The following 68 medical journal studies support the link between vaccination and autism. So the next time a doctor or government official tells you that this link is non-existent, please point them to this list of references.
[UPDATE 2018, PLEASE NOTE: many of the links below no longer go to the studies described. We’ll leave them here so you can see the titles and can search for them yourself if you wish (please understand we are volunteers and don’t have unlimited time to check that all links on old blog posts are working).]
Please visit our Autism And Vaccines page for an up-to-date list of studies.
Thank you to Ginger Taylor and the Age of Autism for compiling these references.
1. Metabolic biomarkers of increased oxidative stress and impaired methylation capacity in children with autism
American Journal of Clinical Nutrition, Vol. 80, No. 6, 1611-1617, December 2004
Department of Pediatrics, University of Arkansas for Medical Sciences, and the Arkansas Children’s Hospital Research Institute
Background: Autism is a complex neurodevelopmental disorder that usually presents in early childhood and that is thought to be influenced by genetic and environmental factors. Although abnormal metabolism of methionine and homocysteine has been associated with other neurologic diseases, these pathways have not been evaluated in persons with autism.
Objective: The purpose of this study was to evaluate plasma concentrations of metabolites in the methionine transmethylation and transsulfuration pathways in children diagnosed with autism.
Design: Plasma concentrations of methionine, S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH), adenosine, homocysteine, cystathionine, cysteine, and oxidized and reduced glutathione were measured in 20 children with autism and in 33 control children. On the basis of the abnormal metabolic profile, a targeted nutritional intervention trial with folinic acid, betaine, and methylcobalamin was initiated in a subset of the autistic children.
Results: Relative to the control children, the children with autism had significantly lower baseline plasma concentrations of methionine, SAM, homocysteine, cystathionine, cysteine, and total glutathione and significantly higher concentrations of SAH, adenosine, and oxidized glutathione. This metabolic profile is consistent with impaired capacity for methylation (significantly lower ratio of SAM to SAH) and increased oxidative stress (significantly lower redox ratio of reduced glutathione to oxidized glutathione) in children with autism. The intervention trial was effective in normalizing the metabolic imbalance in the autistic children.
Conclusions: An increased vulnerability to oxidative stress and a decreased capacity for methylation may contribute to the development and clinical manifestation of autism.
2. Porphyrinuria in childhood autistic disorder: Implications for environmental toxicity
Toxicology and Applied Pharmacology, 2006
Robert Natafa, Corinne Skorupkab, Lorene Ametb, Alain Lama, Anthea Springbettc and Richard Lathed, aLaboratoire Philippe Auguste, Paris, France, Association ARIANE, Clichy, France, Department of Statistics, Roslin Institute, Roslin, UK, Pieta Research,
This new study from France utilizes a new and sophisticated measurement for environmental toxicity by assessing porphyrin levels in autistic children. It provides clear and unequivocal evidence that children with autism spectrum disorders are more toxic than their neurotypical peers.
Excerpt: “Coproporphyrin levels were elevated in children with autistic disorder relative to control groups…the elevation was significant. These data implicate environmental toxicity in childhood autistic disorder.”
Abstract: To address a possible environmental contribution to autism, we carried out a retrospective study on urinary porphyrin levels, a biomarker of environmental toxicity, in 269 children with neurodevelopmental and related disorders referred to a Paris clinic (2002–2004), including 106 with autistic disorder. Urinary porphyrin levels determined by high-performance liquid chromatography were compared between diagnostic groups including internal and external control groups. Coproporphyrin levels were elevated in children with autistic disorder relative to control groups. Elevation was maintained on normalization for age or to a control heme pathway metabolite (uroporphyrin) in the same samples. The elevation was significant (P < 0.001). Porphyrin levels were unchanged in Asperger’s disorder, distinguishing it from autistic disorder. The atypical molecule precoproporphyrin, a specific indicator of heavy metal toxicity, was also elevated in autistic disorder (P < 0.001) but not significantly in Asperger’s. A subgroup with autistic disorder was treated with oral dimercaptosuccinic acid (DMSA) with a view to heavy metal removal. Following DMSA there was a significant (P = 0.002) drop in urinary porphyrin excretion. These data implicate environmental toxicity in childhood autistic disorder.
3. Uncoupling of ATP-mediated Calcium Signaling and Dysregulated IL-6 Secretion in Dendritic Cells by Nanomolar Thimerosal
Environmental Health Perspectives, July 2006.
Samuel R. Goth, Ruth A. Chu Jeffrey P. Gregg
This study demonstrates that very low-levels of Thimerosal can contribute to immune system disregulation.
Excerpt: “Our findings that DCs primarily express the RyR1 channel complex and that this complex is uncoupled by very low levels of THI with dysregulated IL-6 secretion raise intriguing questions about a molecular basis for immune dyregulation and the possible role of the RyR1 complex in genetic susceptibility of the immune system to mercury.”
4. Comparison of Blood and Brain Mercury Levels in Infant Monkeys Exposed to Methylmercury or Vaccines Containing Thimerosal
Environmental Health Perspectives, Aug 2005.
Thomas Burbacher, PhD [University of Washington].
This study demonstrates clearly and unequivocally that ethyl mercury, the kind of mercury found in vaccines, not only ends up in the brain, but leaves double the amount of inorganic mercury as methyl mercury, the kind of mercury found in fish. This work is groundbreaking because little is known about ethyl mercury, and many health authorities have asserted that the mercury found in vaccines is the “safe kind.” This study also delivers a strong rebuke of the Institute of Medicine’s recommendation in 2004 to no longer pursue the mercury-autism connection.
Excerpt: “A recently published IOM review (IOM 2004) appears to have abandoned the earlier recommendation [of studying mercury and autism] as well as back away from the American Academy of Pediatrics goal [of removing mercury from vaccines]. This approach is difficult to understand, given our current limited knowledge of the toxicokinetics and developmental neurotoxicity of thimerosal, a compound that has been (and will continue to be) injected in millions of newborns and infants.”
6. Increases in the number of reactive glia in the visual cortex of Macaca fascicularis following subclinical long-term methyl mercury exposure.
Toxicology and Applied Pharmacology, 1994
Charleston JS, Bolender RP, Mottet NK, Body RL, Vahter ME, Burbacher TM., Department of Pathology, School of Medicine, University of Washington
The number of neurons, astrocytes, reactive glia, oligodendrocytes, endothelia, and pericytes in the cortex of the calcarine sulcus of adult female Macaca fascicularis following long-term subclinical exposure to methyl mercury (MeHg) and mercuric chloride (inorganic mercury; IHg) has been estimated by use of the optical volume fractionator stereology technique. Four groups of monkeys were exposed to MeHg (50 micrograms Hg/kg body wt/day) by mouth for 6, 12, 18, and 12 months followed by 6 months without exposure (clearance group). A fifth group of monkeys was administered IHg (as HgCl2; 200 micrograms Hg/kg body wt/day) by constant rate intravenous infusion via an indwelling catheter for 3 months. Reactive glia showed a significant increase in number for every treatment group, increasing 72% in the 6-month, 152% in the 12-month, and 120% in the 18-month MeHg exposed groups, and the number of reactive glia in the clearance group remained elevated (89%). The IHg exposed group showed a 165% increase in the number of reactive glia. The IHg exposed group and the clearance group had low levels of MeHg present within the tissue; however, the level of IHg was elevated in both groups. These results suggest that the IHg may be responsible for the increase in reactive glia. All other cell types, including the neurons, showed no significant change in number at the prescribed exposure level and durations. The identities of the reactive glial cells and the implications for the long-term function and survivability of the neurons due to changes in the glial population following subclinical long-term exposure to mercury are discussed.
7. Neuroglial Activation and Neuroinflammation in the Brain of Patients with Autism
Annals of Neurology, Feb 2005.
Diana L. Vargas, MD [Johns Hopkins University].
This study, performed independently and using a different methodology than Dr. Herbert (see above) reached the same conclusion: the brains of autistic children are suffering from inflammation.
Excerpt: “Because this neuroinflammatory process appears to be associated with an ongoing and chronic mechanism of CNS dysfunction, potential therapeutic interventions should focus on the control of its detrimental effects and thereby eventually modify the clinical course of autism.”
8. Autism: A Brain Disorder, or A Disorder That Affects the Brain?
Clinical Neuropsychiatry, 2005
Martha R. Herbert M.D., Ph.D., Harvard University
Autism is defined behaviorally, as a syndrome of abnormalities involving language, social reciprocity and hyperfocus or reduced behavioral flexibility. It is clearly heterogeneous, and it can be accompanied by unusual talents as well as by impairments, but its underlying biological and genetic basis in unknown. Autism has been modeled as a brain-based, strongly genetic disorder, but emerging findings and hypotheses support a broader model of the condition as a genetically influenced and systemic. These include imaging, neuropathology and psychological evidence of pervasive (and not just specific) brain and phenotypic features; postnatal evolution and chronic persistence of brain, behavior and tissue changes (e.g. inflammation) and physical illness symptomatology (e.g. gastrointestinal, immune, recurrent infection); overlap with other disorders; and reports of rate increases and improvement or recovery that support a role for modulation of the condition by environmental factors (e.g. exacerbation or triggering by toxins, infectious agents, or others stressors, or improvement by treatment). Modeling autism more broadly encompasses previous work, but also encourages the expansion of research and treatment to include intermediary domains of molecular and cellular mechanisms, as well as chronic tissue, metabolic and somatic changes previously addressed only to a limited degree. The heterogeneous biologies underlying autism may conceivably converge onto the autism profile via multiple mechanisms on the one hand and processing and connectivity abnormalities on the other may illuminate relevant final common pathways and contribute to focusing on the search for treatment targets in this biologically and etiologically heterogeneous behavioral syndrome.
9. Activation of Methionine Synthase by Insulin-like Growth Factor-1 and Dopamine: a Target for Neurodevelopmental Toxins and Thimerosal
Molecular Psychiatry, July 2004.
Richard C. Deth, PhD [Northeastern University].
This study demonstrates how Thimerosal inhibits methylation, a central driver of cellular communication and development. Excerpt:
“The potent inhibition of this pathway [methylation] by ethanol, lead, mercury, aluminum, and thimerosal suggests it may be an important target of neurodevelopmental toxins.”
10. Validation of the Phenomenon of Autistic Regression Using Home Videotapes
Archives of General Psychiatry, 2005
Emily Werner, PhD; Geraldine Dawson, PhD, University of Washington
Objective To validate parental report of autistic regression using behavioral data coded from home videotapes of children with autism spectrum disorder (ASD) vs typical development taken at 12 and 24 months of age.
Design Home videotapes of 56 children’s first and second birthday parties were collected from parents of young children with ASD with and without a reported history of regression and typically developing children. Child behaviors were coded by raters blind to child diagnosis and regression history. A parent interview that elicited information about parents’ recall of early symptoms from birth was also administered.
Setting Participants were recruited from a multidisciplinary study of autism conducted at a major university.
Participants Fifteen children with ASD with a history of regression, 21 children with ASD with early-onset autism, and 20 typically developing children and their parents participated.
Main Outcome Measures Observations of children’s communicative, social, affective, repetitive behaviors, and toy play coded from videotapes of the toddlers’ first and second birthday parties.
Results Analyses revealed that infants with ASD with regression show similar use of joint attention and more frequent use of words and babble compared with typical infants at 12 months of age. In contrast, infants with ASD with early onset of symptoms and no regression displayed fewer joint attention and communicative behaviors at 12 months of age. By 24 months of age, both groups of toddlers with ASD displayed fewer instances of word use, vocalizations, declarative pointing, social gaze, and orienting to name as compared with typically developing 24-month-olds.
