Rapid and highly sensitive detection of cadmium chloride induced cytotoxicity using the HSP70B' promoter in live cells.
One unique to detect cytotoxicity is to utilize reporter gene assays for promoters that respond to stress-induced effects. In the present study, we discovered that the DNA sequence from nt -287 to +110 of the heat shock protein 70B' (HSP70B') gene could be used as a functional promoter to detect cytotoxicity of cadmium chloride. We thus detected cytotoxicity induced by cadmium chloride with the luciferase assay using this functional HSP70B' promoter, as well as the cell viability test based on the quantification of intracellular ATP. The luciferase assay using the functional HSP70B' promoter resulted in nearly maximal luciferase activity after only 12 h of exposure to cadmium chloride, however, with intracellular ATP quantification, the decrease in cell viability only reached a plateau after 24 h of exposure. Cytotoxicity detection limits for cadmium chloride with the functional HSP70B' promoter assay or cell viability based on ATP quantification were 130 ng/mL and 530 ng/mL, respectively. Our results therefore suggest that the novel reporter gene assay using a functional region of the HSP70B' promoter has significant advantages for the detection of cytotoxicity in terms of both speed and sensitivity, when compared to the cell viability test based on ATP quantification.
Wada
Ken-Ichi
K
Bionic Materials Technology Group, Biomaterials Center, National Institute for Materials Science, Tsukuba, Ibaraki, Japan.
Taniguchi
Akiyoshi
A
Xu
Liming
L
Okano
Teruo
T
D001681:Biological Assay / Q000379:methods; D019256:Cadmium Chloride / Q000032:analysis* / Q000494:pharmacology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics; D006801:Humans; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000187:drug effects; D012680:Sensitivity and Specificity
D018840:HSP70 Heat-Shock Proteins; C491129:HSPA7 protein, human; D008156:Luciferases; D019256:Cadmium Chloride