Cadmium-induced expression of immediate early genes in LLC-PK1 cells.

To identify molecular mechanisms underlying renal cell damage by cadmium, the effect of this heavy metal on the level of immediate early genes (IEGs) transcripts in LLC-PK1 cells was studied. Cadmium chloride (CdCl2) induced the expression of four IEGs examined, but with differing time courses. The level of c-fos mRNA peaked at 30 minutes, and then decreased. The levels of c-jun and c-myc transcripts reached a maximum at one hour, and remained elevated up to four hours. Egr-1 mRNA level peaked at one hour, and returned to the control level by three hours. Experiments with cycloheximide and actinomycin D showed, respectively, that induction of IEGs by cadmium occurred in a protein synthesis-independent and transcriptional activation-dependent manner. Cadmium induction of c-fos mRNA was reduced markedly by the intracellular calcium chelator, bis-(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid tetra(acetoxymethyl)-ester (BAPTA/AM), and was decreased partially by a protein kinase C (PKC) inhibitor, 1-(5-isoquinolinylsulfonyl)-2- methylpiperazine (H-7). These data indicate that IEG induction by cadmium requires intracellular calcium mobilization and occurs in part by a PKC-dependent pathway. Exposure of LLC-PK1 cells to CdCl2 (20 microM for 1 to 24 hr) resulted loss of cell viability and DNA fragmentation, which was indicative of apoptosis.