Modulation of reactive oxygen levels and gene expression in sensitive and resistant tumoral cells by C-phyocyanin.

In this work we investigated the possible cellular changes induced by C-phycocyanin (C-PC) in erythroleukemic cells with (K562-Lucena and FEPS) and without (K562) the multidrug resistance (MDR) phenotype. The reactive oxygen levels (ROS) (evaluated by fluorimetry) were increased relative to the control in K562 and K562-Lucena cells treated with 100 µg/mL and were not increased in FEPS cells. The expression of the following genes related to resistance was evaluated by real-time PCR: COX2, ALOX5, ABCB1 and ABCC1. Treatment with 100 µg/mL of C-PC increased COX2 and ABCB1 expression in K562-Lucena cells and reduced expression of ALOX5 in K562-Lucena and FEPS cells. ROS levels appear to be involved in biological responses to C-PC in K562 and K562-Lucena cells, although expression of the genes studied here was only modified by C-PC in the K562-Lucena cell line. Thus, it is possible to suggest that C-PC modulates the expression of COX2 and ABCB1 for the K562-Lucena in a ROS-dependent manner and the expression of ALOX5 for the FEPS in a ROS-independent manner; however, more studies are needed to elucidate these mechanisms.

C-phycocyanin to overcome the multidrug resistance phenotype in human erythroleukemias with or without interaction with ABC transporters.

The phenotype of multidrug resistance (MDR) is one of the main causes of chemotherapy failure. Our study investigated the effect of C-phycocyanin (C-PC) in three human erythroleukemia cell lines with or without the MDR phenotype: K562 (non-MDR; no overexpression of drug efflux proteins), K562-Lucena (MDR; overexpression of ATP-binding cassette, sub-family B/ABCB1), and FEPS (MDR; overexpression of ABCB1 and ATP-binding cassette, sub-family C/ABCC1). Using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, we showed that 20 and 200 µg/mL C-PC decreased K562 viable cells after 24 h and 200 µg/mL C-PC decreased K562-Lucena cell proliferation after 48 h. C-PC did not decrease viable cells of FEPS cells. On the other hand, the MTT assay showed that exposure of 2, 20, and 200 µg/mL C-PC for 24 or 48 h was not cytotoxic to peritoneal macrophages. At 72 h, the trypan blue exclusion assay showed that 20 µg/mL C-PC decreased K562 and K562-Lucena cell proliferation and in FEPS cells, only 200 µg/mL C-PC decreased proliferation. In addition, protein-protein docking showed differences in energy and binding sites of ABCB1 and ABCC1 for C-PC, and these results were confirmed by the efflux protein activity assay. Only ABCC1 activity was altered in the presence of C-PC and FEPS cells showed lower C-PC accumulation, suggesting C-PC extrusion by ABCC1, conferring C-PC resistance. In combination with chemotherapy (vincristine [VCR] and daunorubicin [DNR]), the sensitivity of K562-Lucena cells for C-PC + VCR did not increase, whereas FEPS cell sensitivity for C-PC + DNR was increased. In molecular docking experiments, the estimated free energies of binding for C-PC associated with chemotherapy were similar (VCR: -6.9 kcal/mol and DNR: -7.2 kcal/mol) and these drugs were located within the C-PC cavity. However, C-PC exhibited specificity for tumor cells and K562 cells were more sensitive than K562-Lucena cells, followed by FEPS cells. Thus, C-PC is a possible chemotherapeutic agent for cells with the MDR phenotype, both alone in K562-Lucena cells (resistance due to ABCB1), or in combination with other drugs for cells similar to FEPS (resistance due to ABCC1). Moreover, C-PC did not damage healthy cells (peritoneal macrophages of Mus musculus).

Integrated biological responses of zebrafish (Danio rerio) to analyze water quality in regions under anthropogenic influence.

This study analyzed water quality in regions around Patos lagoon (Southern Brazil) that are under anthropogenic pressure. Water samples were collected from five different sites, including one used as a source for human consumption (COR) and others known to be influenced by human activities (IP). Danio rerio (Teleostei, Cyprinidae) organisms were exposed for 24h to these water samples, plus a control group. It was observed that: (1) reactive oxygen species levels were lower in COR and IP than in the control group; (2) glutamate-cysteine ligase (catalytic subunit) expression was higher in COR than in other sites; (3) exposure to all water samples affected long-term memory (LTM) when compared to control group. Thus, some water samples possess the ability to modulate the antioxidant system and to induce a decline in cognitive functions, as measured by LTM. The obtained results indicate that a combination of variables of different organization level (molecular, biochemical and behavioral) can be employed to analyze water quality in impacted regions.