1,25-DIHYDROXYVITAMIN-D-3 INDUCES MORPHOLOGICAL AND BIOCHEMICAL MARKERS OF APOPTOSIS IN MCF-7 BREAST-CANCER CELLS

1,25-Dihydroxyvitamin D-3 [1,25(OH)(2)D-3], the active metabolite of v itamin D, is a potent inhibitor of breast cancer cell growth both in v ivo and in vitro. To complement data which documents the anti-prolifer ative effects of 1,25(OH)(2)D-3 we assessed the role of apoptosis in v itamin D-mediated growth arrest of MCF-7 cells. Time course studies in dicated that 100 nM 1,25(OH)(2)D-3 significantly reduces MCF-7 cell nu mbers within 48 h of treatment. Morphological assessment demonstrated that MCF-7 cells treated with 1,25(OH)(2)D-3 for 48 h exhibit characte ristic apoptotic features, including cytoplasmic condensation, pyknoti c nuclei, condensed chromatin and nuclear matrix re-organization. In s itu end labelling with terminal transferase indicated that cells exhib iting apoptotic morphology in 1,25(OH)(2)D-3-treated cultures were pos itive for DNA strand breaks. These morphological features of apoptosis were accompanied by an increase in the cell. death rate assessed as s oluble DNA-histone complexes indicative of DNA fragmentation. To compl ement the morphological data, we assessed the temporal expression of t wo proteins which have been associated with apoptosis in mammary cells and tumors. The steady state mRNA levels for TRPM-2/clusterin and cat hepsin B mRNA were significantly up-regulated in MCF-7 cells treated w ith 1,25(OH)(2)D-3 compared to control cells. Time-dependent increases in the expression of TRPM-2/clusterin and cathepsin B proteins were d etected by Western blotting in 1,25(OH)(2)D-3-treated cells. These fin dings indicate that, in addition to its anti-proliferative effects, 1, 25(OH)(2)D-3 activates the apoptotic cell death pathway in MCF-7 breas t cancer cells. Copyright (C) 1996 Elsevier Science Ltd.