Bladder Cancer Cells Exert Pleiotropic Effects on Human Adipose-Derived Stem Cells

Abstract

Stem cell-based therapies are considered one of the most promising disciplines in biomedicine. Bladder cancer patients could benefit from therapies directed to promote healing after invasive surgeries or to lessen urinary incontinence, a common side effect of both cancer itself and the treatment. However, the local delivery of cells producing large amounts of paracrine factors may alter interactions within the microenvironment. For this reason, reconstructive cellular therapies for patients with a history of cancer carry a potential risk of tumor reactivation. We used an indirect co-culture model to characterize the interplay between adipose-derived stem cells and bladder cancer cells. Incubation with ASCs increased MCP-1 secretion by bladder cancer cells (from 2.1-fold to 8.1-fold, depending on the cell line). Cancer cell-derived factors altered ASC morphology. Cells with atypical shapes and significantly enlarged volumes appeared within the monolayer. Incubation in a conditioned medium (CM) containing soluble mediators secreted by 5637 and HB-CLS-1 bladder cancer cell lines decreased ASC numbers by 47.5% and 45.7%. A significant increase in adhesion to ECM components, accompanied by reduced motility and sheet migration, was also observed after incubation in CM from 5637 and HB-CLS-1 cells. No differences were observed when ASCs were co-cultured with HT-1376 cells. Our previous and present results indicate that soluble mediators secreted by ASCs and bladder cancer cells induce opposite effects influencing cells that represent the non-muscle-invasive urinary bladder.

Keywords: adipose-derived stem cells; bladder cancer; cancer recurrence; cell-based therapies.

Figure 1

Growth characteristics of ASCs co-cultured with bladder cancer cells or in cancer cells CM. (A) Morphology of ASCs cultured in the presence of cancer cell-derived factors. Magnification x100. (B) ASC numbers decreased by 37% and 9.5% after co-culture with 5637 and HB-CLS-1 cells. Similarly, incubation in CM decreased the cell number by 47.5% and 45.7%. No significant differences were noted when ASCs were co-cultured with HT-1376 cells. (C) Conditioned medium from 5637 and HB-CLS-1 cells decreased ASC viability by 20.6% and 16.7%. No significant changes were observed after co-culture with bladder cancer cell lines. (D) Soluble mediators secreted by 5637 and HB-CLS-1 cells decreased the number of ASCs by 10.3% and 10.7% compared to the control. Co-culture with HT-1376 cells did not influence the proliferation of ASCs. (E) Culture in CM form 5637 and HB-CLS-1 cells reduced exosomal protein content by 8.1% and 17.7%. No differences were noted after culture in CM from HT-1376 cells. Red/orange stands for 5637 cells, gray for HB-CLS-1, and yellow for HT-1376 cells. All results are presented as a percentage of control. Bars represent standard deviation; * p > 0.05, ** p > 0.01, *** p > 0.001.

Figure 2

Adhesion and migration of ASCs cultured in CM from bladder cancer cells. (A) The migration of ASCs cultured in CM from 5637 and HB-CLS-1 cells decreased by 24.0% and 32.4%, respectively. No differences were observed for ASCs cultured in CM from HT-1376 cells. (B) Soluble mediators secreted by 5637 and HB-CLS-1 cells reduced sheet migration. After 24 h observation, 87.4% and 65.3% of the scratch wounds closed compared to 95.7% for cells cultured in a standard growth medium. No significant changes in growth kinetics were observed for ASCs cultured in CM from HT-1376 cells. (C) ASC adhesion significantly increased after culture in conditioned medium from bladder cancer cell lines. Depending on the ECM component, cell adhesion rose from 36.3% to 65.4% for ASCs cultured in CM from 5637 cells and from 56.1% to 83.0% for ASCs cultured in CM from HB-CLS-1 cells. (D) Culture with soluble mediators secreted by HT-1376 cells reduced ERK and AKT activation in ASCs by 37.4% and 18.8%, respectively. No differences were noted after culture in CM from 5637 cells. Although not statistically significant, decreased activation of P70 S6K was observed after culture in CM from all three bladder cancer cell lines. Red/orange stands for 5637 cells, gray for HB-CLS-1, and yellow for HT-1376 cells. All results are presented as a percentage of control. Bars represent standard deviation; * p > 0.05, ** p > 0.01, *** p > 0.001.