Effect of endotoxin challenge on normal, tumor initiated, and invasive human breast cells -

Abstract

Breast cancer is the most common female cancer worldwide, recording high incidence rates caused by risk factors associated with urbanization and economic development. Inflammatory breast cancer (IBC) is the most lethal type of breast cancer, targeting young women, mainly after chronic inflammation. Interestingly, endotoxin (ET) is known to simulate inflammation-like conditions in several in vitro and in vivo models including mammary epithelial cells; however, little is known about the effect of ET-induced inflammation on breast cancer initiation events. In order to investigate the tumor initiating role of ET-induced inflammation, we monitored inflammatory mediators’ response and cancer progression events of in vitro 2D cultures of normal mouse mammary epithelial cells (SCp2), 2D and 3D breast progression models of both, non-tumorigenic S1 cells, and intermediate stage of tumorigenesis S1-Connexin 43 knockouts (Cx43-KO S1), that can closely mimic the in vivo mammary epithelial morphology. Moderately invasive MCF-7 and highly invasive MDA-MB-231 human breast cancer cells were exploited in order to determine the effect of ET treatment on tumor invasion events. Short-term treatment (48 hours) of SCp2 cells with ET concentrations ranging from 0.1 to 1 µg-ml upregulated the levels of MMP-9 produced by SCp2 cells in a dose-dependent manner. Long-term ET treatment (one month), but not short-term, enhanced the migration of SCp2 cells in wound healing assays. As for 2D cultures of the human S1 cells, 9-day ET treatment with concentrations ranging from 2 to 20 µg-ml upregulated the levels of MMP-9 in the conditioned media in a dose dependent manner. MMP-9 production by S1 cells was also upregulated upon long-term ET treatment (one month) with 10 µg-ml. Interestingly, the rate of S1 cells invasion though matrigel substrata was enhanced by both, 9-day and long-term treatment with 10 µg-ml ET. Moreover, immunofluorescent imaging of S1 acini in 3D cultures suggested lumen disruption and β-