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Methylmercury (MeHg) exposure during pregnancy is a concern because of its potential health risks to fetuses. Intestinal microbiota has important roles in the decomposition and fecal excretion of MeHg. We investigated the effect of nondigestible saccharides on the accumulation and excretion of Hg after MeHg exposure. Female BALB/cByJ mice were fed a basal diet or the same diet supplemented with 5% fructooligosaccharides (FOS) or 2.5% glucomannan. Six weeks after feeding, mice were administered MeHg chloride (4 mg Hg/kg, per os (p.o.)), and urine and feces were collected for 28 d. FOS-fed mice had lower total Hg levels in all tissues (including the brain) compared with that of controls. The glucomannan diet had no effect on tissue Hg levels. No differences in tissue concentrations of inorganic Hg among groups were found. Fecal Hg excretion was markedly higher in FOS-fed mice than that in controls, but urinary Hg excretion was similar. FOS-fed mice had a higher proportion of inorganic Hg in feces than that of controls, with a significant increase in fecal Hg excretion. Analysis of fecal bacterial population showed the relative abundance of Bacteroides in FOS-fed mice to be higher than that in controls. The results suggest that FOS enhanced fecal Hg excretion and decreased tissue Hg levels after MeHg administration, possibly by accelerating MeHg demethylation by intestinal bacteria (the candidate genus Bacteroides). This demethylation also reduces MeHg absorption in the large intestine. In conclusion, daily FOS intake may decrease tissue Hg levels in animals and humans exposed to MeHg.

Cadmium (Cd) is an environmental pollutant present in contaminated water, food and soil. Cd adversely affects fetal development. We exposed pregnant mice to daily oral doses of 5 and 10 mg/kg Cd and examined fetal growth. It was demonstrated that the exposure to Cd (10 mg/kg) during gestation caused fetal growth retardation (FGR). Investigation of the ubiquitin-proteasome system in fetal livers of mice exposed to gestational Cd revealed increased polyubiquitinated protein accumulation, contrasting with decreased levels of monoubiquitin protein. Moreover, the expression level of Ubc (encoding polyubiquitin C protein) was significantly decreased in 5 and 10 mg/kg Cd-treated groups in comparison with the control group. Therefore, we propose that decrease of monoubiquitin level and accumulation of polyubiquitinated protein in the fetal liver may be important factors in Cd-induced FGR.

We compared the cadmium (Cd) concentration in the liver and kidney of different strains of mice after exposure to 50 ppm Cd for 30 days via drinking water. Cd concentration in the liver and kidney of C57BL/6J mice were higher than those of 129/Sv and DBA/2 mice. Since orally ingested heavy metals are absorbed in the small intestine and then widely distributed to target tissues, microarray analyses were performed to compare the expression levels of transport-related genes in the duodenum between C57BL/6J mice and 129/Sv or DBA/2 mice. The expression levels of 9 and 11 genes were elevated more than 2.0-fold and 13 and 12 genes were reduced less than 0.5-fold in 129/Sv mice and DBA/2 mice, respectively. Among these low expressed genes, 10 genes (Slc2a2, Slc5a1, Slc16a2, Slc22a13, Slc22a18, Slc25a11, Slc36a1, Slco6c1, Abca3 and Abcd1) were common between the two types of strains. These results suggest that some of those genes might be involved in Cd absorption and its toxicity.

