Metallothionein in cultured human epithelial cells and synovial rheumatoid fibroblasts after in vitro treatment with auranofin.
Radioimmunoassay (RIA) and reversed-phase high-pressure liquid chromatography (HPLC) were used to investigate gold-binding proteins of possible metallothionein (MT) nature occurring upon auranofin exposure of cultured human cells. An epithelial cell line (HE) and two sub-strains were examined. The HEAF sub-strain had been made resistant to 2 mumole auranofin/l culture medium. The resistance was associated with the appearance of gold-binding substances with gel filtration characteristics like MT. The HE100 sub-strain had been made resistant to 100 mumole CdCl2/l and contained high amounts of cytosolic Cd-induced MT. In addition, cultured synovial fibroblasts, derived from normal (SN) and rheumatoid (SRA) synovial tissues, were investigated. Evidence was obtained by RIA that the low molecular weight (mol.wt. 6000-7000) gold-binding proteins occurring in the HEAF cells and SRA cells following auranofin exposure, were of MT nature. The relative amounts of MT in the epithelial cell lines were: HE:HEAF:HE100 = 1:18:100. The relative amounts in the synovial fibroblasts were: SN:SRA:SRA treated with auranofin = 1:3:10. The HPLC methods used were found suitable for isolation of Cd-MT in the HE100 cells, but not for the Au-MT in the HEAF cells. By HPLC, the Cd-MT in the HE100 cells was resolved into 3 MT-1 and 1 MT-2 iso-proteins exhibiting the amino acid composition typical of MT. Judged by HPLC, the MT in these cells constituted 0.4% of the cytosolic proteins.
D000596:Amino Acids / Q000032:analysis; D000893:Anti-Inflammatory Agents / Q000494:pharmacology*; D001172:Arthritis, Rheumatoid / Q000378:metabolism*; D001310:Auranofin; D006051:Aurothioglucose / Q000031:analogs & derivatives* / Q000494:pharmacology; D002104:Cadmium / Q000378:metabolism; D002478:Cells, Cultured; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D004848:Epithelium / Q000032:analysis; D005347:Fibroblasts / Q000032:analysis; D006046:Gold / Q000031:analogs & derivatives*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008668:Metallothionein / Q000032:analysis*; D011863:Radioimmunoassay; D013583:Synovial Membrane / Q000032:analysis*
D000596:Amino Acids; D000893:Anti-Inflammatory Agents; D002104:Cadmium; D006051:Aurothioglucose; D001310:Auranofin; D006046:Gold; D008668:Metallothionein