SafeMinds Finding the Truth February 2006 #2

Over Half A Million Dollars of Research Sponsored
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The Coalition for SafeMinds (Sensible Action For Ending Mercury-Induced Neurological Disorders) is a private nonprofit organization founded in 2000 to raise awareness and investigate the risks of mercury exposure to infants, children and pregnant women. Mercury is a well known neurotoxin and exposure can result in significant disability, particularly in the fetus, infants and children since their brains are still growing and developing.

Since its inception in 2000, Safe Minds has sponsored over one half million dollars in research related specifically to mercury and adverse neurological outcomes, including autism. This level of financial commitment establishes Safe Minds as the largest private non-profit organization funding mercury and autism related research. Below is a list of recently funded Safe Minds investigations. Research proposals are accepted throughout the year. For additional information contact us.

Neurotoxic Effects of Postnatal Thimerosal are Mouse Strain Dependant, Mady Hornig, MD, PhD, Columbia University

The developing brain is uniquely susceptible to the neurotoxic hazard posed by mercurials. Host differences in maturation, metabolism, nutrition, sex, and autoimmunity influence outcomes. How population-based variability affects the safety of the ethylmercury-containing vaccine preservative, thimerosal, is unknown. Reported increases in the prevalence of autism, a highly heritable neuropsychiatric condition, are intensifying public focus on environmental exposures such as thimerosal. Immune profiles and family history in autism are frequently consistent with autoimmunity. We hypothesized that autoimmune propensity influences outcomes in mice following thimerosal challenges that mimic routine childhood immunizations. Autoimmune disease-sensitive SJL/J mice showed growth delay; reduced locomotion; exaggerated response to novelty; and densely packed, hyperchromic hippocampal neurons with altered glutamate receptors and transporters. Strains resistant to autoimmunity, C57BL/6J and BALB/cJ, were not susceptible. These findings implicate genetic influences and provide a model for investigating thimerosal-related neurotoxicity.

Richard C. Deth, PhD, Northeastern University

It has been proposed that the ethylmercury- containing vaccine preservative thimerosal may contribute to autism, and our earlier studies demonstrated the ability of thimerosal to inhibit methionine synthase-dependent phospholipid methylation (PLM) in cultured human neuroblastoma cells. To investigate the possible contribution of this action of thimerosal to autism, we compared its ability to inhibit PLM measured with [14C]-formate, which labels the cellular pool of 5- methyltetrahydrofolate and therefore selectively measures methionine synthase-dependent PLM. PLM was measured in immortalized lymphoblasts from same-sex siblings who were discordant for autism, as obtained from the Autism Genetic Resource Exchange (AGRE). Basal PLM was not significantly different between lymphoblasts from autistic and non-autistic siblings. Thimerosal (100 nM) did not significantly affect PLM in lymphoblasts from non-autistic siblings, but significantly reduced PLM in lymphoblasts from autistic subjects (p < 0.05). Analysis of MTHFR and transcobalamin polymorphisms in a sample of 18 sib- pairs did not reveal a significant genetic pattern of association with autism. Preliminary studies indicate a trend for autistic subjects to exhibit higher rates of mitochondrial oxygen consumption. Taken together, our results to date provide evidence that methionine synthase-dependent methylation is more sensitive to thimerosal in cells from autistic children, consistent with a potential role of thimerosal in causing autism.

Mechanisms of Thimerosal Toxicity, Jill James, PhD, University of Arkansas

Children with autism have increased vulnerability to pro-oxidant exposures such as ethyl mercury in Thimerosal as a result of increased frequency of genetic polymorphisms that reduce the synthesis of cysteine and glutathione, the major metabolites involved in the detoxification and excretion of mercury. Dr. James will extend preliminary data on plasma levels in children with autism by measuring intracellular levels of thiol metabolites and selected enzyme activities in lymphoblastoid cell lines derived from children with autism and unrelated control children. Iintracellular metabolic profiles will be correlated with genetic profiles of specific polymorphisms that negatively affect methionine, cysteine, and glutathione synthesis. These experiments will allow us to determine whether intracellular metabolites and related enzyme activities are abnormal in children with autism compared to controls and whether the intracellular metabolic profile reflects the profile previously observed in plasma (see preliminary data). If the observed metabolic profiles are associated with increased frequency of polymorphisms in the same metabolic pathway, it will provide support for our hypothesis that children with autism have a genetic vulnerability to heavy metal toxicity.

In addition, we will expose lymphoblastoid cells derived from autistic children and unrelated controls to increasing doses of thimerosal (nanomolar to micromolar levels) and define individual dose-response curves in terms of cytotoxicity, glutathione depletion, and DNA damage. In addition, we will determine whether subtoxic doses of ethylmercury in the presence of subtoxic levels of an additional pro- oxidant heavy metal such as lead, will interact synergistically to reach a threshold of toxicity. If lymphocytes from autistic children exhibit increased sensitivity to Thimerosal toxicity in culture compared to cells from normal children, the dose-response curve should be shifted to the left. An interaction between subtoxic doses of thimerosal and other heavy metals in autistic children, but not normal children, would further support the hypothesis that autistic children have an increased vulnerability to pro-oxidant exposures. In addition, we will be able to determine whether an increase in thimerosal sensitivity is associated with abnormal genetic and metabolic profiles and glutathione depletion. If confirmed, these results would support for the hypothesis that children with autism have an increased sensitivity to thimerosal as a result of reduced intracellular levels of cysteine and glutathione, and consequently, reduced capacity to detoxify and excrete ethylmercury.

Thimerosal Neurotoxicity, Thomas Burbacher, PhD, University of Washington

The specific aim of this research project is to determine the extent of changes in the absolute number of neurons, astrocytes and microglia within six specific regions of the central nervous system of the nonhuman primate (NHP) Macaca fascicularis following a known low-level thimerosal (ethylmercury) exposure. The changes in absolute cell number will be determined by use of modern designed-based stereological methods utilizing the optical disector and fractionator principles. The experimental design will test the hypothesis that exposure to thimerosal will correlate with changes in cell number within specific CNS regions, suggesting thimerosal may cause structural damage to the CNS. The six regions to be examined will include sub regions of the frontal cortex (principle sulcus- memory processing, higher function), occipital pole (calcarine sulcus-visual cortex), thalamus (functional integration), hippocampus (memory), amygdala (emotion integration), and the cerebellum (coordination, motor skills). These regions have been selected for investigation because they are well-characterized anatomical regions of the NHP brain, and extensive information about these regions has been developed describing CNS effects of methylmercury exposure. Ultimately, the results from the investigation proposed in this study will help clarify issues about the safety of ethylmercury exposure. In addition, this proposed project will seek to determine the distribution of inorganic mercury within the six specific brain regions by use of an autometallographic technique capable of localizing mercury deposits within specific cell types in histology tissue sections. Previous mercury quantification has demonstrated that inorganic mercury is present in the brain of these animals following thimerosal exposure, suggesting ethylmercury may be demethylated in the brain in a manner similar to demethylation of methylmercury that we have previously reported. Prior to sampling of the brains for the stereology and autometallography methods described above, the intact brains will be scanned with magnetic resonance imaging (MRI) techniques to allow for the future determination of potential volumetric changes of (i) total brain volume, (ii) all segmented divisions of total brain volume (cerebral cortex, cerebral white matter, cerebellum, caudate, globus pallidus– putamen, diencephalon, brainstem), (iii) lobes of the cerebral cortex and (iv) individual cortical lobe sub regions (parcellation units) for the entire cerebral cortex. In addition, specific anti-body based histochemistry methods will be used to identify reactive glial cells and immune cells within these brain samples.