This study investigated the influence of forest walking on blood pressure (BP), profile of mood states (POMS) and salivary cortisol in both young and aged people. Twenty-three young people (Men=11, Women=12) with a mean age of 22 yrs and twenty-five aged people (M=10, W=15) with a mean age of 59 yrs participated in this study voluntarily. Each participant walked about three hours in the forest according to their comfortable walking pace. After forest walking, systolic BP (SBP) and mean BP (MAP) had significantly decreased in aged people (-10±3 mmHg in SBP, and -5±2 mmHg in MBP, P

Metallothionein (MT) is a low-molecular-weight cysteine-rich protein which has a high affinity for metals and plays important roles in the protection against metal toxicity. As little information is available concerning the mechanism of MT induction by lead (Pb) compounds, we investigated the induction of MT by Pb acetate both at mRNA and protein levels in mice. Administration of Pb increased the levels of MT-I mRNA in the liver and kidney in six strains of mice. However, MT protein was detected only in the liver, and little or no increases in MT protein were detected in the kidney of any strains of mice. Speciation of metals in the liver cytosol showed that the major metal bound to MT was zinc but not Pb. The increases in plasma concentrations of interleukin-6 suggest that the production of interleukin-6 by Pb administration is involved in the induction of MT in the liver. Treatment of renal cells with Pb in vitro also resulted in the increase in MT mRNA but little increase in MT protein. These data suggest that Pb exerts a dual effect on MT expression; enhancement of MT gene transcription both in the liver and kidney and suppression of MT mRNA translation in the kidney.

Hepatic stellate (HS) cells were isolated from the livers of metallothionein (MT)-null and control mice and used to establish IMS/MT(-) and IMS/N cell lines, respectively, using SV40 virus transformation. Cellular morphology, incorporation of vitamin A and expression of alpha-SMA, desmin and SV40 T-antigen were used to confirm that both cell lines were immortal HS cells. The growth rates of both cell lines were similar and there was little difference between cell line sensitivity to zinc. MT-null IMS/MT(-) cells were more sensitive to cadmium and mercury, although both cell lines accumulated almost equal amounts of cadmium during a 24-hr culture period. As HS cells play an important role in hepatic fibrosis and are activated by heavy metals such as cadmium or reactive oxygen, the MT-null HS cell line derived in this study should be a useful experimental model for examination of the role of MT in HS cell activation.

Metallothionein (MT) is known to play an important role in the resistance of tumor cells to cisdiamminedichloroplatinum (II) (CDDP, cisplatin). To identify non-MT factors that play important roles in CDDP resistance, we established CDDP-resistant cell lines from simian virus 40-transformed MT null cells. Subclones of CDDP-resistant MT null cells, designated as MKCr-3, -12, and -18, exhibited 24- to 62-fold resistance to CDDP compared with parental cells. The contents of glutathione (GSH) and the activities of GSH-related enzymes and antioxidant enzymes in MKCr-12 and -18 were higher than those in parental cells. However, MKCr-3 cells did not show any significant increase in the levels of GSH nor in enzyme activities except for superoxide dismutase. Accumulation of platinum in CDDP-resistant subclones was 15-26% of that in parental cells 24 h after administration of CDDP. Eleven other subclones of CDDP-resistant MT null cells also exhibited low accumulation of platinum. These results suggest that the decreased accumulation of CDDP may be a predominant factor in the resistance to CDDP in the absence of MT, an intracellular metal- and free radical-scavenger.

Intestinal flora plays an important role in the decomposition and fecal excretion of methylmercury. The assumed mechanism is that decomposition of organic mercury (o-Hg) to inorganic mercury (i-Hg) by intestinal flora in cecum decreases the reabsorption of mercury in large intestines. To confirm this hypothesis, we examined the large intestinal mercury absorption in vitro from intestinal contents of methylmercury administered mice. Methylmercury (2 mg Hg/kg) was administered intraperitoneally to adult female mice, and the contents of small intestine (ileum) and cecum were taken out 24 hr after the administration. Ratios of organic mercury to total mercury in small intestinal content and cecal content were 86% and 49%, respectively. Contents fo both samall intestine and cecum were packed into cecum and colon of normal mice, respectively, and were incubated in a medium (Tyrode’s solution) at 37°C for 2 hr. 73% of total mercury (t-Hg) was absorbed from small intestinal content into the cecum and medium after the incubation. On the other hand, only 34% of t-Hg was absorbed from cecal content; most i-Hg remained unabsorbed. However, the absorption rate of t-Hg increased to 57% when the cecal content from antibiotics treated mice was used, because of the high percentage (90%) of o-Hg contained. These results suggest that o-Hg in small intestinal and cecal contents can be reabsorbed by cecum and colon in vivo, and that the decomposition of o-Hg to i-Hg by intestinal flora decreases the intestinal reabsorption rate of t-Hg in methylmercury exposed mice.

