Citotoxicity and uptake of mercury by human mammary cells

MARIÁNGELES ÁVILA MANIERO; JORGE SHORTREDE; MARIEL A. FANELLI; RODOLFO WUILLOUD

Chascomus, Buenos Aires

Congreso; iV LABIC, Fourth Latin American Meeting on Biological Inorganic Chemistry. V WOQUIBIO, FIFTH Workshop on Bioinorgánica Chemistry; 2014

Mercury is a widespread pollutant and one ofthe most toxic heavy metal1, classified as 3thclass carcinogen by the International Agency forResearch on Cancer (IARC)2. Mercury chloride(HgCl2), falling into the subclass of bivalentcationic metals, is considered a metalloestrogenbecause it may activate estrogens receptor (ER).Moreover, metallostrogen effects of Hg havenot been investigated as cadmium3. Since Hgremains a serious threat to human health, it is animportant goal to study the effects of this metalon biological systems.MCF-7 mammary cells were the best studymodel to use because they are positive for ER,and a good contrast for this group were MCF-10A cells, as a control, because they arenegative for ER. The aims of this work were to determine theviability of MCF-7 and MCF-10A cells after a 3-hours treatment with HgCl2, testing for possible differences in Hg uptake by both celllines. This work intends to open future metallomics  of mercury in MCF-typecells.  Cell lines and treatment: Two human cells lines were used: MCF-7 from mammary gland adenocarcinoma ER+ and MCF-10A from fibrocystic disease ER-. Cells were treated with different μM HgCl2 concentrations. Citotoxicity assay: MTT assay: 20.000 cells were grown in 96-wells plate for 48 hs andthey were treated for 3 hs with 0; 0,5; 5; 10;25; 50; 100; 150; 200 and 250 μM HgCl2.The MTT assay was performed according to the manufacturer's protocol Protein assay: BCA method was applied to evaluate protein concentration in this cell lines using the manufacturer´s protocol (Pierce Thermo scientific). Mercury uptake: Mammary cell lines were subjected to 0; 10; 25; 50; 75 and 100 μM HgCl2. Intracellular mercury was measured by ETAAS with graphite furnace (Perkin Elmer 5100). This analytical technique allowed sensitive Hg etermination by injecting uL-volume of samples. The results showed that at low Hgconcentrations there was a higher Hg resistance of MCF-10A than MCF-7 cells, but at high Hg concentration, MCF-7 cells were more resistant.Since Hg is a metalloestrogen, it may enter into MCF-7 cells by the ER, which has this type ofreceptors. Furthermore, in MCF-10A cells, which do not have ER, Hg introduction into the cells might be mediated by another mechanism The results of intracellular Hg indicate that there are differences between the two cell lines in its Hg uptake capacity. MCF-10A cells showed higher Hg concentration than MCF-7 upon higher metal exposure. Despite this, MCF-7 cells had a lower IC50 than MCF10A,indicating that this cell line might have a moreeffective detoxification mechanism than MCF-10A ones. CONCLUSION Initial research on Hg citotoxicity and uptake by MCF-type cells is presented in this work. Fromthe results obtained, it can be concluded that MCF-7 and MCF-10A cells have different uptake mechanism for Hg. Thus, there is higher accumulation in mammary cell line from fibrocystic disease than in mammary cell line from denocarcinoma, however the last ones are better defended at high Hg concentration. Thisfinding opens further opportunities and challenges to understand specific molecular mechanisms of Hg toxicity in mammary cells.