Parent interview data suggested that some children with regression displayed difficulties in regulatory behavior before the regression occurred.
Conclusion This study validates the existence of early autistic regression.
11. Blood Levels of Mercury Are Related to Diagnosis of Autism: A Reanalysis of an Important Data Set
Journal of Child Neurology, Vol. 22, No. 11, 1308-1311 (2007)
M. Catherine DeSoto, PhD, Robert T. Hitlan, PhD -Department of Psychology, University of Northern Iowa, Cedar Falls, Iowa
Excerpt: “We have reanalyzed the data set originally reported by Ip et al. in 2004 and have found that the original p value was in error and that a significant relation does exist between the blood levels of mercury and diagnosis of an autism spectrum disorder. Moreover, the hair sample analysis results offer some support for the idea that persons with autism may be less efficient and more variable at eliminating mercury from the blood.”
12. Empirical Data Confirm Autism Symptoms Related to Aluminum and Acetaminophen Exposure
Entropy, November 7, 2012
Stephanie Seneff, Robert M. Davidson and Jingjing Liu
Computer Science and Artificial Intelligence Laboratory, Massachusetts Institute of Technology, Cambridge, MA 02139, USA, Internal Medicine Group Practice, PhyNet, Inc., Longview, TX 75604, USA
Abstract: Autism is a condition characterized by impaired cognitive and social skills, associated with compromised immune function. The incidence is alarmingly on the rise, and environmental factors are increasingly suspected to play a role. This paper investigates word frequency patterns in the U.S. CDC Vaccine Adverse Events Reporting System (VAERS) database. Our results provide strong evidence supporting a link between autism and the aluminum in vaccines. A literature review showing toxicity of aluminum in human physiology offers further support. Mentions of autism in VAERS increased steadily at the end of the last century, during a period when mercury was being phased out, while aluminum adjuvant burden was being increased. Using standard log-likelihood ratio techniques, we identify several signs and symptoms that are significantly more prevalent in vaccine reports after 2000, including cellulitis, seizure, depression, fatigue, pain and death, which are also significantly associated with aluminum-containing vaccines. We propose that children with the autism diagnosis are especially vulnerable to toxic metals such as aluminum and mercury due to insufficient serum sulfate and glutathione. A strong correlation between autism and the MMR (Measles, Mumps, Rubella) vaccine is also observed, which may be partially explained via an increased sensitivity to acetaminophen administered to control fever.
13. Developmental Regression and Mitochondrial Dysfunction in a Child With Autism
Journal of Child Neurology / Volume 21, Number 2, February 2006
Jon S. Poling, MD, PhD, Department of Neurology and Neurosurgery
Johns Hopkins Hospital
This article showed that 38% of Kennedy Krieger Institute autism patients studied had one marker for impaired oxidative phosphorylation (mitochondrial dysfunction), and 47% had a second marker.
Excerpt: “Children who have (mitochondrial-related) dysfunctional cellular energy metabolism might be more prone to undergo autistic regression between 18 and 30 months of age if they also have infections or immunizations at the same time.”
14. Oxidative Stress in Autism: Elevated Cerebellar 3-nitrotyrosine Levels
American Journal of Biochemistry and Biotechnology 4 (2): 73-84, 2008
Elizabeth M. Sajdel-Sulkowska, – Dept of Psychiatry, Harvard Medical School
Shows a potential link between mercury and the autopsied brains of young people with autism. A marker for oxidative stress was 68.9% higher in autistic brain issue than controls (a statistically significant result), while mercury levels were 68.2% higher.
Excerpt: The preliminary data suggest a need for more extensive studies of oxidative stress, its relationship to the environmental factors and its possible attenuation by antioxidants in autism.”
15. Large Brains in Autism: The Challenge of Pervasive Abnormality
The Neuroscientist, Volume 11, Number 5, 2005.
Martha Herbert, MD, PhD [Harvard University].
This study helps refute the notion that the brains of autistic children are simply wired differently and notes, “neuroinflammation appears to be present in autistic brain tissue from childhood through adulthood.” Dr. Herbert suggests that chronic disease or an external environmental source (like heavy metals) may be causing the inflammation.
Excerpt: “Oxidative stress, brain inflammation, and microgliosis have been much documented in association with toxic exposures including various heavy metals…the awareness that the brain as well as medical conditions of children with autism may be conditioned by chronic biomedical abnormalities such as inflammation opens the possibility that meaningful biomedical interventions may be possible well past the window of maximal neuroplasticity in early childhood because the basis for assuming that all deficits can be attributed to fixed early developmental alterations in neural architecture has now been undermined.”
16. Evidence of Toxicity, Oxidative Stress, and Neuronal Insult in Autism
Journal of Toxicology and Environmental Health, Nov-Dec 2006.
Janet Kern, Anne Jones
“This article discusses the evidence for the case that some children with autism may become autistic from neuronal cell death or brain damage sometime after birth as result of insult; and addresses the hypotheses that toxicity and oxidative stress may be a cause of neuronal insult in autism… the article discusses what may be happening over the course of development and the multiple factors that may interplay and make these children more vulnerable to toxicity, oxidative stress, and neuronal insult.”
17. Oxidative Stress in Autism
Abha Chauhan, Ved Chauhan
This study provides a helpful overview of the growing evidence supporting the link between oxidative stress and autism.
Excerpt: “Upon completion of this article, participants should be able to: 1. Be aware of laboratory and clinical evidence of greater oxidative stress in autism. 2. Understand how gut, brain, nutritional, and toxic status in autism are consistent with greater oxidative stress. 3. Describe how anti-oxidant nutrients are used in the contemporary treatment of autism.”
18. Thimerosal Neurotoxicity is Associated with Glutathione Depletion: Protection with Glutathione Precursors
Neurotoxicology, Jan 2005.
S. Jill James, PhD [University of Arkansas].
This recent study demonstrates that Thimerosal lowers or inhibits the body’s ability to produce Glutathione, an antioxidant and the body’s primary cellular-level defense against mercury.
Excerpt: “Thimerosal-induced cytotoxicity was associated with depletion of intracellular Glutathione in both cell lines…The potential effect of Glutathione or N-acetylcysteine against mercury toxicity warrants further research as possible adjunct therapy to individuals still receiving Thimerosal-containing vaccines.”
19. Aluminum adjuvant linked to gulf war illness induces motor neuron death in mice
Neuromolecular Medicine, 2007
Christopher Shaw, Ph.D. [Department of Ophthalmology and Program in Neuroscience, University of British Columbia, Vancouver, British Columbia, Canada]
This study demonstrates the extreme toxicity of the aluminum adjuvant used as a preservative in vaccines.
Excerpt: “testing showed motor deficits in the aluminum treatment group that expressed as a progressive decrease in strength measured…Significant cognitive deficits in water-maze learning were observed in the combined aluminum and squalene group…Apoptotic neurons were identified in aluminum-injected animals that showed significantly increased activated caspase-3 labeling in lumbar spinal cord (255%) and primary motor cortex (192%) compared with the controls. Aluminum-treated groups also showed significant motor neuron loss (35%) and increased numbers of astrocytes (350%) in the lumbar spinal cord.
20. Environmental mercury release, special education rates, and autism disorder: an ecological study of Texas
Health & Place, 2006
Raymond F. Palmer, University of Texas Health Science Center
This study demonstrated the correlation between environmental mercury and autism rates in Texas.
Excerpt: “On average, for each 1,000 lb of environmentally released mercury, there was a 43% increase in the rate of special education services and a 61% increase in the rate of autism. The association between environmentally released mercury and special education rates were fully mediated by increased autism rates. This ecological study suggests the need for further research regarding the association between environmentally released mercury and developmental disorders such as autism.”
21. Autism Spectrum Disorders in Relation to Distribution of Hazardous Air Pollutants in the SF Bay Area
Environmental Health Perspectives – Vol. 114 No. 9, September, 2006
Gayle Windham, Div. of Environmental and Occupational Disease Control, California Department of Health Services
284 ASD children & 657 controls, born in 1994 in Bay Area, were assigned exposure levels by birth tract for 19 chemicals. Risks for autism were elevated by 50% in tracts with the highest chlorinated solvents and heavy metals. The highest risk compounds were mercury, cadmium, nickel, trichloroethylene, and vinyl chloride, and the risk from heavy metals was almost twice as high as solvents.
Excerpt: “Our results suggest a potential association between autism and estimated metal concentrations, and possibly solvents, in ambient air around the birth residence.”
22. A Case Series of Children with Apparent Mercury Toxic Encephalopathies Manifesting with Clinical Symptoms of Regressive Autistic Disorder
Journal of Toxicology and Environmental Health, 2007
David A. Geier, Mark R. Geier
This study reviewed the case histories and medical profiles of nine autistic children and concluded that eight of the nine children were mercury toxic and this toxicity manifested itself in a manner consistent with Autism Spectrum Disorders.
Excerpt: “…these previously normally developing children suffered mercury toxic encephalopathies that manifested with clinical symptoms consistent with regressive ASDs. Evidence for mercury intoxication should be considered in the differential diagnosis as contributing to some regressive ASDs.”
23. Attention-deficit hyperactivity disorder and blood mercury level: a case-control study in chinese children
Neuropediatrics, August 2006 – P.R. Kong [Department of Pediatrics and Adolescent Medicine, The University of Hong Kong].
This study demonstrates that blood mercury levels are higher for children with ADHD.
Excerpt: “There was significant difference in blood mercury levels between cases and controls, which persists after adjustment for age, gender and parental occupational status. The geometric mean blood mercury level was also significantly higher in children with inattentive and combined subtypes of ADHD. High blood mercury level was associated with ADHD. Whether the relationship is causal requires further studies.”
24. The Changing Prevalence of Autism In California
Journal of Autism and Developmental Disorders, April 2003
Mark Blaxill, MBA
This study helps to refute the supposition made by some researchers that autism’s epidemic may only be due to “diagnostic substitution”.
Excerpt: “They have suggested that ‘diagnostic substitution’ accounts for an apparent increase in the incidence of autism in California that is not real. This hypothesized substitution is not supported by proper and detailed analyses of the California data.”
25. Mitochondrial Energy-Deficient Endophenotype in Autism
American Journal of Biochemistry and Biotechnology 4 (2): 198-207, 2008
J. Jay Gargus and Faiqa Imtiaz
Department of Physiology and Biophysics and Department of Pediatrics, Section of Human Genetics, School of Medicine, University of California, Irvine, Arabian Diagnostics Laboratory, King Faisal Specialist Hospital and Research Centre
Abstract: While evidence points to a multigenic etiology of most autism, the pathophysiology of the disorder has yet to be defined and the underlying genes and biochemical pathways they subserve remain unknown. Autism is considered to be influenced by a combination of various genetic, environmental and immunological factors; more recently, evidence has suggested that increased
vulnerability to oxidative stress may be involved in the etiology of this multifactorial disorder.
Furthermore, recent studies have pointed to a subset of autism associated with the biochemical endophenotype of mitochondrial energy deficiency, identified as a subtle impairment in fat and carbohydrate oxidation. This phenotype is similar, but more subtle than those seen in classic mitochondrial defects. In some cases the beginnings of the genetic underpinnings of these mitochondrial defects are emerging, such as mild mitochondrial dysfunction and secondary carnitine deficiency observed in the subset of autistic patients with an inverted duplication of chromosome 15q11-q13. In addition, rare cases of familial autism associated with sudden infant death syndrome (SIDS) or associated with abnormalities in cellular calcium homeostasis, such as malignant hyperthermia or cardiac arrhythmia, are beginning to emerge. Such special cases suggest that the pathophysiology of autism may comprise pathways that are directly or indirectly involved in mitochondrial energy production and to further probe this connection three new avenues seem worthy of exploration: 1) metabolomic clinical studies provoking controlled aerobic exercise stress to expand the biochemical phenotype, 2) high-throughput expression arrays to directly survey activity of the genes underlying these biochemical pathways and 3) model systems, either based upon neuronal stem cells or model genetic organisms, to discover novel genetic and environmental inputs into these pathways.
