Metallothionein and its importance relative to glutathione in cardiac protection against doxorubicin toxicity

Abstract

Controversial results have been reported regarding whether metallothionein (MT) functions in doxorubicin (DOX) detoxification in the heart. To determine unequivocally the role of MT in this cardiac function, a primary culture of myocardial cells was established from 3-day-old transgenic mouse hearts in which MT was over-expressed more than 40-fold higher than normal. DOX was added directly into the cultures to a final concentration of 0, 0.5, 1.0, 2.0 or 4.0 µM after the cells were cultured for 6 days. As compared to nontransgenic controls, trans-genic myocardial cells displayed a significant resistance to DOX cytotoxicity as measured by the leakage of lactate dehydrogenase (LDH) 72 h after treatment, and by cell survival using a tetrazolium colorimetric (MTT) assay 12 h after treatment. To determine the importance of MT relative to glutathione (GSH) in cardiac protection against DOX toxicity, seven-week-old mice from the same transgenic line were treated with buthionine sulfoximine (BSO) by ip injection at 5 mmol/kg, two times with a 12-h interval, before being treated with DOX at a single dose of 15 mg/kg for 4 days. Cardiac GSH was depleted by about 70% in both transgenic and non-transgenic mice. The blood level of creatine kinase released from the heart, an important indicator of cardiac toxicity, was dramatically increased in the BSO-treated non-transgenic mice. This increase was completely inhibited in the BSO-treated transgenic mice. These results thus demonstrated that cardiac MT overexpressing transgenic mice are resistant to DOX cardiotoxicity, and this resistance remains the same under the condition of cardiac GSH depletion. Therefore, MT can compensate for the loss of protection from depletion of GSH in the heart. Selective modulations of decreasing DOX resistance in tumors by BSO and increasing cardioprotection by MT induction may provide an alternative approach to the improvement of DOX chemotherapeutic efficacy.