Adipocytes are the main stromal cells in the mammary microenvironment, and crosstalk between adipocytes and breast cancer cells may play a critical and important role in cancer maintenance and progression. Tumor-induced differentiation to beige/brown adipose tissue is an important contribution to the hypermetabolic state of breast cancer. However, the effect of epithelial cell-beige adipocyte communication on tumor progression remains unclear. To contribute to the understanding of this phenomenon, we characterized components present in conditioned media (CM) from beige adipocytes (BAs) or white adipocytes (WAs), and evaluated the effects of BA-and WA-CM on both adhesion and migration of tumor (LM3, 4T1 and MC4-L1) and non-tumor (NMuMG) mouse mammary epithelial cell lines. Additionally, we analyzed the expression of ObR, CD44, vimentin, MMP-9, MCT1 and LDH in tumor and non-tumor mouse mammary epithelial cell lines incubated with BA-CM, WA-CM or Ctrol-CM (control conditioned media). 3T3-L1 preadipocytes differentiated into beige adipocytes upon PPARγ activation (rosiglitazone) displaying characteristics that morphologically resembled brown/beige adipocytes. Levels of UCP1, CIDEA, GLUT4, leptin, MCT4 and FABP4 were increased, while adiponectin, caveolin 1 and perilipin 1 levels were decreased in BAs with respect to WAs. Tumor cell lines revealed lower cell adhesion and increased cell migration after incubation with BA-and WA-CM vs. Ctrol-CM. ObR and MMP-9 in MC4-L1 cells were significantly increased after incubation with BA-CM vs. WA-and Ctrol-CM. In addition, MC4-L1 and LM3 cells significantly increased their migration in the presence of BAs, suggesting that new signals originating from the crosstalk between BAs and tumor cells, could be responsible for this change. Our results indicate that beige adipocytes are able to regulate the behavior of both tumor and non-tumor mouse mammary epithelial cells, favoring tumor progression.

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BackgroundStromal adipocytes and tumor breast epithelial cells undergo a mutual metabolic adaptation within tumor microenvironment. Therefore, browning and lipolysis occur in cancer associated adipocytes (CAA). However, the paracrine effects of CAA on lipid metabolism and microenvironment remodeling remain poorly understood.MethodsTo analyze these changes, we evaluated the effects of factors in conditioned media (CM) derived from explants of human breast adipose tissue from tumor (hATT) or normal (hATN) on morphology, degree of browning, the levels of adiposity, maturity, and lipolytic-related markers in 3T3-L1 white adipocytes by Western blot, indirect immunofluorescence and lipolytic assay. We analyzed subcellular localization of UCP1, perilipin 1 (Plin1), HSL and ATGL in adipocytes incubated with different CM by indirect immunofluorescence. Additionally, we evaluated changes in adipocyte intracellular signal pathways.ResultsWe found that adipocytes incubated with hATT-CM displayed characteristics that morphologically resembled beige/brown adipocytes with smaller cell size and higher number of small and micro lipid droplets (LDs), with less triglyceride content. Both, hATT-CM and hATN-CM, increased Pref-1, C/EBPβ LIP/LAP ratio, PPARγ, and caveolin 1 expression in white adipocytes. UCP1, PGC1α and TOMM20 increased only in adipocytes that were treated with hATT-CM. Also, hATT-CM increased the levels of Plin1 and HSL, while decreased ATGL. hATT-CM modified the subcellular localization of the lipolytic markers, favoring their relative content around micro-LDs and induced Plin1 segregation. Furthermore, the levels of p-HSL, p-ERK and p-AKT increased in white adipocytes after incubation with hATT-CM.ConclusionsIn summary, these findings allow us to conclude that adipocytes attached to the tumor could induce white adipocyte browning and increase lipolysis as a means for endocrine/paracrine signaling. Thus, adipocytes from the tumor microenvironment exhibit an activated phenotype that could have been induced not only by secreted soluble factors from tumor cells but also by paracrine action from other adipocytes present in this microenvironment, suggesting a “domino effect”.