26. Bridging from Cells to Cognition in Autism Pathophysiology: Biological
Pathways to Defective Brain Function and Plasticity
American Journal of Biochemistry and Biotechnology 4 (2): 167-176, 2008
Matthew P. Anderson, Brian S. Hooker and Martha R. Herbert
Departments of Neurology and Pathology, Harvard Medical School/Beth Israel Deaconess Medical Center, Harvard Institutes of Medicine, High Throughput Biology Team, Fundamental Science Directorate, Pacific Northwest National Laboratory, Pediatric Neurology/Center for Morphometric Analysis, Massachusetts General Hospital/Harvard Medical School, and
Center for Child and Adolescent Development, Cambridge Health Alliance/Harvard Medical School
Abstract: We review evidence to support a model where the disease process underlying autism may begin when an in utero or early postnatal environmental, infectious, seizure, or autoimmune insult triggers an immune response that increases reactive oxygen species (ROS) production in the brain that leads to DNA damage (nuclear and mitochondrial) and metabolic enzyme blockade and that these inflammatory and oxidative stressors persist beyond early development (with potential further exacerbations), producing ongoing functional consequences. In organs with a high metabolic demand such as the central nervous system, the continued use of mitochondria with damaged DNA and impaired metabolic enzyme function may generate additional ROS which will cause persistent activation of the innate immune system leading to more ROS production. Such a mechanism would self-sustain and possibly progressively worsen. The mitochondrial dysfunction and altered redox signal transduction pathways found in autism would conspire to activate both astroglia and microglia. These activated cells can then initiate a broad-spectrum proinflammatory gene response. Beyond the direct effects of ROS on neuronal function, receptors on neurons that bind the inflammatory mediators may serve to inhibit neuronal signaling to protect them from excitotoxic damage during various pathologic insults (e.g., infection). In autism, over-zealous neuroinflammatory responses could not only influence neural developmental processes, but may more significantly impair neural signaling involved in cognition in an ongoing fashion. This model makes specific predictions in patients and experimental animal models and suggests a number of targets sites of intervention. Our model of potentially reversible pathophysiological mechanisms in autism motivates our hope that effective therapies may soon appear on the horizon.
27. Heavy-Metal Toxicity—With Emphasis on Mercury
John Neustadt, ND, and Steve Pieczenik, MD, PhD
Conclusion: Metals are ubiquitous in our environment, and exposure to them is inevitable. However, not all people accumulate toxic levels of metals or exhibit symptoms of metal toxicity, suggesting that genetics play a role in their potential to damage health. Metal toxicity creates multisystem dysfunction, which appears to be mediated primarily through mitochondrial damage from glutathione depletion.
Accurate screening can increase the likelihood that patients with potential metal toxicity are identified. The most accurate screening method for assessing chronic-metals exposure and metals load in the body is a provoked urine test.
28. Evidence of Mitochondrial Dysfunction in Autism and Implications for Treatment
American Journal of Biochemistry and Biotechnology 4 (2): 208-217, 2008
Daniel A. Rossignol, J. Jeffrey Bradstreet, International Child Development Resource Center,
Abstract: Classical mitochondrial diseases occur in a subset of individuals with autism and are usually caused by genetic anomalies or mitochondrial respiratory pathway deficits. However, in many cases of autism, there is evidence of mitochondrial dysfunction (MtD) without the classic features associated with mitochondrial disease. MtD appears to be more common in autism and presents with less severe signs and symptoms. It is not associated with discernable mitochondrial pathology in muscle biopsy specimens despite objective evidence of lowered mitochondrial functioning. Exposure to environmental toxins is the likely etiology for MtD in autism. This dysfunction then contributes to a number of diagnostic symptoms and comorbidities observed in autism including: cognitive impairment, language deficits, abnormal energy metabolism, chronic gastrointestinal problems, abnormalities in fatty acid oxidation, and increased oxidative stress. MtD and oxidative stress may also explain the high male to female ratio found in autism due to increased male vulnerability to these dysfunctions.
Biomarkers for mitochondrial dysfunction have been identified, but seem widely under-utilized despite available therapeutic interventions. Nutritional supplementation to decrease oxidative stress along with factors to improve reduced glutathione, as well as hyperbaric oxygen therapy (HBOT) represent supported and rationale approaches. The underlying pathophysiology and autistic symptoms of affected individuals would be expected to either improve or cease worsening once effective treatment for MtD is implemented.
29. Proximity to point sources of environmental mercury release as a predictor of autism prevalence
Health & Place, 2008
Raymond F. Palmer, Stephen Blanchard, Robert Wood
University of Texas Health Science Center, San Antonio Department of Family and Community Medicine, Our Lady of the Lake University, San Antonio Texas, Chair, Department of Sociology
This study should be viewed as hypothesis-generating – a first step in examining the potential role of environmental mercury and childhood developmental disorders. Nothing is known about specific exposure routes, dosage, timing, and individual susceptibility. We suspect that persistent low-dose exposures to various environmental toxicants, including mercury, that occur during critical windows of neural development among genetically susceptible children (with a diminished capacity for metabolizing accumulated toxicants) may increase the risk for developmental disorders such as autism. Successfully identifying the specific combination of environmental exposures and genetic susceptibilities can inform the development of targeted prevention intervention strategies.
30. Epidemiology of autism spectrum disorder in Portugal: prevalence, clinical characterization, and medical conditions
Developmental Medicine & Child Neurology, 2007
Guiomar Oliveira MD PhD, Centro de Desenvolvimento da Criança, Hospital Pediátrico de Coimbra; Assunção Ataíde BSc, Direcção Regional de Educação do Centro Coimbra;
Carla Marques MSc, Centro de Desenvolvimento da Criança, Hospital Pediátrico de Coimbra; Teresa S Miguel BSc, Direcção Regional de Educação do Centro, Coimbra;
Ana Margarida Coutinho BSc, Instituto Gulbenkian de Ciência, Oeiras; Luísa Mota-Vieira PhD, Unidade de Genética e Patologia moleculares, Hospital do Divino Espírito Santo, Ponta Delgada, Açores; Esmeralda Gonçalves PhD; Nazaré Mendes Lopes PhD, Faculdade de Ciências e Tecnologia, Universidade de Coimbra; Vitor Rodrigues MD PhD; Henrique Carmona da Mota MD PhD, Faculdade de Medicina, Universidade de Coimbra, Coimbra; Astrid Moura Vicente PhD, Instituto Gulbenkian de Ciência, Oeiras, Portugal.
*Correspondence to first author at Hospital Pediátrico de Coimbra, Av Bissaya Barreto, 3000-076 Coimbra, Portugal. E-mail: [email protected]
Abstract: The objective of this study was to estimate the prevalence of autistic spectrum disorder (ASD) and identify its clinical characterization, and medical conditions in a paediatric population in Portugal. A school survey was conducted in elementary schools, targeting 332 808 school-aged children in the mainland and 10 910 in the Azores islands. Referred children were directly assessed using the Diagnostic and Statistical Manual of Mental Disorders (4th edn), the Autism Diagnostic Interview–Revised, and the Childhood Autism Rating Scale. Clinical history and a laboratory investigation was performed. In parallel, a systematic multi-source search of children known to have autism was carried out in a restricted region. The global prevalence of ASD per 10 000 was 9.2 in mainland, and 15.6 in the Azores, with intriguing regional differences. A diversity of associated medical conditions was documented in 20%, with an unexpectedly high rate of mitochondrial respiratory chain disorders.
31. Thimerosal induces neuronal cell apoptosis by causing cytochrome c and apoptosis-inducing factor release from mitochondria.
International Journal of Molecular Medicine, 2006
Yel L, Brown LE, Su K, Gollapudi S, Gupta S.Department of Medicine, University of California, Irvine, CA 92697, USA. [email protected]
There is a worldwide increasing concern over the neurological risks of thimerosal (ethylmercury thiosalicylate) which is an organic mercury compound that is commonly used as an antimicrobial preservative. In this study, we show that thimerosal, at nanomolar concentrations, induces neuronal cell death through the mitochondrial pathway. Thimerosal, in a concentration- and time-dependent manner, decreased cell viability as assessed by calcein-ethidium staining and caused apoptosis detected by Hoechst 33258 dye. Thimerosal-induced apoptosis was associated with depolarization of mitochondrial membrane, generation of reactive oxygen species, and release of cytochrome c and apoptosis-inducing factor (AIF) from mitochondria to cytosol. Although thimerosal did not affect cellular expression of Bax at the protein level, we observed translocation of Bax from cytosol to mitochondria. Finally, caspase-9 and caspase-3 were activated in the absence of caspase-8 activation. Our data suggest that thimerosal causes apoptosis in neuroblastoma cells by changing the mitochondrial microenvironment.
32. Mitochondrial mediated thimerosal-induced apoptosis in a human neuroblastoma cell line (SK-N-SH).
Humphrey ML, Cole MP, Pendergrass JC, Kiningham KK. Department of Pharmacology, Joan C. Edwards School of Medicine, Marshall University, Huntington, WV 25704-9388, USA.
Environmental exposure to mercurials continues to be a public health issue due to their deleterious effects on immune, renal and neurological function. Recently the safety of thimerosal, an ethyl mercury-containing preservative used in vaccines, has been questioned due to exposure of infants during immunization. Mercurials have been reported to cause apoptosis in cultured neurons; however, the signaling pathways resulting in cell death have not been well characterized. Therefore, the objective of this study was to identify the mode of cell death in an in vitro model of thimerosal-induced neurotoxicity, and more specifically, to elucidate signaling pathways which might serve as pharmacological targets. Within 2 h of thimerosal exposure (5 microM) to the human neuroblastoma cell line, SK-N-SH, morphological changes, including membrane alterations and cell shrinkage, were observed. Cell viability, assessed by measurement of lactate dehydrogenase (LDH) activity in the medium, as well as the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay, showed a time- and concentration-dependent decrease in cell survival upon thimerosal exposure. In cells treated for 24 h with thimerosal, fluorescence microscopy indicated cells undergoing both apoptosis and oncosis/necrosis. To identify the apoptotic pathway associated with thimerosal-mediated cell death, we first evaluated the mitochondrial cascade, as both inorganic and organic mercurials have been reported to accumulate in the organelle. Cytochrome c was shown to leak from the mitochondria, followed by caspase 9 cleavage within 8 h of treatment. In addition, poly(ADP-ribose) polymerase (PARP) was cleaved to form a 85 kDa fragment following maximal caspase 3 activation at 24 h. Taken together these findings suggest deleterious effects on the cytoarchitecture by thimerosal and initiation of mitochondrial-mediated apoptosis.
33. Possible Immunological Disorders in Autism: Concomitant Autoimmunity and Immune Tolerance
The Egyptian Journal of Immunology, 2006
Maha I. Sh. Kawashti, Omnia R. Amin Nadia G. Rowehy
Microbiology Department, Faculty of Medicine (For Girls), Al Azhar University, Cairo, Egypt, Psychiatry Department, Faculty of Medicine, Cairo University, Cairo, Egypt and Serology Lab King Fahad General Hospital, Jeddah, K.S.A.
