Crosstalk between adipocytes and breast cancer cells: modulation of nuclear receptor coactivator 3 (rac3) in fat cells

Abstract

Adipocytes account for the largest proportion among the cells that comprise breast tissue. Although they are considered to be a critical cell type in the tumor microenvironment of breast cancer, there is still unclear the molecular mechanisms that control their behavior in this context. We have demonstrated that the transcriptional coactivator RAC3 decreases during adipogenesis and its high expression in adipose tissue adjacent to breast tumors correlate with different markers of tumoral progression in patient samples. Therefore, the aim of this work was to further study the role of this molecule in an in vitro model. Conditioned media (CM) were collected from non-tumoral or tumoral breast human cell lines with the following RAC3 mRNA levels: MCF10A: 1.55 ± 0.73, MDA-MB-231: 167.20 ± 6.91, T47D: 571.00 ± 12.59. 3T3-L1 derived-murine adipocytes were stimulated with CM or serum-free medium. RAC3 expression levels measured by qPCR resulted significantly higher in adipocytes stimulated with CM from high RAC3 levels-expressing T47D cells (9.25 ± 1.02) compared to the other conditions: basal (1.00 ± 0.00), MCF10A (2.33 ± 0.96), MDA-MB-231 (2.80 ± 1.19) (p<0.05). As RAC3 is a NF-kB coactivator we studied by immunofluorescence the presence of its phosphorylated subunit p65. We observed a greater fluorescence intensity in nucleus when adipocytes were stimulated with CM from tumoral cell lines (MDA-MB-231 or T47D) compared to the other conditions. Even more, since TNF is a NF-kB target gene, we evaluated by dot plot its levels in CM from adipocytes post-stimulation. Adipocytes secreted more TNF after being stimulated with CM from T47D (1.41 ± 0.03) compared to all other conditions: basal (1.00 ± 0.00), MCF10A (1.08 ± 0.11), MDA-MB-231 (1.02 ± 0.04) (p<0.05). The values were relativized to basal condition and results are shown as the mean ± SEM. These results validate our previous findings and suggest RAC3 as a molecular key to understand changes in the adipokines pattern in the tumor context.