CHARACTERISTICS OF THE ACTION OF OXYSTEROLS 7-KETOCHOLESTEROL AND CHOLESTANE-3β,5α,6β-TRIOL ON ABCS AND LRP TRANSPORT PROTEINS OF MESENCHYMAL STEM CELLS DERIVED FROM BONE MARROW OF PATIENTS WITH ACUTE MYELOID LEUKEMIA

Abstract

ABCs and LRP transport proteins are involved in several metabolic processes, among which the transport of lipids and xenobiotics can be highlighted. Thus, the overexpression of some of these proteins, such as ABC-G2 and the LRP protein, are associated with the phenotype of resistance to multiple drugs, one of the main causes of relapses and relapses in patients with acute myeloid leukemia. Mesenchymal stem cells (MSC) are multipotent cells present in the bone marrow and are part of the hematopoietic niche. Phenotypic changes in MSCs are related to AML progression. The oxysterols, 7-KC and Triol, are derived from the auto-oxidation of the cholesterol molecule; and have functions in cell signaling, survival and death. Therefore, the objective of this work was to evaluate the action of oxysterols, in subtoxic concentrations, of 7-KC (30 μM, 50 μM and 70 μM) and Triol (20 μM, 30 μM and 40 μM) on the ABCs and LRP transport proteins of MSCs derived from bone marrow from patients with AML (MSC-D), being used as a comparator MSC of bone marrow from healthy individuals (MSC-S). Using the indirect immunofluorescence technique, it was observed that the amount of ABC-A1 protein was higher in MSC-S strains and that treatment with 30 μM of 7-KC increased ABC-A1 only in MSC-S strains. On the other hand, treatment with Triol decreased the concentration of ABC-A1 in MSC-S strains, and this decrease in ABC-A1 was associated with a decrease in the amount of the LXR-β receptor. Furthermore, treatment with 70 μM of 7-KC decreased LRP protein expression in MSC-S strains, while in MSC-D strains there was no difference. Treatment with Triol showed no significant effect on LRP protein expression in MSC-S, only in MSC-D strains, where there was an increase in the concentration of 20 μM, followed by a decrease in the concentrations of 30 μM and 40 μM of Triol. Both 7-KC and Triol treatment increased the concentration of SMO protein in the cytoplasm and its translocation to the nucleus in MSC-S cell lines. This effect was not observed in MSC-D strains. Treatment with Triol increased the concentration of ABC-D4 protein in the cytoplasm. Furthermore, the presence of ABC-D4 protein was observed in the nucleus of MSC-S and MSC-D strains. Our results demonstrated that in the MSCs lineages the effect of 7-KC and Triol is mainly metabolic, in which an increase in the oxidized glutathione content, alteration in the processes of mitochondrial dynamics with induction of genes related to the fission and fusion process in both 7-KC and Triol-treated MSC lines. Increased expression of the BCL2 gene and cytoplasmic survivin was also observed in MSCs treated with 7-KC. In conclusion, it is possible to observe differences in the expression of ABC transporters and LRP between CTM-S and CTM-D strains. Furthermore, at the same concentrations of 7-KC and Triol used, the observed effect was different between MSC-S and MSC-D, which demonstrates that there are different mechanisms between mesenchymal stem cell lines. In summary, our results are characteristic of the metabolic action of the oxysterols 7-KC and Triol on MSCs.