Abstract: Autism is a pervasive developmental disorder that affect children early in their life. Immunological disorders is one of several contributing factors that have been suggested to cause autism. Thirty autistic children aged 3-6 years and thirty non-autistic psychologically-free siblings were studied. Circulating IgA and IgG autoantibodies to casein and gluten dietary proteins were detected by enzyme-immunoassays (EIA). Circulating IgG antibodies to measles, mumps and rubella vaccine (M.M.R) and cytomeglovirus were investigated by EIA. Results revealed high seropositivity for autoantibodies to casein and gluten: 83.3% and 50% respectively in autistic children as compared to 10% and 6.7% positivity in the control group. Surprisingly, circulating anti-measles, anti-mumps and anti-rubella IgG were positive in only 50%, 73.3% and 53.3% respectively as compared to 100% positivity in the control group. Anti-CMV IgG was positive in 43.3% of the autistic children as compared to 7% in the control group. It is concluded that, autoimmune response to dietary proteins and deficient immune response to measles, mumps and rubella vaccine antigens might be associated with autism, as a leading cause or a resulting event. Further research is needed to confirm these findings.
34. Pediatric Vaccines Influence Primate Behavior, and Amygdala Growth and Opioid Ligand Binding Friday, May 16, 2008: IMFAR
L. Hewitson , Obstetrics, Gynecology and Reproductive Sciences, University of Pittsburgh, Pittsburgh, PA B. Lopresti , Radiology, University of Pittsburgh, Pittsburgh, PA C. Stott , Thoughtful House Center for Children, Austin, TX J. Tomko , Pittsburgh Development Center, University of Pittsburgh, Pittsburgh, PA L. Houser , Pittsburgh Development Center, University of Pittsburgh, Pittsburgh, PA E. Klein , Division of Laboratory Animal Resources, University of Pittsburgh, Pittsburgh, PA C. Castro , Obstetrics, Gynecology and Reproductive Sciences, University of Pittsburgh, Pittsburgh, PA G. Sackett , Psychology, Washington National Primate Research Center, Seattle, WA S. Gupta , Medicine, Pathology & Laboratory Medicine, University of California – Irvine, Irvine, CA D. Atwood , Chemistry, University of Kentucky, Lexington, KY L. Blue , Chemistry, University of Kentucky, Lexington, KY E. R. White , Chemistry, University of Kentucky, Lexington, KY A. Wakefield , Thoughtful House Center for Children, Austin, TX
Background: Macaques are commonly used in pre-clinical vaccine safety testing, but the combined childhood vaccine regimen, rather than individual vaccines, has not been studied. Childhood vaccines are a possible causal factor in autism, and abnormal behaviors and anomalous amygdala growth are potentially inter-related features of this condition.
Objectives: The objective of this study was to compare early infant cognition and behavior with amygdala size and opioid binding in rhesus macaques receiving the recommended childhood vaccines (1994-1999), the majority of which contained the bactericidal preservative ethylmercurithiosalicylic acid (thimerosal).
Methods: Macaques were administered the recommended infant vaccines, adjusted for age and thimerosal dose (exposed; N=13), or saline (unexposed; N=3). Primate development, cognition and social behavior were assessed for both vaccinated and unvaccinated infants using standardized tests developed at the Washington National Primate Research Center. Amygdala growth and binding were measured serially by MRI and by the binding of the non-selective opioid antagonist [11C]diprenorphine, measured by PET, respectively, before (T1) and after (T2) the administration of the measles-mumps-rubella vaccine (MMR).
Results: Compared with unexposed animals, significant neurodevelopmental deficits were evident for exposed animals in survival reflexes, tests of color discrimination and reversal, and learning sets. Differences in behaviors were observed between exposed and unexposed animals and within the exposed group before and after MMR vaccination. Compared with unexposed animals, exposed animals showed attenuation of amygdala growth and differences in the amygdala binding of [11C]diprenorphine. Interaction models identified significant associations between specific aberrant social and non-social behaviors, isotope binding, and vaccine exposure.
Conclusions: This animal model, which examines for the first time, behavioral, functional, and neuromorphometric consequences of the childhood vaccine regimen, mimics certain neurological abnormalities of autism. The findings raise important safety issues while providing a potential model for examining aspects of causation and disease pathogenesis in acquired disorders of behavior and development.
35. Thimerosal exposure in infants and neurodevelopmental disorders: An assessment of computerized medical records in the Vaccine Safety Datalink.
Young HA, Geier DA, Geier MR.
The George Washington University School of Public Health and Health Services, Department of Epidemiology and Biostatistics, United States.
The study evaluated possible associations between neurodevelopmental disorders (NDs) and exposure to mercury (Hg) from Thimerosal-containing vaccines (TCVs) by examining the automated Vaccine Safety Datalink (VSD). A total of 278,624 subjects were identified in birth cohorts from 1990-1996 that had received their first oral polio vaccination by 3 months of age in the VSD. The birth cohort prevalence rate of medically diagnosed International Classification of Disease, 9th revision (ICD-9) specific NDs and control outcomes were calculated. Exposures to Hg from TCVs were calculated by birth cohort for specific exposure windows from birth-7 months and birth-13 months of age. Poisson regression analysis was used to model the association between the prevalence of outcomes and Hg doses from TCVs. Consistent significantly increased rate ratios were observed for autism, autism spectrum disorders, tics, attention deficit disorder, and emotional disturbances with Hg exposure from TCVs. By contrast, none of the control outcomes had significantly increased rate ratios with Hg exposure from TCVs. Routine childhood vaccination should be continued to help reduce the morbidity and mortality associated with infectious diseases, but efforts should be undertaken to remove Hg from vaccines. Additional studies should be conducted to further evaluate the relationship between Hg exposure and NDs.
36. Glutathione, oxidative stress and neurodegeneration
Schulz JB, Lindenau J, Seyfried J, Dichgans J.
Neurodegeneration Laboratory, Department of Neurology, University of Tübingen, Germany.
Eur J Biochem. 2000 Aug;267(16):4904-11.
There is significant evidence that the pathogenesis of several neurodegenerative diseases, including Parkinson’s disease, Alzheimer’s disease, Friedreich’s ataxia and amyotrophic lateral sclerosis, may involve the generation of reactive oxygen species and mitochondrial dysfunction. Here, we review the evidence for a disturbance of glutathione homeostasis that may either lead to or result from oxidative stress in neurodegenerative disorders. Glutathione is an important intracellular antioxidant that protects against a variety of different antioxidant species. An important role for glutathione was proposed for the pathogenesis of Parkinson’s disease, because a decrease in total glutathione concentrations in the substantia nigra has been observed in preclinical stages, at a time at which other biochemical changes are not yet detectable. Because glutathione does not cross the blood-brain barrier other treatment options to increase brain concentrations of glutathione including glutathione analogs, mimetics or precursors are discussed.
37. Hepatitis B triple series vaccine and developmental disability in US children aged 1-9 years
Carolyn Gallagher a; Melody Goodman, Graduate Program in Public Health, Stony Brook University Medical Center, Health Sciences Center, New York, USA
Journal Toxicological & Environmental Chemistry, Volume 90, Issue 5 September 2008 , pages 997 – 1008
This study investigated the association between vaccination with the Hepatitis B triple series vaccine prior to 2000 and developmental disability in children aged 1-9 years (n = 1824), proxied by parental report that their child receives early intervention or special education services (EIS). National Health and Nutrition Examination Survey 1999-2000 data were analyzed and adjusted for survey design by Taylor Linearization using SAS version 9.1 software, with SAS callable SUDAAN version 9.0.1. The odds of receiving EIS were approximately nine times as great for vaccinated boys (n = 46) as for unvaccinated boys (n = 7), after adjustment for confounders. This study found statistically significant evidence to suggest that boys in United States who were vaccinated with the triple series Hepatitis B vaccine, during the time period in which vaccines were manufactured with thimerosal, were more susceptible to developmental disability than were unvaccinated boys.
38. Induction of metallothionein in mouse cerebellum and cerebrum with low-dose thimerosal injection.
Minami T, Miyata E, Sakamoto Y, Yamazaki H, Ichida S., Department of Life Sciences, School of Science & Engineering, Kinki University, 3-4-1 Kowakae, Higashi-osaka, Osaka, 577-8502, Japan, [email protected]
Cell Biology and Toxicology. 2009 Apr 9. [Epub ahead of print]
Thimerosal, an ethyl mercury compound, is used worldwide as a vaccine preservative. We previously observed that the mercury concentration in mouse brains did not increase with the clinical dose of thimerosal injection, but the concentration increased in the brain after the injection of thimerosal with lipopolysaccharide, even if a low dose of thimerosal was administered. Thimerosal may penetrate the brain, but is undetectable when a clinical dose of thimerosal is injected; therefore, the induction of metallothionein (MT) messenger RNA (mRNA) and protein was observed in the cerebellum and cerebrum of mice after thimerosal injection, as MT is an inducible protein. MT-1 mRNA was expressed at 6 and 9 h in both the cerebrum and cerebellum, but MT-1 mRNA expression in the cerebellum was three times higher than that in the cerebrum after the injection of 12 microg/kg thimerosal. MT-2 mRNA was not expressed until 24 h in both organs. MT-3 mRNA was expressed in the cerebellum from 6 to 15 h after the injection, but not in the cerebrum until 24 h. MT-1 and MT-3 mRNAs were expressed in the cerebellum in a dose-dependent manner. Furthermore, MT-1 protein was detected from 6 to 72 h in the cerebellum after 12 microg/kg of thimerosal was injected and peaked at 10 h. MT-2 was detected in the cerebellum only at 10 h. In the cerebrum, little MT-1 protein was detected at 10 and 24 h, and there were no peaks of MT-2 protein in the cerebrum. In conclusion, MT-1 and MT-3 mRNAs but not MT-2 mRNA are easily expressed in the cerebellum rather than in the cerebrum by the injection of low-dose thimerosal. It is thought that the cerebellum is a sensitive organ against thimerosal. As a result of the present findings, in combination with the brain pathology observed in patients diagnosed with autism, the present study helps to support the possible biological plausibility for how low-dose exposure to mercury from thimerosal-containing vaccines may be associated with autism.
39. Mercury induces inflammatory mediator release from human mast cells
Duraisamy Kempuraj, Shahrzad Asadi, Bodi Zhang, Akrivi Manola, Jennifer Hogan,
Erika Peterson, Theoharis C Theoharides
Journal of Neuroinflammation 2010, 7:20 doi:10.1186/1742-2094-7-20
Background: Mercury is known to be neurotoxic, but its effects on the immune system are less well known. Mast cells are involved in allergic reactions, but also in innate and acquired immunity, as well as in inflammation. Many patients with Autism Spectrum Disorders (ASD) have “allergic” symptoms; moreover, the prevalence of ASD in patients with mastocytosis, characterized by numerous hyperactive mast cells in most tissues, is 10-fold higher than the general population suggesting mast cell involvement. We, therefore, investigated the effect of mercuric chloride (HgCl2) on human mast cell activation.
Methods: Human leukemic cultured LAD2 mast cells and normal human umbilical cord bloodderived cultured mast cells (hCBMCs) were stimulated by HgCl2 (0.1-10 μM) for either 10 min for beta-hexosaminidase release or 24 hr for measuring vascular endothelial growth factor (VEGF) and IL-6 release by ELISA.
Results: HgCl2 induced a 2-fold increase in β-hexosaminidase release, and also significant VEGF release at 0.1 and 1 μM (311±32 pg/106 cells and 443±143 pg/106 cells, respectively) from LAD2 mast cells compared to control cells (227±17 pg/106 cells, n=5, p<0.05). Addition of HgCl2 (0.1 μM) to the proinflammatory neuropeptide substance P (SP, 0.1 μM) had synergestic action in inducing VEGF from LAD2 mast cells. HgCl2 also stimulated significant VEGF release (360 ± 100 pg/106 cells at 1 μM, n=5, p<0.05) from hCBMCs compared to control cells (182 ±57 pg/106 cells), and IL-6 release (466±57 pg/106 cells at 0.1 μM) compared to untreated cells (13±25 pg/106 cells, n=5, p<0.05). Addition of HgCl2 (0.1 μM) to SP (5 μM) further increased IL-6 release. Conclusions: HgCl2 stimulates VEGF and IL-6 release from human mast cells. This phenomenon could disrupt the blood-brain-barrier and permit brain inflammation. As a result, the findings of the present study provide a biological mechanism for how low levels of mercury may contribute to ASD pathogenesis.
