Ascites IL-10 Promotes Ovarian Cancer Cell Migration

Abstract

Ovarian cancer (OC) ascites is an inflammatory and immunosuppressive tumor environment characterized by the presence of various cytokines, chemokines and growth factors. The presence of high concentrations of these cytokines/chemokines in ascites is associated with a more aggressive tumor phenotype. IL-10 is an immunosuppressive cytokine for which high expression has been associated with poor prognosis in some cancers. However, its role on OC tumor cells has not been explored. Therefore, the aim of the current study was to elucidate the role of ascites IL-10 on the proliferation, migration and survival of OC cell lines. Here, we show that IL-10 levels are markedly increased in patients with advanced serous OC ascites relative to serous stage I/II ascites and peritoneal effusions from women with benign conditions. Ascites and IL-10 dose-dependently enhanced the proliferation and migration of OC cell lines CaOV3 and OVCAR3 but had no effect on cell survival. IL-10 levels in ascites positively correlated with the ability of ascites to promote cell migration but not proliferation. Depletion of IL-10 from ascites markedly inhibited ascites-induced OC cell migration but was not crucial for ascites-mediated cell proliferation. Taken together, our findings establish an important role for IL-10, as a component of ascites, in the migration of tumor cells.

Keywords: Ascites; Cell migration; Cell proliferation; IL-10; Ovarian carcinoma.

Fig. 1

IL-10 levels in ovarian cancer ascites. a The concentration of IL-10 in benign fluids (n = 10), stage I/II serous ovarian cancer ascites (n = 7) and stage III/IV ascites (n = 50) was determined by ELISA. Box plot representing levels of IL-10. b Kaplan-Meier curve of ascites IL-10 levels. The median IL-10 level (46.6 pg/ml) was taken as the cutoff point

Fig. 2

IL-10 enhance the migration of tumor cells. a The migration of ovarian cancer CaOV3 cells (left panel) and OVCAR3 cells (right panel) was evaluated at 24 h using Boyden chambers with either serum-free medium (control) or different concentrations (0–25 ng/ml) of IL-10 as chemoattractants. b On day 0, 5000 cells were incubated in media supplemented with no serum or 0–1000 pg/ml of IL-10 for 48 h. Cell growth was determined by XTT assay. CaOV3 cells (left panel) and OVCAR3 cells (right panel). * indicates P < 0.05. Values are presented as mean ± SEM and represent three independent experiments (c) CaOV3 cells (left panel and OVCAR3 cells (right panel) were incubated in media with no serum containing IL-10 (10 ng/ml) for 24 h. Cells were then challenged with increasing concentration of cisplatin (0 to 50,000 ng/ml). Cell viability was determined by XTT assay after 48 h. Values are presented as mean ± SEM and represent three independent experiments

Fig. 3

Ovarian cancer ascites promotes cell migration. a The migration of ovarian cancer CaOV3 cells was evaluated at 24 h using Boyden chambers with either serum-free medium (control) or different serous OC ascites as chemoattractants (concentration 10% v/v). b Migration of CaOV3 cells with increasing concentration of OVC439 ascites evaluated at 24 h. c Migration of ovarian cancer OVCAR3 cells in the presence of either OVC439 or OVC469 (10% v/v). d On day 0, 5000 cells were incubated in media supplemented with no serum or 10% v/v serous OC ascites for 48 h. Cell growth was determined by Trypan Blue exclusion assay. CaOV3 cells (left panel) and OVCAR3 cells (right panel). e Proliferation of CaOV3 cells in the presence of increasing concentrations of OVC439 (0 to 25% v/v). * indicates P < 0.05. Values are presented as mean ± SEM and represent three independent experiments

Fig. 4

IL-10 levels in ascites correlates with ability of ascites to stimulate CaOV3 cell migration. The correlation coefficient (r) was determined by Pearson’s correlation coefficient test. a Correlation between ascites-induced CaOV3 cell migration and ascites IL-10 concentration. b Correlation between ascites-induced CaOV3 cell proliferation and ascites IL-10 concentration

Fig. 5

Ascites IL-10 contributes to ascites-induced cell migration. CaOV3 (left panel) and OVCAR3 cells (right panel) were incubated in serum-free media containing ascites (OVC439 and OVC469 with IL-10 blocking antibody (20 μg/ml). Cell migration was determined 24 h later. Values are presented as mean ± SEM and represent three independent experiments. * indicates P < 0.05