QUANTIFICATION OF METALLOTHIONEIN AS A BIOMARKER FOR CADMIUM EXPOSURE IN TERRESTRIAL GASTROPODS

摘要

A useful cadmium saturation method, the cadmium-Chelex assay, is adopted and modified for quantification of me-tallothionein induction and protein cadmium saturation in midgut gland of cadmium-exposed Roman snails (Helix pomatia). The assay is based on denaturation of nonmetallothionein, cadmium-binding ligands; complete saturation of metallothionein by adding excess amounts of nonradioactive cadmium; and chelating of excessive amounts of cytosolic metal by the Chelex-100 cation exchange resin. After short-term cadmium feeding, snails quickly responded by showing rising metallothionein levels, protein concentrations in the midgut gland increasing from about 300 μg/g tissue (wet weight) to 750 μg/g within a period of 2 d. At the same time cadmium loading of protein rose form 20% in control snails to 50% saturation in exposed individuals. After long-term cadmium exposure, maximal metallothionein concentrations of about 1,000 μg/g tissue (wet weight) and a relative metallothionein cadmium saturation of 70% were reached in midgut glands of exposed snails. It is proposed that metallothionein quantification in H. pomatia might be used as a tool for biomarker studies in three ways. First, the slope of metallothionein induction might be used as a biomarker for incipient cadmium exposure and for the responsiveness of an invertebrate to metal exposure stress. Second, the steady-state level of metallothionein concentration in the midgut gland of H. pomatia might serve as an integrating biomarker reflecting single or repeated exposure events occurring over a prolonged period of time. Third, the percentage cadmium saturation of metallothionein could be utilized as a biomarker indicating if and by how much the detoxification capacity of H. pomatia is becoming exhausted due to elevated cadmium exposure.