40. Influence of pediatric vaccines on amygdala growth and opioid ligand binding in rhesus macaque infants: A pilot study
Acta Neurobiol Exp 2010, 70: 147–164 Polish Neuroscience Society – PTBUN, Nencki Institute of Experimental Biology
Laura Hewitson1,2,*, Brian J. Lopresti3, Carol Stott4, N. Scott Mason3 and Jaime Tomko1
Department of Obstetrics and Gynecology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA; Thoughtful House Center for Children, Austin, TX, USA; Department of Radiology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA; 4Independent Chartered Scientist, Cambridge, UK;
This longitudinal, case-control pilot study examined amygdala growth in rhesus macaque infants receiving the complete US childhood vaccine schedule (1994-1999). Longitudinal structural and functional neuroimaging was undertaken to examine central effects of the vaccine regimen on the developing brain. Vaccine-exposed and saline-injected control infants underwent MRI and PET imaging at approximately 4 and 6 months of age, representing two specific timeframes within the vaccination schedule. Volumetric analyses showed that exposed animals did not undergo the maturational changes over time in amygdala volume that was observed in unexposed animals. After controlling for left amygdala volume, the binding of the opioid antagonist [11C]diprenorphine (DPN) in exposed animals remained relatively constant over time, compared with unexposed animals, in which a significant decrease in [11C]DPN binding occurred. These results suggest that maturational changes in amygdala volume and the binding capacity of [11C]DPN in the amygdala was significantly altered in infant macaques receiving the vaccine schedule. The macaque infant is a relevant animal model in which to investigate specific environmental exposures and structural/functional neuroimaging during neurodevelopment.
41. Cultured lymphocytes from autistic children and non-autistic siblings up-regulate heat shock protein RNA in response to thimerosal challenge.
Neurotoxicology. 2006 Sep;27(5):685-92. Epub 2006 Jun 16.
Walker SJ, Segal J, Aschner M.
Department of Physiology and Pharmacology, Wake Forest University School of Medicine, Winston-Salem, NC 27156, USA. [email protected]
There are reports suggesting that some autistic children are unable to mount an adequate response following exposure to environmental toxins. This potential deficit, coupled with the similarity in clinical presentations of autism and some heavy metal toxicities, has led to the suggestion that heavy metal poisoning might play a role in the etiology of autism in uniquely susceptible individuals. Thimerosal, an anti-microbial preservative previously added routinely to childhood multi-dose vaccines, is composed of 49.6% ethyl mercury. Based on the levels of this toxin that children receive through routine immunization schedules in the first years of life, it has been postulated that thimerosal may be a potential triggering mechanism contributing to autism in susceptible individuals. One potential risk factor in these individuals may be an inability to adequately up-regulate metallothionein (MT) biosynthesis in response to presentation of a heavy metal challenge. To investigate this hypothesis, cultured lymphocytes (obtained from the Autism Genetic Resource Exchange, AGRE) from autistic children and non-autistic siblings were challenged with either 10 microM ethyl mercury, 150 microM zinc, or fresh media (control). Following the challenge, total RNA was extracted and used to query “whole genome” DNA microarrays. Cultured lymphocytes challenged with zinc responded with an impressive up-regulation of MT transcripts (at least nine different MTs were over-expressed) while cells challenged with thimerosal responded by up-regulating numerous heat shock protein transcripts, but not MTs. Although there were no apparent differences between autistic and non-autistic sibling responses in this very small sampling group, the differences in expression profiles between those cells treated with zinc versus thimerosal were dramatic. Determining cellular response, at the level of gene expression, has important implications for the understanding and treatment of conditions that result from exposure to neurotoxic compounds.
42. Hepatitis B Vaccination of Male Neonates and Autism
Annals of Epidemiology , Vol. 19, No. 9 ABSTRACTS (ACE), September 2009: 651-680,
CM Gallagher, MS Goodman, Graduate Program in Public
Health, Stony Brook University Medical Center, Stony Brook, NY
PURPOSE: Universal newborn immunization with hepatitis B vaccine was recommended in 1991; however, safety findings are mixed. The Vaccine Safety Datalink Workgroup reported no association between hepatitis B vaccination at birth and febrile episodes or neurological adverse events. Other studies found positive associations between
hepatitis B vaccination and ear infection, pharyngitis, and chronic arthritis; as well as receipt of early intervention/special education services (EIS); in probability samples of U.S. children. Children with autistic spectrum disorder (ASD) comprise a growing caseload for EIS. We evaluated the association between hepatitis B vaccination of male neonates and parental report of ASD.
METHODS: This cross-sectional study used U.S. probability samples obtained from National Health Interview Survey 1997-2002 datasets. Logistic regression modeling was used to estimate the effect of neonatal hepatitis B vaccination on ASD risk amongboys age 3-17 years with shot records, adjusted for race, maternal education, and two-parent household.
RESULTS:Boyswho received the hepatitis B vaccine during the first month of life had 2.94 greater odds for ASD (nZ31 of 7,486; OR Z 2.94; p Z 0.03; 95% CI Z 1.10, 7.90)
compared to later- or unvaccinated boys.Non-Hispanicwhite boys were 61%less likely to haveASD(ORZ0.39; pZ0.04; 95% CIZ0.16, 0.94) relative to non-white boys.
CONCLUSION: Findings suggest that U.S. male neonates vaccinated with hepatitis B vaccine had a 3-fold greater risk of ASD; risk was greatest for non-white boys.
43. Neonatal administration of a vaccine preservative, thimerosal, produces lasting impairment of nociception and apparent activation of opioid system in rats.
Brain Res. 2009 Dec 8;1301:143-51. Epub 2009 Sep 9.
Olczak M, Duszczyk M, Mierzejewski P, Majewska MD. Department of Pharmacology and Physiology of the Nervous System, Institute of Psychiatry and Neurology, Warsaw, Poland.
Thimerosal (THIM), an organomercury preservative added to many child vaccines is a suspected factor in pathogenesis of neurodevelopmental disorders. We examined the pharmacokinetics of Hg in the brain, liver and kidneys after i.m. THIM injection in suckling rats and we tested THIM effect on nociception. THIM solutions were injected to Wistar and Lewis rats in a vaccination-like mode on PN days 7, 9, 11 and 15 in four equal doses. For Wistar rats these were: 12, 48, 240, 720, 1440, 2160, 3000 microg Hg/kg and for Lewis: 54, 216, 540 and 1080 microg Hg/kg. Pharmacokinetic analysis revealed that Hg from THIM injections accumulates in the rat brain in significant amounts and remains there longer than 30 days after the injection. At the 6th week of age animals were examined for pain sensitivity using the hot plate test. THIM treated rats of both strains and sexes manifested statistically significantly elevated pain threshold (latency for paw licking, jumping) on a hot plate (56 degrees C). Wistar rats were more sensitive to this effect than Lewis rats. Protracted THIM-induced hypoalgesia was reversed by naloxone (5 mg/kg, i.p.) injected before the hot plate test, indicative of involvement of endogenous opioids. This was confirmed by augmented catalepsy after morphine (2.5 mg/kg, s.c.) injection. Acute THIM injection to 6-week-old rats also produced hypoalgesia, but this effect was transient and was gone within 14 days. Present findings show that THIM administration to suckling or adult rats impairs sensitivity to pain, apparently due to activation the endogenous opioid system.
44. Sorting out the spinning of autism: heavy metals and the question of incidence
Acta Neurobiol Exp 2010, 70: 165–176
Mary Catherine DeSoto* and Robert T. Hitlan, Department of Psychology, University of Northern Iowa, Cedar Falls, Iowa, USA
The reasons for the rise in autism prevalence are a subject of heated professional debate. Featuring a critical appraisal of some research used to question whether there is a rise in cases and if rising levels of autism are related to environmental
exposure to toxins (Soden et al. 2007, Thompson et al. 2007, Barbaresi et al. 2009) we aim to evaluate the actual state of scientific knowledge. In addition, we surveyed the empirical research on the topic of autism and heavy metal toxins. Overall, the various causes that have led to the increase in autism diagnosis are likely multi-faceted, and understanding the causes is one of the most important health topics today. We argue that scientific research does not support rejecting the link between the neurodevelopmental disorder of autism and toxic exposures.
45. Urinary Porphyrin Excretion in Neurotypical and Autistic Children
Environ Health Perspect. 2010 Oct;118(10):1450-7. Epub 2010 Jun 24.
Woods JS, Armel SE, Fulton DI, Allen J, Wessels K, Simmonds PL, Granpeesheh D, Mumper E, Bradstreet JJ, Echeverria D, Heyer NJ, Rooney JP., Department of Environmental and Occupational Health Sciences, University of Washington
BACKGROUND: Increased urinary concentrations of pentacarboxyl-, precopro- and copro-porphyrins have been associated with prolonged mercury (Hg) exposure in adults, and comparable increases have been attributed to Hg exposure in children with autism (AU).
OBJECTIVES: This study was designed to measure and compare urinary porphyrin concentrations in neurotypical (NT) children and same-age children with autism, and to examine the association between porphyrin levels and past or current Hg exposure in children with autism.
METHODS: This exploratory study enrolled 278 children 2-12 years of age. We evaluated three groups: AU, pervasive developmental disorder-not otherwise specified (PDD-NOS), and NT. Mothers/caregivers provided information at enrollment regarding medical, dental, and dietary exposures. Urine samples from all children were acquired for analyses of porphyrin, creatinine, and Hg. Differences between groups for mean porphyrin and Hg levels were evaluated. Logistic regression analysis was conducted to determine whether porphyrin levels were associated with increased risk of autism.
RESULTS: Mean urinary porphyrin concentrations are naturally high in young children and decline by as much as 2.5-fold between 2 and 12 years of age. Elevated copro- (p < 0.009), hexacarboxyl- (p < 0.01) and pentacarboxyl- (p < 0.001) porphyrin concentrations were significantly associated with AU but not with PDD-NOS. No differences were found between NT and AU in urinary Hg levels or in past Hg exposure as determined by fish consumption, number of dental amalgam fillings, or vaccines received. CONCLUSIONS:These findings identify disordered porphyrin metabolism as a salient characteristic of autism. Hg exposures were comparable between diagnostic groups, and a porphyrin pattern consistent with that seen in Hg-exposed adults was not apparent.
