; peritoneal.

Conflict of interest statement

Disclosure The authors report no conflicts of interest in this work.

Figures

Figure 1
Figure 1
The morphological and biological changes in mesothelial cells in the peritoneal metastases of ovarian cancer. Notes: (A) Mesothelial cells of normal omental tissues are closely packed. (B) Mesothelial cells of metastatic omental tissues exhibited fibrosis change with microscopy. Magnification for (A and B) ×200. (C) The mesothelial cells of normal omentum were arranged tightly and covered with microvilli on the surface. (D) The MFT mesothelial cells formed local bare areas under SEM. Magnification for (C and D) ×2000. (E) Western blot analysis for the levels of E-cadherin and α-SMA proteins in the HMrSV5 and SKOV3 cells. compared with free serum cultured (NC) and normal cultured mesothelial cell line HMrSV5 (HM); the levels of E-cadherin protein were downregulated and the levels of α-SMA protein were upregulated in the HMrSV5 cells that were cultured with supernatants from the SKOV3 (HM-Ca). Abbreviations: MFT, mesothelial cell fibroblast transformation; NC, negative control; SEM, scanning electron microscopy.
Figure 2
Figure 2
The major cytokines induced mesothelial cell fibroblast transformation. Notes: The expression of TGF-β1, IL-10, IL-8, TNF-α, VEGF, EGF, and MMPs in ovarian cancer cell lines (SKOV3 and HO8910) using ELISA kits (A and B). The cellular morphology of HMrSV5 cells examined by retreating with different concentrations of TGF-β1 (0, 10, and 100 ng/mL) (CE); original magnification ×400.
Figure 3
Figure 3
TGF-β1 could induce MFT in a time- and dose-dependent manner. Notes: (A) Western blot analysis for the expression of α-SMA, E-cadherin, and cytokeratin-8 in the HMrSV5 cells treated with TGF-β1 (10 and 100 ng/mL) for 24–72 h. (B) The Western blot data were quantified with a densitometer, the expression of α-SMA was significantly increased, and the expression of E-cadherin and cytokeratin-8 were significantly decreased in the HMrSV5 cells treated with TGF-β1 (100 ng/mL) compared to the control (*P<0.05). Abbreviation: MFT, mesothelial cell fibroblast transformation.
Figure 4
Figure 4
MFT promotes the proliferation and invasion of ovarian cancer cells. Notes: The MTT results showed that supernatants of fibrotic HMrSV5 cell (Fibro-HM) stimulated the proliferation of ovarian cancer cells. (A) SKOV3 and (B) HO8910; results are represented as mean ± SD of three experiments. *P<0.01. Cell migration was monitored using a Transwell chamber assay. The SKOV3 was seeded into the upper chamber. The cells were induced with medium from the lower chamber to migrate without FBS for 20 h and then replaced with (C) HMrSV5 cell suspension in serum-free medium, (D) HMrSV5 cell suspension in SKOV3 medium, (E) HMrSV5 cell suspension in serum-free medium +100 ng/mL of TGF-β1. (F) The number of migrated cancer cells was significantly increased in MFT models (D and E) compared with controls. Magnification ×200. Abbreviation: MFT, mesothelial cell fibroblast transformation.
Figure 5
Figure 5
Chemokines of mesothelial cells’ promotion of ovarian cancer metastasis. Notes: (A) The expression of CXCL12, CXCL16, CXCL19, and CXCL21 was examined in the supernatants of fibrotic HMrSV5 cell (Fibro-HM) and HMrSV5 by ELISA, the free serum cultured HMrSV5 (NC) was the control group. The data on the protein levels were summarized as mean ± SE of each sample. (B) The level of chemokine receptor proteins was examined in supernatants of SKOV3 cells. Compared with the free serum cultured (NC) and normal cultured SKOV3 cells (SKOV3), the levels of CXCR4 and CXCR6 were significantly increased in SKOV3 co-cultured with HMrSV5 supernatants (SKOV3-HM). (C) The Western blot data were quantified with a densitometer (*P<0.05). Abbreviation: NC, negative control.

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