β2ARs stimulation in osteoblasts promotes breast cancer cell adhesion to bone marrow endothelial cells in an IL-1β and selectin-dependent manner

Progression and recurrence of breast cancer, as well as reduced survival of patients with breast cancer, are associated with chronic stress, a condition known to impact the hypothalamic-pituitary axis and the autonomic nervous system. Preclinical and clinical evidence support the involvement of the sympathetic nervous system in the control of bone remodeling and in pathologies of the skeleton, including bone metastasis. In experimental mouse models of skeletal metastasis, administration of the beta AR agonist isoproterenol (ISO), used as a surrogate of norepinephrine, the main neurotransmitter of sympathetic neurons, was shown to favor bone colonization of metastatic breast cancer cells via an increase bone marrow vascularity. However, successful extravasation of cancer cells into a distant organ is known to be favored by an activated endothelium, itself stimulated by inflammatory signals. Based on the known association between high sympathetic outflow, the expression of inflammatory cytokines and bone metastasis, we thus asked whether beta AR stimulation in osteoblasts may alter the vascular endothelium to favor cancer cell engraftment within the skeleton. To address this question, we used conditioned medium (CM) from PBS or ISO-treated bone marrow stromal cells (BMSCs) in adhesion assays with bone marrow endothelial cells (BMECs) or the endothelial cell line C166. We found that ISO treatment in differentiated BMSCs led to a robust induction of the pro-inflammatory cytokines interleukin-1 beta (IL-1 beta) and interleukin-6 (IL-6). The CM from ISO-treated BMSCs increased the expression of E-and P-selectin in BMECs and the adhesion of human MDA-MB-231 breast cancer cells to these cells in short-term static and dynamic adhesion assays, and a blocking antibody against IL-1 beta, but not IL-6, reduced this effect. Direct IL-1 beta treatment of BMECs had a similar effect, whereas the impact of IL-6 treatment on the expression of adhesion molecules by BMECs and on the adhesion of cancer cells to BMECs was negligible. Collectively, these in vitro results suggest that in the context of the multicellular and dynamic bone marrow environment, sympathetic activation and subsequent beta AR stimulation in osteoblasts may profoundly remodel the density but also the activation status of bone marrow vessels to favor the skeletal engraftment of circulating breast cancer cells.