Cytotoxicity and stress gene microarray analysis in cadmium-exposed CRL-1439 normal rat liver cells

Cadmium is a biologically non-essential divalent hazardous metal. Previous studies demonstrated that cadmium toxic effect was caused by reactive oxygen species. Since gene expression is influenced by the presence of these reactive oxygen species, the association between metal intoxication and gene expression has recently become a major focus of research. We examined the effect of cadmium chloride on cell viability at 4, 8 and 24 h. Our results indicate that cadmium chloride did not alter cell viability at 4 or 8 h, but decreased the viability in a dose-dependent manner (p > 0.01) at 24 h. Using DNA microarray, we studied the profile of stress gene expression in rat primary hepatocytes treated with cadmium for different time periods using a 100 mu M cadmium chloride concentration. Microarray analysis indicated that cadmium treatment caused different patterns of gene expression profiles at each time point of incubation. Of the 207 stress genes on the microarray, only 32 genes were regulated. Since microarrays were hybridized by radioactive cDNA which was less sensitive than fluorescent-labeled cDNA, an experimental/control ratio > 1.3 or < 0.7 (30% increase or decrease) was taken as significant up- or down-regulation. Exposure of cells to cadmium for 4 h resulted in the expression of three up-regulated genes and six down-regulated genes. Longer exposure to cadmium for 8 h resulted in an increase in up-regulated genes to six and down-regulated genes to 14. After 24 h of cadmium exposure, 15 genes were down-regulated and six genes were up-regulated. Our findings suggest that the cells maintained complete viability up to 8 h with cadmium due to expression of various heat shock proteins and stress response proteins like heme oxygenase. Longer exposure periods, due to the down-regulation of the basic cell function proteins and cell-cycle regulating proteins, led to toxicity in cells and eventually to cell death.