46. Mitochondrial dysfunction in autism spectrum disorders: a systematic review and meta-analysis
Molecular Psychiatry advance online publication 25 January 2011;doi: 10.1038/mp.2010.136
D A Rossignol and R E Frye
A comprehensive literature search was performed to collate evidence of mitochondrial dysfunction in autism spectrum disorders (ASDs) with two primary objectives. First, features of mitochondrial dysfunction in the general population of children with ASD were identified. Second, characteristics of mitochondrial dysfunction in children with ASD and concomitant mitochondrial disease (MD) were compared with published literature of two general populations: ASD children without MD, and non-ASD children with MD. The prevalence of MD in the general population of ASD was 5.0% (95% confidence interval 3.2, 6.9%), much higher than found in the general population (~0.01%). The prevalence of abnormal biomarker values of mitochondrial dysfunction was high in ASD, much higher than the prevalence of MD. Variances and mean values of many mitochondrial biomarkers (lactate, pyruvate, carnitine and ubiquinone) were significantly different between ASD and controls. Some markers correlated with ASD severity. Neuroimaging, in vitro and post-mortem brain studies were consistent with an elevated prevalence of mitochondrial dysfunction in ASD. Taken together, these findings suggest children with ASD have a spectrum of mitochondrial dysfunction of differing severity. Eighteen publications representing a total of 112 children with ASD and MD (ASD/MD) were identified. The prevalence of developmental regression (52%), seizures (41%), motor delay (51%), gastrointestinal abnormalities (74%), female gender (39%), and elevated lactate (78%) and pyruvate (45%) was significantly higher in ASD/MD compared with the general ASD population. The prevalence of many of these abnormalities was similar to the general population of children with MD, suggesting that ASD/MD represents a distinct subgroup of children with MD. Most ASD/MD cases (79%) were not associated with genetic abnormalities, raising the possibility of secondary mitochondrial dysfunction. Treatment studies for ASD/MD were limited, although improvements were noted in some studies with carnitine, co-enzyme Q10 and B-vitamins. Many studies suffered from limitations, including small sample sizes, referral or publication biases, and variability in protocols for selecting children for MD workup, collecting mitochondrial biomarkers and defining MD. Overall, this evidence supports the notion that mitochondrial dysfunction is associated with ASD. Additional studies are needed to further define the role of mitochondrial dysfunction in ASD.
47. Sensitization effect of thimerosal is mediated in vitro via reactive oxygen species and calcium signaling.
Toxicology. 2010 July – August;274(1-3):1-9. Epub 2010 May 10.
Migdal C, Foggia L, Tailhardat M, Courtellemont P, Haftek M, Serres M.
Thimerosal, a mercury derivative composed of ethyl mercury chloride (EtHgCl) and thiosalicylic acid (TSA), is widely used as a preservative in vaccines and cosmetic products and causes cutaneous reactions. Since dendritic cells (DCs) play an essential role in the immune response, the sensitization potency of chemicals was studied in vitro using U937, a human promyelomonocytic cell line that is used as a surrogate of monocytic differentiation and activation. Currently, this cell line is under ECVAM (European Center for the Validation of Alternative Methods) validation as an alternative method for discriminating chemicals. Thimerosal and mercury derivatives induced in U937 an overexpression of CD86 and interleukin (IL)-8 secretion similarly to 1-chloro-2,4-dinitrobenzene (DNCB), a sensitizer used as a positive control for DC activation. Non-sensitizers, dichloronitrobenzene (DCNB), TSA and sodium dodecyl sulfate (SDS), an irritant, had no effect. U937 activation was prevented by cell pretreatment with N-acetyl-l-cysteine (NAC) but not with thiol-independent antioxidants except vitamin E which affected CD86 expression by preventing lipid peroxidation of cell membranes. Thimerosal, EtHgCl and DNCB induced glutathione (GSH) depletion and reactive oxygen species (ROS) within 15min; another peak was detected after 2h for mercury compounds only. MitoSOX, a specific mitochondrial fluorescent probe, confirmed that ROS were essentially produced by mitochondria in correlation with its membrane depolarization. Changes in mitochondrial membrane permeability induced by mercury were reversed by NAC but not by thiol-independent antioxidants. Thimerosal and EtHgCl also induced a calcium (Ca(2+)) influx with a peak at 3h, suggesting that Ca(2+) influx is a secondary event following ROS induction as Ca(2+) influx was suppressed after pretreatment with NAC but not with thiol-independent antioxidants. Ca(2+) influx was also suppressed when culture medium was deprived of Ca(2+) confirming the specificity of the measure. In conclusion, these data suggest that thimerosal induced U937 activation via oxidative stress from mitochondrial stores and mitochondrial membrane depolarization with a primordial effect of thiol groups. A cross-talk between ROS and Ca(2+) influx was demonstrated.
50. Theoretical aspects of autism: Causes—A review
Journal of Immunotoxicology, January-March 2011, Vol. 8, No. 1 , Pages 68-79
Helen V. Ratajczak, PhD
Autism, a member of the pervasive developmental disorders (PDDs), has been increasing dramatically since its description by Leo Kanner in 1943. First estimated to occur in 4 to 5 per 10,000 children, the incidence of autism is now 1 per 110 in the United States, and 1 per 64 in the United Kingdom, with similar incidences throughout the world. Searching information from 1943 to the present in PubMed and Ovid Medline databases, this review summarizes results that correlate the timing of changes in incidence with environmental changes. Autism could result from more than one cause, with different manifestations in different individuals that share common symptoms. Documented causes of autism include genetic mutations and/or deletions, viral infections, and encephalitis following vaccination. Therefore, autism is the result of genetic defects and/or inflammation of the brain. The inflammation could be caused by a defective placenta, immature blood-brain barrier, the immune response of the mother to infection while pregnant, a premature birth, encephalitis in the child after birth, or a toxic environment.
51. A Positive Association found between Autism Prevalence and Childhood Vaccination uptake across the U.S. Population
Journal of Toxicology and Environmental Health, Part A: Current Issues
Volume 74, Issue 14, 2011, Pages 903 – 916
Author: Gayle DeLonga
The reason for the rapid rise of autism in the United States that began in the 1990s is a mystery. Although individuals probably have a genetic predisposition to develop autism, researchers suspect that one or more environmental triggers are also needed. One of those triggers might be the battery of vaccinations that young children receive. Using regression analysis and controlling for family income and ethnicity, the relationship between the proportion of children who received the recommended vaccines by age 2 years and the prevalence of autism (AUT) or speech or language impairment (SLI) in each U.S. state from 2001 and 2007 was determined. A positive and statistically significant relationship was found: The higher the proportion of children receiving recommended vaccinations, the higher was the prevalence of AUT or SLI. A 1% increase in vaccination was associated with an additional 680 children having AUT or SLI. Neither parental behavior nor access to care affected the results, since vaccination proportions were not significantly related (statistically) to any other disability or to the number of pediatricians in a U.S. state. The results suggest that although mercury has been removed from many vaccines, other culprits may link vaccines to autism. Further study into the relationship between vaccines and autism is warranted. To read the abstract click HERE.
52. Ancestry of pink disease (infantile acrodynia) identified as a risk factor for autism spectrum disorders.
J Toxicol Environ Health A. 2011 Sep 15;74(18):1185-94.
Shandley K, Austin DW.
Swinburne Autism Bio-Research Initiative (SABRI), Brain and Psychological Sciences Research Centre , Swinburne University of Technology , Hawthorn , Victoria , Australia.
Pink disease (infantile acrodynia) was especially prevalent in the first half of the 20th century. Primarily attributed to exposure to mercury (Hg) commonly found in teething powders, the condition was developed by approximately 1 in 500 exposed children. The differential risk factor was identified as an idiosyncratic sensitivity to Hg. Autismspectrum disorders (ASD) have also been postulated to be produced by Hg. Analogous to the pink disease experience, Hg exposure is widespread yet only a fraction of exposed children develop an ASD, suggesting sensitivity to Hg may also be present in children with an ASD. The objective of this study was to test the hypothesis that individuals with a known hypersensitivity to Hg (pink disease survivors) may be more likely to have descendants with an ASD. Five hundred and twenty-two participants who had previously been diagnosed with pink disease completed a survey on the health outcomes of their descendants. The prevalence rates of ASD and a variety of other clinical conditions diagnosed in childhood (attention deficit hyperactivity disorder, epilepsy, Fragile X syndrome, and Down syndrome) were compared to well-established general population prevalence rates. The results showed the prevalence rate of ASD among the grandchildren of pink disease survivors (1 in 22) to be significantly higher than the comparable general population prevalence rate (1 in 160). The results support the hypothesis that Hg sensitivity may be a heritable/genetic risk factor for ASD.
53. Do aluminum vaccine adjuvants contribute to the rising prevalence of autism?
J Inorg Biochem. 2011 Nov;105(11):1489-99. Epub 2011 Aug 23.
Tomljenovic L, Shaw CA.
Neural Dynamics Research Group, Department of Ophthalmology and Visual Sciences, University of British Columbia, 828 W. 10th Ave, Vancouver, BC, Canada V5Z 1L8.
Autism spectrum disorders (ASD) are serious multisystem developmental disorders and an urgent global public health concern. Dysfunctional immunity and impaired brain function are core deficits in ASD. Aluminum (Al), the most commonly used vaccine adjuvant, is a demonstrated neurotoxin and a strong immune stimulator. Hence, adjuvant Al has the potential to induce neuroimmune disorders. When assessing adjuvant toxicity in children, two key points ought to be considered: (i) children should not be viewed as “small adults” as their unique physiology makes them much more vulnerable to toxic insults; and (ii) if exposure to Al from only few vaccines can lead to cognitive impairment and autoimmunity in adults, is it unreasonable to question whether the current pediatric schedules, often containing 18 Al adjuvanted vaccines, are safe for children? By applying Hill’s criteria for establishing causality between exposure and outcome we investigated whether exposure to Al from vaccines could be contributing to the rise in ASD prevalence in the Western world. Our results show that: (i) children from countries with the highest ASD prevalence appear to have the highest exposure to Al from vaccines; (ii) the increase in exposure to Al adjuvants significantly correlates with the increase in ASD prevalence in the United States observed over the last two decades (Pearson r=0.92, p<0.0001); and (iii) a significant correlation exists between the amounts of Al administered to preschool children and the current prevalence of ASD in seven Western countries, particularly at 3-4months of age (Pearson r=0.89-0.94, p=0.0018-0.0248). The application of the Hill’s criteria to these data indicates that the correlation between Al in vaccines and ASD may be causal. Because children represent a fraction of the population most at risk for complications following exposure to Al, a more rigorous evaluation of Al adjuvant safety seems warranted.
54. Lasting neuropathological changes in rat brain after intermittent neonatal administration of thimerosal.
Folia Neuropathol. 2010;48(4):258-69. Olczak M, Duszczyk M, Mierzejewski P, Wierzba-Bobrowicz T, Majewska MD.
Department of Pharmacology and Physiology of the Nervous System, Institute of Psychiatry and Neurology, ul. Sobieskiego 9, Warsaw, Poland.
Thimerosal, an organomercurial added as a preservative to some vaccines, is a suspected iatrogenic factor, possibly contributing to paediatric neurodevelopmental disorders including autism. We examined the effects of early postnatal administration of thimerosal (four i.m. injections, 12 or 240 μg THIM-Hg/kg, on postnatal days 7, 9, 11 and 15) on brain pathology in Wistar rats. Numerous neuropathological changes were observed in young adult rats which were treated postnatally with thimerosal. They included: ischaemic degeneration of neurons and “dark” neurons in the prefrontal and temporal cortex, the hippocampus and the cerebellum, pathological changes of the blood vessels in the temporal cortex, diminished synaptophysin reaction in the hippocampus, atrophy of astroglia in the hippocampus and cerebellum, and positive caspase-3 reaction in Bergmann astroglia. These findings document neurotoxic effects of thimerosal, at doses equivalent to those used in infant vaccines or higher, in developing rat brain, suggesting likely involvement of this mercurial in neurodevelopmental disorders.
55. Persistent behavioral impairments and alterations of brain dopamine system after early postnatal administration of thimerosal in rats.
Behav Brain Res. 2011 Sep 30;223(1):107-18. doi: 10.1016/j.bbr.2011.04.026. Epub 2011 Apr 28.
Olczak M, Duszczyk M, Mierzejewski P, Meyza K, Majewska MD. Department of Pharmacology and Physiology of the Nervous System, Institute of Psychiatry and Neurology, 02-957 Warsaw, Poland.
