Increased ROS production in non-polarized mammary epithelial cells induces monocyte infiltration in 3D culture

Loss of epithelial cell polarity promotes cell invasion and cancer dissemination. Therefore, identification of factors that disrupt polarized acinar formation is critical. Reactive oxygen species (ROS) drive cancer progression and promote inflammation. Here, we show that the non-polarized breast cancer cell line, T4-2, generates significantly higher ROS levels than polarized S1 and T4R cells in 3D (three dimensional) culture, accompanied by induction of the nuclear factor κB (NF-κB) pathway and cytokine expression. Minimizing ROS in T4-2 cells with antioxidants reestablished basal polarity and inhibited cell proliferation. Introducing constitutively activated RAC1 disrupted cell polarity and increased ROS levels, indicating that RAC1 is a critical regulator that links cell polarity and ROS generation. We also linked monocyte infiltration with disruption of polarized acinar structure using a 3D co-culture system. Gain- and loss-of function experiments demonstrated that increased ROS in non-polarized cells is necessary and sufficient to enhance monocyte recruitment. ROS also induced cytokine expression and NF-κB activity. These results suggest that increased ROS production in mammary epithelial cell leads to disruption of cell polarity and promotes monocyte infiltration.