BackgroundRecent studies demonstrate that interstitial inorganic phosphate is significantly elevated in the breast cancer microenvironment as compared to normal tissue. In addition it has been shown that breast cancer cells express high levels of the NaPi-IIb carrier (SLC34A2), suggesting that this carrier may play a role in breast cancer progression. However, the biochemical behavior of inorganic phosphate (Pi) transporter in this cancer type remains elusive.MethodsIn this work, we characterize the kinetic parameters of Pi transport in the aggressive human breast cancer cell line, MDA-MB-231, and correlated Pi transport with cell migration and adhesion.ResultsWe determined the influence of sodium concentration, pH, metabolic inhibitors, as well as the affinity for inorganic phosphate in Pi transport. We observed that the inorganic phosphate is dependent on sodium transport (K0,5 value = 21.98 mM for NaCl). Furthermore, the transport is modulated by different pH values and increasing concentrations of Pi, following the Michaelis-Menten kinetics (K0,5 = 0.08 mM Pi). PFA, monensin, furosemide and ouabain inhibited Pi transport, cell migration and adhesion.ConclusionsTaken together, these results showed that the uptake of Pi in MDA-MB-231 cells is modulated by sodium and by regulatory mechanisms of intracellular sodium gradient.General Significance: Pi transport might be regarded as a potential target for therapy against tumor progression.
Synovial fluid holds a population of mesenchymal stem cells (MSC) that could be used for clinical treatment. Our goal was to characterize the inflammatory and metabolomic profile of the synovial fluid from osteoarthritic patients and to identify its modulatory effect on synovial fluid cells. Synovial fluid was collected from non-OA and OA patients, which was centrifuged to isolate cells. Cells were cultured for 21 days, characterized with specific markers for MSC, and exposed to a specific cocktail to induce chondrogenic, osteogenic, and adipogenic differentiation. Then, we performed a MTT assay exposing SF cells from non-OA and OA patients to a medium containing non-OA and OA synovial fluid. Synovial fluid from non-OA and OA patients was submitted to ELISA to evaluate BMP-2, BMP-4, IL-6, IL-10, TNF-α, and TGF-β1 concentrations and to a metabolomic evaluation using 1H-NMR. Synovial fluid cells presented spindle-shaped morphology in vitro. Samples from OA patients formed a higher number of colonies than the ones from non-OA patients. After 21 days, the colony-forming cells from OA patients differentiated into the three mesenchymal cell lineages, under the appropriated induction protocols. Synovial fluid cells increased its metabolic activity after being exposed to the OA synovial fluid. ELISA assay showed that OA synovial fluid samples presented higher concentration of IL-10 and TGF-β1 than the non-OA, while the NMR showed that OA synovial fluid presents higher concentrations of glucose and glycerol. In conclusion, SFC activity is modulated by OA synovial fluid, which presents higher concentration of IL-10, TGF-β, glycerol, and glucose.
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