The neurotoxic organomercurial thimerosal (THIM), used for decades as vaccine preservative, is a suspected factor in the pathogenesis of some neurodevelopmental disorders. Previously we showed that neonatal administration of THIM at doses equivalent to those used in infant vaccines or higher, causes lasting alterations in the brain opioid system in rats. Here we investigated neonatal treatment with THIM (at doses 12, 240, 1440 and 3000 μg Hg/kg) on behaviors, which are characteristically altered in autism, such as locomotor activity, anxiety, social interactions, spatial learning, and on the brain dopaminergic system in Wistar rats of both sexes. Adult male and female rats, which were exposed to the entire range of THIM doses during the early postnatal life, manifested impairments of locomotor activity and increased anxiety/neophobia in the open field test. In animals of both sexes treated with the highest THIM dose, the frequency of prosocial interactions was reduced, while the frequency of asocial/antisocial interactions was increased in males, but decreased in females. Neonatal THIM treatment did not significantly affect spatial learning and memory. THIM-exposed rats also manifested reduced haloperidol-induced catalepsy, accompanied by a marked decline in the density of striatal D₂ receptors, measured by immunohistochemical staining, suggesting alterations to the brain dopaminergic system. Males were more sensitive than females to some neurodisruptive/neurotoxic actions of THIM. These data document that early postnatal THIM administration causes lasting neurobehavioral impairments and neurochemical alterations in the brain, dependent on dose and sex. If similar changes occur in THIM/mercurial-exposed children, they could contribute do neurodevelopmental disorders.
56. Risk Factors for Autistic Regression: Results of an Ambispective Cohort Study.
J Child Neurol. 2012 Jan 30. [Epub ahead of print]
Zhang Y, Xu Q, Liu J, Li SC, Xu X., Department of Child Health Care, Children’s Hospital of Fudan University,Shanghai, China.
A subgroup of children diagnosed with autism experience developmental regression featured by a loss of previously acquired abilities. The pathogeny of autistic regression is unknown, although many risk factors likely exist. To better characterize autistic regression and investigate the association between autistic regression and potential influencing factors in Chinese autistic children, we conducted an ambispective study with a cohort of 170 autistic subjects. Analyses by multiple logistic regression showed significant correlations between autistic regression and febrile seizures (OR = 3.53, 95% CI = 1.17-10.65, P = .025), as well as with a family history of neuropsychiatric disorders (OR = 3.62, 95% CI = 1.35-9.71, P = .011). This study suggests that febrile seizures and family history of neuropsychiatric disorders are correlated with autistic regression.
57. Adverse events following 12 and 18 month vaccinations: a population-based, self-controlled case series analysis.
PLoS One. 2011;6(12):e27897. Epub 2011 Dec 12.
Wilson K, Hawken S, Kwong JC, Deeks S, Crowcroft NS, Van Walraven C, Potter BK, Chakraborty P, Keelan J, Pluscauskas M, Manuel D. Department of Medicine, Ottawa Hospital Research Institute, University of Ottawa, Ottawa, Canada. [email protected]
Live vaccines have distinct safety profiles, potentially causing systemic reactions one to 2 weeks after administration. In the province of Ontario, Canada, live MMR vaccine is currently recommended at age 12 months and 18 months.
Using the self-controlled case series design we examined 271,495 12 month vaccinations and 184,312 18 month vaccinations to examine the relative incidence of the composite endpoint of emergency room visits or hospital admissions in consecutive one day intervals following vaccination. These were compared to a control period 20 to 28 days later. In a post-hoc analysis we examined the reasons for emergency room visits and the average acuity score at presentation for children during the at-risk period following the 12 month vaccine.
Four to 12 days post 12 month vaccination, children had a 1.33 (1.29-1.38) increased relative incidence of the combined endpoint compared to the control period, or at least one event during the risk interval for every 168 children vaccinated. Ten to 12 days post 18 month vaccination, the relative incidence was 1.25 (95%, 1.17-1.33) which represented at least one excess event for every 730 children vaccinated. The primary reason for increased events was statistically significant elevations in emergency room visits following all vaccinations. There were non-significant increases in hospital admissions. There were an additional 20 febrile seizures for every 100,000 vaccinated at 12 months.
There are significantly elevated risks of primarily emergency room visits approximately one to two weeks following 12 and 18 month vaccination. Future studies should examine whether these events could be predicted or prevented.
58. Administration of thimerosal to infant rats increases overflow of glutamate and aspartate in the prefrontal cortex: protective role of dehydroepiandrosterone sulfate.
Neurochem Res. 2012 Feb;37(2):436-47. Epub 2011 Oct 21.
Duszczyk-Budhathoki M, Olczak M, Lehner M, Majewska MD. Marie Curie Chairs Program, Department of Pharmacology and Physiology of Nervous System, Institute of Psychiatry and Neurology, 02-957, Warsaw, Poland.
Thimerosal, a mercury-containing vaccine preservative, is a suspected factor in the etiology of neurodevelopmental disorders. We previously showed that its administration to infant rats causes behavioral, neurochemical and neuropathological abnormalities similar to those present in autism. Here we examined, using microdialysis, the effect of thimerosal on extracellular levels of neuroactive amino acids in the rat prefrontal cortex (PFC). Thimerosal administration (4 injections, i.m., 240 μg Hg/kg on postnatal days 7, 9, 11, 15) induced lasting changes in amino acid overflow: an increase of glutamate and aspartate accompanied by a decrease of glycine and alanine; measured 10-14 weeks after the injections. Four injections of thimerosal at a dose of 12.5 μg Hg/kg did not alter glutamate and aspartate concentrations at microdialysis time (but based on thimerosal pharmacokinetics, could have been effective soon after its injection). Application of thimerosal to the PFC in perfusion fluid evoked a rapid increase of glutamate overflow. Coadministration of the neurosteroid, dehydroepiandrosterone sulfate (DHEAS; 80 mg/kg; i.p.) prevented the thimerosal effect on glutamate and aspartate; the steroid alone had no influence on these amino acids. Coapplication of DHEAS with thimerosal in perfusion fluid also blocked the acute action of thimerosal on glutamate. In contrast, DHEAS alone reduced overflow of glycine and alanine, somewhat potentiating the thimerosal effect on these amino acids. Since excessive accumulation of extracellular glutamate is linked with excitotoxicity, our data imply that neonatal exposure to thimerosal-containing vaccines might induce excitotoxic brain injuries, leading to neurodevelopmental disorders. DHEAS may partially protect against mercurials-induced neurotoxicity.
59. Neonatal Administration of Thimerosal Causes Persistent Changes in Mu Opioid Receptors in the Rat Brain
Neurochem Res. 2010 November; 35(11): 1840–1847.
Mieszko Olczak, Michalina Duszczyk, Pawel Mierzejewski, Teresa Bobrowicz, and Maria Dorota Majewska1, Department of Pharmacology and Physiology of the Nervous System, Institute of Psychiatry and Neurology, Sobieskiego 9 str., 02-957 Warsaw, Poland, Department of Forensic Medicine, Medical University of Warsaw, Oczki 1 str., 02-007 Warsaw, Poland, Department of Neuropathology, Institute of Psychiatry and Neurology, 02-957 Warsaw, Poland, Department of Biology and Environmental Science, University of Cardinal Stefan Wyszynski, Wóycickiego Str. 1/3, 01-815 Warsaw, Poland
Thimerosal added to some pediatric vaccines is suspected in pathogenesis of several neurodevelopmental disorders. Our previous study showed that thimerosal administered to suckling rats causes persistent, endogenous opioid-mediated hypoalgesia. Here we examined, using immunohistochemical staining technique, the density of μ-opioid receptors (MORs) in the brains of rats, which in the second postnatal week received four i.m. injections of thimerosal at doses 12, 240, 1,440 or 3,000 μg Hg/kg. The periaqueductal gray, caudate putamen and hippocampus were examined. Thimerosal administration caused dose-dependent statistically significant increase in MOR densities in the periaqueductal gray and caudate putamen, but decrease in the dentate gyrus, where it was accompanied by the presence of degenerating neurons and loss of synaptic vesicle marker (synaptophysin). These data document that exposure to thimerosal during early postnatal life produces lasting alterations in the densities of brain opioid receptors along with other neuropathological changes, which may disturb brain development.
60. Unanswered Questions: A Review of Compensated Cases of Vaccine-Induced Brain Injury
Pace Environmental Law Review, vol. 28, no. 2, 2011
Mary Holland, Louis Conte, Robert Krakow and Lisa Colin
In 1986, Congress created the Vaccine Injury Compensation Program (VICP) under the National Childhood Vaccine Injury Act (1986 Law). This Program has original jurisdiction for children’s claims of vaccine injury. Because almost all children receive multiple vaccinations for daycare and school, it is critically important that the Program provides fundamental fairness, due process and transparency.
This empirical investigation, published in a peer-reviewed law journal, examines claims that the VICP compensated for vaccine-induced encephalopathy and seizure disorder. The VICP has compensated approximately 2,500 claims of vaccine injury since the inception of the program. This study found 83 cases of acknowledged vaccine-induced brain damage that include autism, a disorder that affects speech, social communication and behavior. In 21 published cases of the Court of Federal Claims, which administers the VICP, the Court stated that the petitioners had autism or described autism unambiguously. In 62 remaining cases, the authors identified settlement agreements where Health and Human Services (HHS) compensated children with vaccine-induced brain damage, who also have autism or an autism spectrum disorder.
Parents reported the existence of autism in telephone interviews and supplied supplemental materials including medical diagnoses, school records, and completed, standard autism screening questionnaires to verify their reports. In 39 of the 83 cases, or 47% of the cases of vaccine injury reviewed, there is confirmation of autism or autism spectrum disorder beyond parental report.
This finding of autism in compensated cases of vaccine injury is significant. U.S. government spokespeople have been asserting no vaccine-autism link for more than a decade. This finding calls into question the decisions of the Court of Federal Claims in the Omnibus Autism Proceeding in 2009 and 2010 and the statement of Health and Human Services on its website that “HHS has never concluded in any case that autism was caused by vaccination.”
Using publicly available information, the investigation shows that the VICP has been compensating cases of vaccine-induced brain damage associated with autism for more than twenty years. This investigation suggests that officials at HHS, the Department of Justice and the Court of Federal Claims may have been aware of this association but failed to publicly disclose it.
The study calls on Congress to thoroughly investigate the VICP, including a medical investigation of compensated claims of vaccine injury. This investigation calls on Congress to get answers to these critically important unanswered questions.
61. Abnormal measles-mumps-rubella antibodies and CNS autoimmunity in children with autism.
J Biomed Sci. 2002 Jul-Aug;9(4):359-64.
Singh VK, Lin SX, Newell E, Nelson C., Department of Biology and Biotechnology Center, Utah State University, Logan, Utah 84322, USA. [email protected]
Autoimmunity to the central nervous system (CNS), especially to myelin basic protein (MBP), may play a causal role in autism, a neurodevelopmental disorder. Because many autistic children harbor elevated levels of measles antibodies, we conducted a serological study of measles-mumps-rubella (MMR) and MBP autoantibodies. Using serum samples of 125 autistic children and 92 control children, antibodies were assayed by ELISA or immunoblotting methods. ELISA analysis showed a significant increase in the level of MMR antibodies in autistic children. Immunoblotting analysis revealed the presence of an unusual MMR antibody in 75 of 125 (60%) autistic sera but not in control sera. This antibody specifically detected a protein of 73-75 kD of MMR. This protein band, as analyzed with monoclonal antibodies, was immunopositive for measles hemagglutinin (HA) protein but not for measles nucleoprotein and rubella or mumps viral proteins. Thus the MMR antibody in autistic sera detected measles HA protein, which is unique to the measles subunit of the vaccine. Furthermore, over 90% of MMR antibody-positive autistic sera were also positive for MBP autoantibodies, suggesting a strong association between MMR and CNS autoimmunity in autism. Stemming from this evidence, we suggest that an inappropriate antibody response to MMR, specifically the measles component thereof, might be related to pathogenesis of autism.
