Caracterização do meio condicionado e das vesículas extracelulares pelo tecido adiposo de indivíduos obesos e seus efeitos de células tumorais de mama

Abstract

Obesity is a multifactorial disease characterized by a meta-inflammation sustained by adipose tissue (AT). Evidence has shown that obesity is associated with a worse prognosis and an increased risk of death in women with breast cancer, the most common type of cancer among women. In addition to releasing adipokines, the AT also secrets extracellular vesicles (EVs), which are small membrane vesicles capable of transferring several molecules to target cells. We have previously shown that CM and EVs released by white AT modulate the migratory and proliferative activity of breast cancer cells, thus suggesting their role in modifying the tumor microenvironment to a more malignant profile. One of the processes related to the increase in tumor malignancy is the epithelial-mesenchymal transition (EMT), which is defined by losing epithelial characteristics and acquiring the mesenchymal phenotype, increasing migratory and invasive capacity. In this study, we aimed to characterize the secretome (CM and EVs) released by white visceral AT (VAT) and subcutaneous AT (SAT) explants from lean and obese patients and to investigate the content from obese AT secretome that could be related to the increased malignancy of human breast adenocarcinoma lineages: MCF -7 (luminal A-like) and MDA-MB-231 (basal-like). This model, already established in our laboratory, is performed from the culture of AT explants obtained from obese or lean patients who underwent bariatric or plastic surgery, respectively. In the CM, the lipid profile was analyzed by gas chromatography; Milliplex evaluated the adipokine panel; the protein load present in the EVs was analyzed by proteomics and Western blotting; the expression of EMT-related markers in breast tumor cells was evaluated by Western blotting; the migration assay was performed using the wound healing assay. Our results showed that comparing SAT depots, those derived from obese patients presented higher levels of TGF-β1, showing a positive correlation with BMI. Comparing VAT deposits, obese VAT presented higher levels of IL-6, TGF-β 2, and 3, without correlation with BMI. In addition, obese VAT showed more significant release of saturated fatty acids such as palmitic, lauric, myristic, and stearic, besides oleic (monounsaturated) and linoleic (polyunsaturated) fatty acids. Compared to lean individuals, EVs released by the AT from obese individuals showed proteins related to tumor progression, such as S100A4, S100A11, ANXA, TGF-β, and vimentin. The CM and EVs from obese AT induced an increase in MCF-7 cells migration compared to the control and decreased the expression of the epithelial marker E-cadherin. In MDA-MB-231 cells, we did not observe changes in the mesenchymal marker vimentin between groups. The EVs from obese AT induced a decrease in CDH1, Slug, and an increase in Snail in MCF-7 cells, while in MDA-MB-231 cells, it also induced an increase in Snail. Furthermore, EVs from obese AT also induced activation of the TGF-β/SMAD pathway in MCF-7 cells. Our data indicate that ET-derived EVs from obese individuals carry proteins with pro-tumor properties that increase the malignancy of tumor cells through a decrease in E-cadherin expression and increasing migration.