Abstract

One of the possible pathways of action of melatonin is its effect on the cytoskeleton. In this work we looked for alterations in the cytoskeleton of cells treated with melatonin at physiological concentrations. T-47D, Hs-578T (human breast carcinoma cell lines), and MDCK (normal dog kidney) cells were maintained in MCDB 153 supplemented with 1% fetal bovine serum (FBS), or in Dulbecco's modified Eagle's medium (DMEM) supplemented with 5% FBS and treated with melatonin (10(-9) M or 10(-10) M) for 2 and 5 days, with or without 10(-8) M estradiol. F-actin was stained with phalloidin-fluorescein isothiocyanate (FITC). Cytokeratin 19 and beta-tubulin filaments were detected with specific monoclonal antibodies and secondary antibodies bound to FITC. Melatonin-treated T-47D cells observed in a transmission electronic microscope (TEM) showed an irregular nuclear shape and intermediate filaments disposed around the nucleus, which was not observed in control cells. Immunofluorescence analysis of cytokeratin filaments did not show significant differences between their distribution in control and treated cells. Melatonin did not induce significant alterations in cytokeratin filaments of T-47D, Hs578T or MDCK cells in DMEM and MCDB 153, or T-47D cells in DMEM. Melatonin induced the derangement of F-actin both in T-47D and MDCK cells kept in MCDB 153. The same was not observed when estradiol was also present. We did not observe significant alterations in the distribution of F-actin in T-47D or Hs-578T cells grown in DMEM. In DMEM, melatonin-treated MDCK cells were more elongated, with a slight concentration of F-actin on the cell boundary. Melatonin induced very slight alterations in microtubule organization of all cell lines studied.