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The expression and induction of metallothionein has been associated with protection against oxidative stress and apoptosis. This study examines the effect of tumour suppressor protein p53 on metallothionein expression following CdCl2 treatment in eight human epithelial breast cancer cell lines differing in p53 and oestrogen-receptor status. Cells were treated with 10 microM CdCl2 for 24 h and metallothionein protein levels were measured by cadmium binding assay. MCF7 cells which are p53-positive (p53+) and oestrogen-receptor-positive showed a large induction in metallothionein synthesis by 10.79+/-1.36-fold. Other breast cancer cell lines which are p53-negative (p53-) and oestrogen-receptor-negative or weakly oestrogen-receptor-positive showed a small induction ranging from 1.40+/-0.10 to 3.65+/-0.30-fold. RT-PCR analysis showed an induction of metallothionein mRNA in MCF7 cells by about 1.61+/-0.08-fold, while in HCC1806 cells (p53-, oestrogen-receptor-negative) by 1.11+/-0.13-fold, and in MDA-MB-231 (p53-, oestrogen-receptor-negative) by 1.25+/-0.06-fold. Metallothionein localisation was determined by immunohistochemical staining. Prior to metal treatment, metallothionein was localised mainly in the cytoplasm of MCF7 and MDA-MB-231 cells. After treatment with 10 microM CdCl2 for 24 h, MCF7 cells showed intense nuclear and cytoplasmic staining for metallothionein, while MDA-MB-231 cells showed staining in the cytoplasm with weak nuclear staining. Apoptosis induced by 10-40 microM CdCl2 at time points between 4 and 48 h was examined with TUNEL assay. In MCF7 cells, apoptosis increased with higher concentrations of CdCl2, it peaked at 6-8 h and appeared again at 48 h for all concentrations of CdCl2 tested. In MDA-MB-231 cells, apoptosis remained at low levels for 10-40 microM CdCl2 at all time points. Studies on cadmium uptake showed similar uptake and accumulation of cadmium at 8 and 24 h in all the cell lines. The data demonstrate that treatment of epithelial breast cancer cells with 10 microM CdCl2 for 24 h caused a greater induction of metallothionein protein and mRNA expression in p53+ and oestrogen-receptor-positive cells as compared to p53- and oestrogen-receptor-negative or weakly oestrogen-receptor-positive cells. This effect may be associated with the occurrence of apoptosis and suggests a role for p53 and oestrogen-receptor on the expression and induction of metallothionein in epithelial cells.

Potential role of p53 on metallothionein induction in human epithelial breast cancer cells