In order to clarify the mechanism of metallothionein (MT) induction by bacterial endotoxin, the induction of MT mRNA in cultured human hepatic (Chang) cells by a human MT cDNA was investigated. Although endotoxin added to the culture medium did not induce MT mRNA, the conditioned medium from endotoxin-activated macrophages induced MT mRNA in Chang cells. Conditioned medium containing actinomycin D inhibited the induction of MT mRNA, while conditioned medium containing cycloheximide did not. The induction of MT mRNA was found to be a primary induction. MT inducibility was also tested with IL-1, IFN, IL-6, and TNF, all of which are substances released from endotoxin-activated macrophages. Since they did not induce MT mRNA in Chang cells, endotoxin-activated macrophages release some other MT inducer (s), which are heat stable and synthesized de novo in endetoxin-activated macrophages. To elucidate the details of MT mRNA induction, the kinetics of MT mRNA induction by conditioned medium in Chang cells was studied and compared with the kinetics of MT mRNA induction by zinc and dexamethasone in Chang cells. MT mRNA was induced by conditioned medium dose dependently, and the level of MT mRNA was increased within 3hr, reached a maximum at 12hr, and then gradually decreased. Since the induction of MTmRNA in regulated by factor (s) released from endotoxin-activated macrophages, MT may be involved in the host defense mechanism.
Northern blot hybridization