Metallothionein (MT) is a low-molecular-mass, cysteine-rich metal binding protein thought to be involved in the detoxification of heavy metals and scavenging of free radicals. MT is directly induced not only by heavy metals, but also by hormones and cytokines. The present study, which uses mice with genetic deletions of the MT proteins (MT −/− mice), was designed to evaluate the effects of MT on the expression of pro-inflammatory cytokines in macrophages. We found that the production of tumour necrosis factor (TNF) induced by lipopolysaccharide (LPS) in peritoneal macrophages is up-regulated by MT via the modulation of nuclear factor κB (NF-κB) activity. This conclusion is supported by the following observations : (1) LPS stimulated the secretion of less TNF activity from MT −/−

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Metallothionein (MT) is a low-molecular-mass, cysteine-rich metal binding protein thought to be involved in the detoxification of heavy metals and scavenging of free radicals. MT is directly induced not only by heavy metals, but also by hormones and cytokines. The present study, which uses mice with genetic deletions of the MT proteins (MT(-/-) mice), was designed to evaluate the effects of MT on the expression of pro-inflammatory cytokines in macrophages. We found that the production of tumour necrosis factor (TNF) induced by lipopolysaccharide (LPS) in peritoneal macrophages is up-regulated by MT via the modulation of nuclear factor kappaB (NF-kappaB) activity. This conclusion is supported by the following observations: (1) LPS stimulated the secretion of less TNF activity from MT(-/-) peritoneal exudate macrophages (PEMs) than from wild-type controls (MT(+/+) mice) without a difference in the pattern of kinetics; (2) LPS-stimulated expression of TNF-alpha mRNA was decreased in MT(-/-) PEMs; (3) LPS-stimulated activation of NF-kappaB was decreased in MT(-/-) PEMs; and (4) production of TNF in PEMs of MT(-/-) mice after LPS treatment in vivo was decreased (compared with MT(+/+) PEMs). Expression of other inflammatory cytokines, interleukin (IL)-1alpha and IL-6 mRNA, which were modulated by NF-kappaB, were also down-regulated in MT(-/-) PEMs. Thus MT plays a key role in the LPS-induced activation of PEMs via the modulation of NF-kappaB activity.

Macrophage colony stimulating factor (M-CSF) plays an important role in the proliferation and differentiation of mononuclear phagocytes. The present study investigates the effect of zinc on M-CSF expression in MC3T3-E1 and L929 cells. Zinc dose-dependently increased M-CSF mRNA levels. The time-course of zinc-induced M-CSF mRNA expression peaked at 6 h. Stability studies of mRNA using actinomycin D revealed that zinc does not affect M-CSF mRNA stability. We examined the function of the M-CSF gene promoter using a luciferase reporter assay. A construct containing the -467/+39 region of the promoter was upregulated by zinc. In the presence of cycloheximide, zinc did not induce a greater increase in the M-CSF mRNA than cycloheximide alone. To confirm the effect of MT on M-CSF mRNA expression, mouse lung fibroblasts (MLFs) were prepared from MT+/+ and MT-/- mice. Zinc induced an increase in the expression of M-CSF in MT+/+ MLFs, but this response was not evident in MT-/- MLFs. Moreover, overexpression of MT upregulated M-CSF mRNA expression as well as M-CSF secretion. Our findings suggest that MT expression mediates zinc regulation of M-CSF gene expression at the transcriptional level.

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