It is widely recognized that heavy metals exert many of their toxic effects via binding to thiols. Moreover, thiol-containing compounds, especially GSH, are critical for heavy metal detoxifi cation and elimination. This is particularly true for mercury, and the term mercaptan is a synonym for thiol-containing compounds, which “cap- ture mercury.” Since thiols play a central role in maintaining cellular redox status, it is not surprising that mercury and other heavy metals would disrupt redox status. Moreover, cell type-specifi c differences in thiol metabolism, as described above, can lead to differences in vulnerability to heavy metals.
Mercury exists in elemental (Hg•), inorganic (Hg2+), or organic (e.g., methyl-
mercury or ethylmercury) states. Hg• is liquid at room temperature and has a high vapor pressure, which allows it to readily enter the gas phase. These unique physical properties increase the spread of mercury throughout the earth and its atmosphere and also facilitate its use in industrial products such as liquid electrical switches and light bulbs. Methylmercury from maternal seafood ingestion and dental amalgams is the primary source of in utero exposure for infants, while ethylmercury from the vaccine preservative thimerosal can be a postnatal source of exposure. Vaccination- associated mercury exposure signifi cantly increased starting in the late 1980s, but was dramatically reduced following a 1999 FDA report (Centers for Disease Control and Prevention, 1999), which led to the availability of nominally thimerosal-free infant vaccines starting in 2001. Removal of thimerosal was not, however, associ- ated with a decrease in rising autism rates (Schechter and Grether, 2008), casting doubt on a causative role for thimerosal (Fonbonne, 2008). Nonetheless, since epi- genetic effects of toxic exposures can be transmitted across generations, signifi cant concerns remain about the impact of mercury exposure during the two previous decades. In addition, aluminum, which shares many of the effects of mercury on thiol metabolism and redox status (Sharma and Mishra, 2006; Verstraeten et al., 2008), remains as an adjuvant additive in a number of vaccines at levels much higher than previous levels of mercury. Lead exposure, from paint, dust, contaminated soil, or other sources is an additional, potentially important contributor to neurotoxicity and autism via its effects on redox status (Quig, 1998; Verstraeten et al., 2008).
Organomercurials have greater access to the brain than inorganic mercury, since methyl and ethyl groups increase hydrophobic character and facilitate diffusion across the blood–brain barrier. However, the methyl and ethyl groups dissociate from mer- cury, leaving inorganic mercury trapped within the brain compartment, where it can remain for years. Studies in nonhuman primates showed that a greater proportion of inorganic mercury remained in the brain from thimerosal than from methylmercury, consistent with the weaker chemical bond of mercury to the ethyl group (Burbacher et al., 2005). Lacking methyl or ethyl groups, aluminum has an intrinsically lower
ability to cross the blood–brain barrier than organomercurials, but signifi cant levels in brain can be detected after vaccination (Flarend et al., 1997).
Within the brain compartment, mercury and other metals affect thiol metabolism in different cell types, including pluripotent stem cells, neurons, astrocytes, micro- glia, and oligodendrocytes. Under mild oxidative stress conditions, an increased proportion of pluripotent stem cells become astrocytes, whereas mild reducing con- ditions increase the proportion of neuronal cells (Prozorovski et al., 2008). Since astrocytes serve as reservoirs of GSH and provide cysteine for neurons (Figure 7.1), this mode of regulation appears to adjust cell fate in response to prevailing redox conditions, and heavy metal-induced oxidative stress would reduce neuronal development. Neuronal stem cells are particularly sensitive to mercury, and low nanomolar concentrations of methylmercury activate caspase-dependent apopto- sis (Tamm et al., 2006). Heavy metal-induced oxidative stress in oligodendrocytes can lead to impaired myelination (Crang and Jacobson, 1982), while it can lead to activation and the release of proinfl ammatory cytokines in microglia (Kim and de Vellis, 2005). Importantly, each of these conditions has been observed in the brain of children with autism.
In 2004, our group fi rst described the potent inhibitory effects of mercury, thimerosal, aluminum, and lead on methylation and methionine synthase activity in SH-SY5Y human neuronal cells (Waly et al., 2004). Subsequently, we determined that inhibition refl ected the ability of these heavy metals to lower GSH levels (M. Waly et al., unpublished observation), resulting in decreased synthesis of meth- ylcobalamin (methylB12), which is required for methionine synthase activity in these neuronal cells, as illustrated in Figure 7.1. These heavy metals potently inhibit EAAT3-mediated uptake of cysteine, which accounts for their ability to decrease GSH, methylB12, and methionine synthase activity. Together, these studies illustrate the critical role of EAAT3 in regulating redox status and methylation activity in human neuronal cells, as well as their vulnerability to heavy metals.
An important breakthrough in understanding the molecular mechanism of mer- cury toxicity was provided from studies carried out by Holmgren and colleagues at the Karolinska Institute in Sweden (Carvalho et al., 2008). They compared the potency of inorganic mercury and methylmercury to inhibit several enzymes, each of which promote a reduced intracellular redox state, including thioredoxin, thiore- doxin reductase, glutathione reductase, and glutaredoxin. Among these, thioredoxin and thioredoxin reductase showed exceptionally high sensitivity to both mercury compounds, strongly suggesting that they are primary targets for mercury-induced neurotoxicity. Thioredoxin has multiple activities, including the ability to release GSH from glutathionylated proteins (i.e., proteins with a thiol-bound GSH), while thioredoxin reductase, a selenoprotein, serves to reactivate thioredoxin after it has carried out deglutathionylation (Figure 7.2). The extent of protein glutathionylation refl ects the level of cellular oxidative stress, and mercury inhibition of the thiore- doxin system will promote the accumulation of glutathionylated proteins, producing and sustaining a state of high oxidative stress. The ultrahigh affi nity of mercury for selenoproteins has long been recognized, and selenium supplementation has been suggested as a treatment for mercury toxicity.