Because precise information as to the toxicity of vanadium is required for practical use of vanadium compounds as antidiabetic drugs, we examined vanadium toxicity in mice fed normal diet or high-fat diet (C57BL/6N, male, 7 weeks) by oral administration of ammonium metavanadate (AMV) with a maximum dose of 20 mgV/kg/day. Marked lipid accumulation in hepatocytes, renal epithelial cells, and mucosal epithelial cells of the small and large intestines and severe degeneration, necrosis, and loss of mucosal epithelial cells in the small intestine were observed. These pathological changes were more severe in mice fed high-fat diet than mice fed normal diet, and the intensity of the changes increased with increase in the administered dose of AMV. By electron microscopy, the number and size of lipid droplets in hepatocytes were increased. In the small intestine, a TUNEL assay showed a decreased number of positive cells, and positive cells for acrolein immunohistochemistry were observed specifically in the mucosal epithelial cells indicating degeneration and necrosis in the AMV-treated group, suggesting that a possible factor responsible for cell necrosis in the small intestine could be oxidative stress. In conclusion, AMV may impair cellular lipid metabolism, resulting in lipid accumulation, and induce mucosal epithelial cell necrosis in the small intestine.

Intestinal flora plays important roles in the decomposition and excretion of organomercurial methylmercury, and these roles in decomposition and excretion of organomercurial phenylmercury were examined in mice given antibiotics orally. Female ICR mice were given antibiotic neomycin and chloramphenicol in drinking water for 2 d before phenylmercury administration in order to decrease intestinal flora, and the antibiotics were given throughout the experiment. Phenylmercuric acetate (2mg Hg/kg body weight) was administered to mice intraperitoneally, and feces and urine were collected daily for 4 d. Mice were sacrificed 5 d after phenylmercury administration. There were no differences between the control and the antibiotic-treated mice in the percentage of inorganic mercury to total mercury in feces, urine, liver or kidney. Total mercury excreted in feces and urine did not differ between the antibiotic-treated mice and the control mice. These results suggest that intestinal flora does not play a role in the decomposition and excretion of phenylmercury in mice.

Metallothionein (MT) is a small sulfydryl-rich protein that binds to and is inducible by heavy metals such as mercury, cadmium, zinc, and copper. However, little is known about the induction of MT by trivalent metals except for bismuth. In this study, we examined the induction of MT synthesis by cerium, a trivalent lanthanoid metal. Administration of cerium chloride (CeCl3) to mice resulted in accumulation of cerium and induction of MT in the liver in a dose-dependent manner. Distribution profiles of metals in the soluble fraction of the liver of CeCl3-treated mice analyzed by high performance liquid chromatography/inductively coupled argon plasma-mass spectrometry (HPLC/ICP-MS) demonstrated that the metal bound to MT-I and MT-II was zinc, but not cerium. Administration of CeCl3 caused increases in the activities of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) and the levels of serum amyloid A (SAA), an acute phase protein. Among inflammatory cytokines examined, interleukin 6 (IL-6) exhibited a marked increase in the serum at 3 h after the CeCl3 administration. In order to evaluate the involvement of IL-6 in the induction of MT by cerium, we examined MT induction by CeCl3 in IL-6 null mice. Both the induction of hepatic MT and the increases in SAA levels were markedly suppressed in IL-6 null mice. These results suggest that IL-6 plays an important role in the induction of hepatic MT by cerium.