Shiraishi N, Hochadel J F, Coogan T P, Koropatnick J, Waalkes M P
Inorganic Carcinogenesis Section, PRI DynCorp, Frederick Cancer Research, National Cancer Institute, Frederick, Maryland 21702-1201.
Toxicol Appl Pharmacol. 1995 Feb;130(2):229-36. doi: 10.1006/taap.1995.1028.
Cadmium is a carcinogenic metal. Although the mechanism of tumor induction is unknown, DNA/metal interactions may be involved. Metallothionein can protect against cadmium toxicity in our previous work it was shown to reduce cadmium genotoxicity in cultured cells. To extend these results, the genotoxicity of cadmium was studied in R2C cells, a rat testicular Leydig cell line. The R2C cells were very sensitive to cadmium-induced single-strand DNA damage (SSD), as measured by alkaline elution. SSD occurred in R2C cells after treatment with 25 and 50 microM CdCl2 for 2 hr. Prior work showed other cells required much higher levels of cadmium (approximately 500 microM) to induce genotoxicity. The genotoxic levels of cadmium (25-50 microM) were not cytotoxic in R2C cells as assessed by a metabolic activity (MTT) assay. Pretreatment of R2C cells with a low cadmium dose (2 microM, 24 hr) had no effect on cadmium-induced SSD, in contrast to prior work in other cells where such pretreatments reduced SSD through metallothionein gene activation. In fact, cadmium or zinc treatments resulted in little or no increase in metallothionein gene expression in R2C cells as determined by Northern blot analysis for metallothionein mRNA using cDNA or oligonucleotide probes and radioimmunoassay for metallothionein protein production. Basal metallothionein mRNA was essentially nondetectable. Induction of a cadmium-binding protein in R2C cells did occur, as determined by Cd-heme assay, but did not induce tolerance to SSD. In vivo, the Leydig cell is a target for cadmium carcinogenicity and its cadmium-binding protein is thought not to be a true metallothionein. These results indicate that R2C cells are sensitive to cadmium-induced genotoxicity and that this sensitivity is associated with minimal expression of the metallothionein gene.
镉是一种致癌金属。尽管肿瘤诱发机制尚不清楚，但可能涉及DNA/金属相互作用。金属硫蛋白可抵御镉毒性，在我们之前的研究中，它能降低培养细胞中的镉遗传毒性。为扩展这些结果，我们在大鼠睾丸间质细胞瘤细胞系R2C细胞中研究了镉的遗传毒性。通过碱性洗脱法测定，R2C细胞对镉诱导的单链DNA损伤（SSD）非常敏感。在用25和50微摩尔/升氯化镉处理2小时后，R2C细胞中出现了SSD。先前的研究表明，其他细胞需要更高水平的镉（约500微摩尔/升）才能诱导遗传毒性。通过代谢活性（MTT）测定评估，镉的遗传毒性水平（25 - 50微摩尔/升）对R2C细胞无细胞毒性。用低剂量镉（2微摩尔/升，24小时）预处理R2C细胞对镉诱导的SSD没有影响，这与之前在其他细胞中的研究结果相反，在其他细胞中这种预处理通过金属硫蛋白基因激活降低了SSD。事实上，通过使用cDNA或寡核苷酸探针进行金属硫蛋白mRNA的Northern印迹分析以及金属硫蛋白蛋白产生的放射免疫测定确定，镉或锌处理导致R2C细胞中金属硫蛋白基因表达几乎没有增加。基础金属硫蛋白mRNA基本无法检测到。通过镉 - 血红素测定确定，R2C细胞中确实诱导产生了一种镉结合蛋白，但并未诱导对SSD的耐受性。在体内，间质细胞是镉致癌作用的靶细胞，其镉结合蛋白被认为不是真正的金属硫蛋白。这些结果表明，R2C细胞对镉诱导的遗传毒性敏感，且这种敏感性与金属硫蛋白基因的最小表达相关。
