The synthesis of metallothionein and cellular adaptation to metal toxicity in primary rat kidney epithelial cell cultures.

Single cells were isolated from rat (100-150 g) kidney cortex slices by mild trypsinization. They were plated on plastic culture dishes and were maintained in a selective Eagle's D-Valine Minimum Essential Media (MEM) to form an epithelial cell monolayer culture. The fibroblast growth was retarded by D-Valine. The cells in monolayer culture accumulated cadmium when they were incubated with a 109CdCl2 (10(-5) M) containing medium. The synthesis of metallothionein by these cells was demonstrated by incorporation of [35S]cysteine and 109CdCl2 in a heat stable protein (mol. wt 10,000) within 8 h of exposure and also by immunoprecipitation using a specific antibody to rat liver metallothionein. The cytotoxic effects of Zn2+, Cu2+, Cd2+ and Hg2+ were studied in culture after addition of various concentrations of metal salts (10(-5)-10(-2) M). Disrupted cellular colonies with severe cell damage were observed after addition of 10(-3) M Cd2+ or Hg2+ as CdCl2 or HgCl2 while similar toxicity was observed only after addition of 10(-2) M Zn2+ or Cu2+ as ZnSO4 or CuSO4 to confluent cell cultures. The cellular damage due to Cd2+ was protected when the cells were pretreated with Cd2+ (10(-5) M) for 24 h and these cells could tolerate much higher concentrations of cadmium (10(-2) M). These results indicate a direct protective role of intracellular metallothionein in the cellular toxicity of cadmium.