Methylmercury selectively damages the central nervous system (CNS). The tumor necrosis factor (TNF) superfamily includes representative cytokines that participate in the inflammatory response as well as cell survival, and apoptosis. In this study, we found that administration of methylmercury selectively induced TNF-α expression in the brain of mice. Although the accumulated mercury concentration in the liver and kidneys was greater than in the brain, TNF-α expression was induced to a greater extent in brain. Thus, it is possible that there may exist a selective mechanism by which methylmercury induces TNF-α expression in the brain. We also found that TNF-α expression was induced by methylmercury in C17.2 cells (mouse neural stem cells) and NF-κB may participate as a transcription factor in that induction. Further, we showed that the addition of TNF-α antagonist (WP9QY) reduced the toxicity of methylmercury to C17.2 cells. In contrast, the addition of recombinant TNF-α to the culture medium decreased the cell viability. We suggest that TNF-α may play a part in the selective damage of the CNS by methylmercury. Furthermore, our results indicate that the higher TNF-α expression induced by methylmercury maybe the cause of cell death, as TNF-α binds to its receptor after being released extracellularly.

We recently found that tumor necrosis factor-α (TNF-α) may be involved in neuronal cell death induced by methylmercury in the mouse brain. Here, we examined the cells involved in the induction of TNF-α expression by methylmercury in the mouse brain by in situ hybridization. TNF-α-expressing cells were found throughout the brain and were identified as microglia by immunostaining for ionized calcium binding adaptor molecule 1 (Iba1). Methylmercury induced TNF-α expression in mouse primary microglia and mouse microglial cell line BV2. Knockdown of apoptosis signal-regulating kinase 1 (ASK1), an inflammatory cytokine up-regulator that is responsible for reactive oxygen species (ROS), decreased methylmercury-induced TNF-α expression through decreased phosphorylation of p38 MAP kinase in BV2 cells. Suppression of methylmercury-induced reactive oxygen species (ROS) by antioxidant treatment largely abolished the induction of TNF-α expression and phosphorylation of p38 by methylmercury in BV2 cells. Finally, in mouse brain slices, the TNF-α antagonist (WP9QY) inhibited neuronal cell death induced by methylmercury, as did the p38 inhibitor SB203580 and liposomal clodronate (a microglia-depleting agent). These results indicate that methylmercury induces mitochondrial ROS that are involved in activation of the ASK1/p38 pathway in microglia and that this is associated with induction of TNF-α expression and neuronal cell death.

We have previously demonstrated that the overexpression of enzymes involved in the production of pyruvate, enolase 2 (Eno2) and D-lactate dehydrogenase (Dld3) renders yeast highly sensitive to methylmercury and that the promotion of intracellular pyruvate synthesis may be involved in intensifying the toxicity of methylmercury. In the present study, we showed that the addition of pyruvate to culture media in non-toxic concentrations significantly enhanced the sensitivity of yeast and human neuroblastoma cells to methylmercury. The results also suggested that methylmercury promoted the transport of pyruvate into mitochondria and that the increased pyruvate concentrations in mitochondria were involved in intensifying the toxicity of methylmercury without pyruvate being converted to acetyl-CoA. Furthermore, in human neuroblastoma cells, methylmercury treatment alone decreased the mitochondrial membrane potential and the addition of pyruvate led to a further significant decrease. In addition, treatment with N-acetylcysteine (an antioxidant) significantly alleviated the toxicity of methylmercury and significantly inhibited the intensification of methylmercury toxicity by pyruvate. Based on these data, we hypothesize that methylmercury exerts its toxicity by raising the level of pyruvate in mitochondria and that mitochondrial dysfunction and increased levels of reactive oxygen species are involved in the action of pyruvate.

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The male reproductive system is being recognized as toxic targets of nanoparticles including titanium dioxide nanoparticles (TiNP). Most of these reports are, however, obtained from the results of long-term exposure of TiNP. In this study, we diversely examined the acute effects of TiNP on the male reproductive system. Male C57BL/6J mice were administered a single intravenous injection of TiNP (10, 50 mg/kg), and were sacrificed at 1, 3, and 9 days post-injection. Testicular functions (estimated by sperm motility and sperm number) were measured via computer-assisted sperm analysis (CASA). Results indicated that sperm motility was significantly reduced from 1 day following TiNP injection (in both dose), and this reduction persisted up to 9 days post-TiNP injection (10 mg/kg injection group). Interestingly, we observed no significant decrease in sperm numbers in both the testis and the cauda epididymis in either treatment groups during the course of the experiment. Therefore, we hypothesized that TiNP may target the mature spermatozoa. In addition, sperm suspensions directly incubated with TiNP showed reduced sperm motility, [³H]-thymidine incorporation, and ATP level. Our results indicated that TiNP possesses “biphasic effects”; the obstacles to mature sperms (short term effect) in addition to the impairment in testis (long-term effect).

Methylmercury (MeHg) is a causative agent of Minamata disease and an environmental pollutant that comprises a large portion of organically occurring mercury. Many aspects of the biological defense mechanisms against MeHg toxicity remain unclear. Recently, nuclear factor-E2-related factor 2 (Nrf2), heat shock factor protein 1 (Hsf1), and hydrogen sulfide were identified as intracellular defense factors against MeHg toxicity. These findings suggest that novel biological defense mechanisms against MeHg toxicity exist in the living organism. In addition, the expression of downstream genes that mediate activation of the transcription factors Nrf2 and Hsf1 was markedly induced by MeHg treatment, suggesting that this action is involved in the reduction of MeHg toxicity. On the other hand, when the gaseous form of hydrogen sulfide (H2S) binds directly to MeHg, bismethylmercury sulfide (MeHg-S-HgMe) as a low toxicity metabolite is formed. This suggests the involvement of the gaseous form of H2S in the reduction of MeHg toxicity. In this topic, we summarize the roles of factors involved in novel biological defense mechanisms against MeHg toxicity.