62. Serological association of measles virus and human herpesvirus-6 with brain autoantibodies in autism.
Clin Immunol Immunopathol. 1998 Oct;89(1):105-8.
Singh VK, Lin SX, Yang VC. College of Pharmacy, University of Michigan, Ann Arbor, Michigan, 48109-1065, USA.
Considering an autoimmunity and autism connection, brain autoantibodies to myelin basic protein (anti-MBP) and neuron-axon filament protein (anti-NAFP) have been found in autistic children. In this current study, we examined associations between virus serology and autoantibody by simultaneous analysis of measles virus antibody (measles-IgG), human herpesvirus-6 antibody (HHV-6-IgG), anti-MBP, and anti-NAFP. We found that measles-IgG and HHV-6-IgG titers were moderately higher in autistic children but they did not significantly differ from normal controls. Moreover, we found that a vast majority of virus serology-positive autistic sera was also positive for brain autoantibody: (i) 90% of measles-IgG-positive autistic sera was also positive for anti-MBP; (ii) 73% of measles-IgG-positive autistic sera was also positive for anti-NAFP; (iii) 84% of HHV-6-IgG-positive autistic sera was also positive for anti-MBP; and (iv) 72% of HHV-6-IgG-positive autistic sera was also positive for anti-NAFP. This study is the first to report an association between virus serology and brain autoantibody in autism; it supports the hypothesis that a virus-induced autoimmune response may play a causal role in autism.
63. Integrating experimental (in vitro and in vivo) neurotoxicity studies of low-dose thimerosal relevant to vaccines.
Neurochem Res. 2011 Jun;36(6):927-38. doi: 10.1007/s11064-011-0427-0. Epub 2011 Feb 25.
Dórea JG, Faculty of Health Sciences, Universidade de Brasília, CP 04322, 70919-970, Brasília, DF, Brazil. [email protected]
There is a need to interpret neurotoxic studies to help deal with uncertainties surrounding pregnant mothers, newborns and young children who must receive repeated doses of Thimerosal-containing vaccines (TCVs). This review integrates information derived from emerging experimental studies (in vitro and in vivo) of low-dose Thimerosal (sodium ethyl mercury thiosalicylate). Major databases (PubMed and Web-of-science) were searched for in vitro and in vivo experimental studies that addressed the effects of low-dose Thimerosal (or ethylmercury) on neural tissues and animal behaviour. Information extracted from studies indicates that: (a) activity of low doses of Thimerosal against isolated human and animal brain cells was found in all studies and is consistent with Hg neurotoxicity; (b) the neurotoxic effect of ethylmercury has not been studied with co-occurring adjuvant-Al in TCVs; (c) animal studies have shown that exposure to Thimerosal-Hg can lead to accumulation of inorganic Hg in brain, and that (d) doses relevant to TCV exposure possess the potential to affect human neuro-development. Thimerosal at concentrations relevant for infants’ exposure (in vaccines) is toxic to cultured human-brain cells and to laboratory animals. The persisting use of TCV (in developing countries) is counterintuitive to global efforts to lower Hg exposure and to ban Hg in medical products; its continued use in TCV requires evaluation of a sufficiently nontoxic level of ethylmercury compatible with repeated exposure (co-occurring with adjuvant-Al) during early life.
64. Hepatitis B vaccine induces apoptotic death in Hepa1-6 cells
Apoptosis. 2012 Jan 17. Hamza H, Cao J, Li X, Li C, Zhu M, Zhao S.
Key Lab of Agricultural Animal Genetics, Breeding, and Reproduction of Ministry of Education, College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, 430070, People’s Republic of China, [email protected]
Vaccines can have adverse side-effects, and these are predominantly associated with the inclusion of chemical additives such as aluminum hydroxide adjuvant. The objective of this study was to establish an in vitro model system amenable to mechanistic investigations of cytotoxicity induced by hepatitis B vaccine, and to investigate the mechanisms of vaccine-induced cell death. The mouse liver hepatoma cell line Hepa1-6 was treated with two doses of adjuvanted (aluminium hydroxide) hepatitis B vaccine (0.5 and 1 μg protein per ml) and cell integrity was measured after 24, 48 and 72 h. Hepatitis B vaccine exposure increased cell apoptosis as detected by flow cytometry and TUNEL assay. Vaccine exposure was accompanied by significant increases in the levels of activated caspase 3, a key effector caspase in the apoptosis cascade. Early transcriptional events were detected by qRT-PCR. We report that hepatitis B vaccine exposure resulted in significant upregulation of the key genes encoding caspase 7, caspase 9, Inhibitor caspase-activated DNase (ICAD), Rho-associated coiled-coil containing protein kinase 1 (ROCK-1), and Apoptotic protease activating factor 1 (Apaf-1). Upregulation of cleaved caspase 3,7 were detected by western blot in addition to Apaf-1 and caspase 9 expressions argues that cell death takes place via the intrinsic apoptotic pathway in which release of cytochrome c from the mitochondria triggers the assembly of a caspase activation complex. We conclude that exposure of Hepa1-6 cells to a low dose of adjuvanted hepatitis B vaccine leads to loss of mitochondrial integrity, apoptosis induction, and cell death, apoptosis effect was observed also in C2C12 mouse myoblast cell line after treated with low dose of vaccine (0.3, 0.1, 0.05 μg/ml). In addition In vivo apoptotic effect of hepatitis B vaccine was observed in mouse liver.
65. Maternal thimerosal exposure results in aberrant cerebellar oxidative stress, thyroid hormone metabolism, and motor behavior in rat pups; sex- and strain-dependent effects.
Cerebellum. 2012 Jun;11(2):575-86. doi: 10.1007/s12311-011-0319-5.
Sulkowski ZL, Chen T, Midha S, Zavacki AM, Sajdel-Sulkowska EM, Department of Psychiatry, Harvard Medical School and Brigham and Women’s Hospital, Boston, MA, USA.
Methylmercury (Met-Hg) and ethylmercury (Et-Hg) are powerful toxicants with a range of harmful neurological effects in humans and animals. While Met-Hg is a recognized trigger of oxidative stress and an endocrine disruptor impacting neurodevelopment, the developmental neurotoxicity of Et-Hg, a metabolite of thimerosal (TM), has not been explored. We hypothesized that TM exposure during the perinatal period impairs central nervous system development, and specifically the cerebellum, by the mechanism involving oxidative stress. To test this, spontaneously hypertensive rats (SHR) or Sprague-Dawley (SD) rat dams were exposed to TM (200 μg/kg body weight) during pregnancy (G10-G15) and lactation (P5-P10). Male and female neonates were evaluated for auditory and motor function; cerebella were analyzed for oxidative stress and thyroid metabolism. TM exposure resulted in a delayed startle response in SD neonates and decreased motor learning in SHR male (22.6%), in SD male (29.8%), and in SD female (55.0%) neonates. TM exposure also resulted in a significant increase in cerebellar levels of the oxidative stress marker 3-nitrotyrosine in SHR female (35.1%) and SD male (14.0%) neonates. The activity of cerebellar type 2 deiodinase, responsible for local intra-brain conversion of thyroxine to the active hormone, 3′,3,5-triiodothyronine (T3), was significantly decreased in TM-exposed SHR male (60.9%) pups. This coincided with an increased (47.0%) expression of a gene negatively regulated by T3, Odf4 suggesting local intracerebellar T3 deficiency. Our data thus demonstrate a negative neurodevelopmental impact of perinatal TM exposure which appears to be both strain- and sex-dependent.
66. The rise in autism and the role of age at diagnosis.
Epidemiology. 2009 Jan;20(1):84-90. doi: 10.1097/EDE.0b013e3181902d15.
Hertz-Picciotto I, Delwiche L., Department of Public Health Sciences, University of California, Davis, California 95616, USA. [email protected]
Autism prevalence in California, based on individuals eligible for state-funded services, rose throughout the 1990s. The extent to which this trend is explained by changes in age at diagnosis or inclusion of milder cases has not been previously evaluated.
Autism cases were identified from 1990 through 2006 in databases of the California Department of Developmental Services, which coordinates services for individuals with specific developmental disorders. The main outcomes were population incident cases younger than age 10 years for each quarter, cumulative incidence by age and birth year, age-specific incidence rates stratified by birth year, and proportions of diagnoses by age across birth years.
Autism incidence in children rose throughout the period. Cumulative incidence to 5 years of age per 10,000 births rose consistently from 6.2 for 1990 births to 42.5 for 2001 births. Age-specific incidence rates increased most steeply for 2- and 3-year olds. The proportion diagnosed by age 5 years increased only slightly, from 54% for 1990 births to 61% for 1996 births. Changing age at diagnosis can explain a 12% increase, and inclusion of milder cases, a 56% increase.
Autism incidence in California shows no sign yet of plateauing. Younger ages at diagnosis, differential migration, changes in diagnostic criteria, and inclusion of milder cases do not fully explain the observed increases. Other artifacts have yet to be quantified, and as a result, the extent to which the continued rise represents a true increase in the occurrence of autism remains unclear.
67. Enrichment of Elevated Plasma F2t-Isoprostane Levels in Individuals with Autism Who Are Stratified by Presence of Gastrointestinal Dysfunction
PLoS ONE 8(7): e68444.
Gorrindo P, Lane CJ, Lee EB, McLaughlin B, Levitt P (July 3, 2013)
Funding: This work was supported in part by National Institutes of Health awards National Institute of Child Health and Human Development R21HD065289 (PL), National Institute of General Medical Sciences T32GM07347 for the Vanderbilt Medical Scientist Training Program (PG), National Center for Research Resources TL1RR024978 (PG), and National Center for Advancing Translational Sciences UL1TR000445 for the Vanderbilt Institute for Clinical and Translational Research. Additional support was provided by the Marino Autism Research Institute, the Pediatric Clinical Research Center at Vanderbilt University, The Scott Family Foundation, and the Vanderbilt Autism Treatment Network Site, a program funded by Autism Speaks.
Etiology is unknown in the majority of individuals with autism spectrum disorder (ASD). One strategy to investigate pathogenesis is to stratify this heterogeneous disorder based on a prominent phenotypic feature that enriches for homogeneity within population strata. Co-occurring gastrointestinal dysfunction (GID) characterizes a subset of children with ASD. Our current objective was to investigate a potential pathophysiological measure to test the hypothesis that children with both ASD and GID have a more severe metabolic dysfunction than children with ASD-only, given that the highly metabolically active brain and gastrointestinal system may additively contribute measurable impairment. Plasma levels of F2t-Isoprostanes (F2-IsoPs), a gold standard biomarker of oxidative stress, were measured in 87 children in four groups: ASD-GID, ASD-only, GID-only and Unaffected. F2-IsoP levels were elevated in all 3 clinical groups compared to the Unaffected group, with the ASD-GID group significantly elevated above the ASD-only group (mean, SD in pg/mg: ASD-GID 53.6, 24.4; ASD-only 36.5, 13.3; p = 0.007). Adjusting for age, sex, and triglyceride levels, F2-IsoP levels remained significantly different between study groups, with a moderate effect size of ηp2 = 0.187 (p = 0.001). Elevation in peripheral oxidative stress is consistent with, and may contribute to, the more severe functional impairments in the ASD-GID group. With unique medical, metabolic, and behavioral features in children with ASD-GID, the present findings serve as a compelling rationale for both individualized approaches to clinical care and integrated studies of biomarker enrichment in ASD subgroups that may better address the complex etiology of ASD.