https://youtu.be/ioMspoSNgmw

Introduction

Autism and Asperger’s disorder (a better prognostic variety) that used to be classified under pervasive developmental disorders (PDD) in DSM-IV (1) are currently grouped under autistic spectrum disorders (ASD) in DSM-5 (2). These disorders were not recognized in the first and second editions of DSM in 1952 (3) and 1968 (4) and the closest term to them was “schizophrenic reaction of childhood type”! DSM-III in 1980 (5) introduced the PDD and classified them to childhood onset PDD, infantile autism, and atypical autism. In 1987, DSM-III-R (6) classified these disorders into PDD-NOS (not otherwise specified) and autistic disorder. In 1994, DSM-IV (7) classified these disorders into PDD-NOS, autistic disorder, Asperger disorder, childhood disintegrative disorder and Rett syndrome, that did not change in DSM-IV-TR in 2000. (8) The new DSM 5 published in may of 2014, with the explanation that all these disorders are along a spectrum and labeled them under the new term “Autistic Spectrum Disorders”.  

Regardless of the terminology, ASD like schizophrenia are neurodevelopmental disorders and that is why in the past was classified under childhood psychoses or schizophrenic reaction of childhood. (9-13) These neurodevelopmental disorders, while share some common features in underlying pathogenesis and clinical manifestations, they have their own specific differences at both pathophysiologic and phenomenological levels. These disorders etiopathologically share an early insult to the developing brain, e.g. the prenatal maternal infections that trigger maternal and fetal immune reactions, damaging the fetal neurodevelopment. Although the insult such as infection could be non-specific, the severity, acuteness, latency of the invasion and the time window of neurodevelopment, determines which disorder to be manifested. (14) Moreover the pathophysiology of these neurodevelopmental disorders seems to be a multi-steps process. In the first step there is the microbial attack at a specific time of neurodevelopment that triggers the second step of maternal and fetal immune reaction response that would be damaging the fetal brain development. In the third step, the microbial invasion causes genetic mutations to pass on its impact to the next generation through genetic inheritance. In this paper, I will strive to present research evidence of the link between infective insults and the causation of ASD, e.g. autism, so hopefully to get us closer to the prevention of such disorders, by intervening in any of the above-mentioned steps, as the current treatments are nothing but symptomatic reliefs at the best!

First: The Prenatal Maternal Infections

The first time in the literature, Desmond et al. (15) in 1967 reported a link between congenital rubella and autism, but without discussing the criteria of autism and the detail of behavioral symptoms. In 1971 Stella Chess, well known for the first theory and study on children temperament with her husband, Alexander Thomas, reported 243 children ages 2.5-5 years old with congenital rubella meeting Kanner’s srict criteria for autism. (16) Six years later, Chess in a follow up report of these children reported significant improvement in the most autistic symptoms, even low intelligence. But Chess does not claim that viral infection of the brain is the cause of autism and suspects “autism may be the final behavioral consequence of many different causes”. (17) Soon cases of congenital cytomegalovirus infection were linked with autistic symptoms as early as 6 months of age, (18-20) and the infection hypothesis of autism was established as early as late 70’s and early 80’s! (21) most recently Engman et al. (22) from Sweden reported that only one of 33 children with autism spectrum disorder and intellectual disability of 115 preschool children with ASD had cytomegalovirus infection. In recent years also cytomegalovirus infection as an etiology of autism has been reported by Yamashita et al. (23) from Japan.

 The infection theory of autism has raised some concerns about the impact of common vaccines to children, specially MMR (Measles-mumps-rubella) vaccine. Although some biological assays have lent support to the association between measles virus or MMR and autism, some epidemiologic studies have shown no association between MMR and autism. This suspicion and concern led the American Academy of Pediatrics in 2000 to convene a conference including parents, practitioners, and scientists presenting for and against the link between MMR vaccine and ASD (Autistic Spectrum Disorders). (24) Despite admitting to the possibility of such link, the reporters of the conference concluded that there are not sufficient evidence to support such hypothesis and there would be no difference between risk of combination of that MMR vaccine and separate administration of measles, mumps, and rubella vaccines to children.

Atladóttir et al. (25) in their large and longitudinal study on all children born in Denmark from 1980, through 2005, have reported no association between any maternal infection and diagnosis of ASDs in the child when looking at the total period of pregnancy, but high admission rates to hospital due to maternal viral infection in the first trimester and maternal bacterial infection in the second trimester associated with diagnosis of ASDs in the offspring. Lintas et al. (26) by nested polymerase chain reaction (PCR) ) and DNA sequence analysis in the postmortem brains of autistic patients found more significant polyomavirus of BK virus (BKV), JC virus (JCV), and simian virus 40 (SV40) types among autistic patients compared to controls (67% versus 23%, respectively; P < .05). Also polyviral infections tend to occur more frequently in the brains of autistic patients compared to controls (40% versus 7.7%, respectively; P =.08). It should be noted that JCV causes multifocal leukoencephalopathy, and BKV causes respiratory infection, nephropathy and immunosuppression in humans, and the polyomaviruses are potentially oncogenic by causing persistent latent infections in a host without causing disease {polyoma refers to the viruses’ ability to produce multiple (poly-) tumors (-oma)}. (27-28)

Animal models of autism by injecting viral particles have also supported the infectious hypothesis of autism. (29) Autistic like behaviors in the offsprings of rodents have been shown after the maternal injections of influenza virus, Borna virus, cytomegalovirus, and synthetic viral particles, e.g. polyinosine:cytosine (poly(I:C)) or lipopolysaccharide (LPS). (30-34) Higher titers of autoantibodies to brain proteins/antigens have been reported in autistic subjects. (34-35) Also cytokines, such as interleukin (IL)-6, tumor necrosis factor (TNF)-and IL-1have been found in the brain and blood of autistic patients. (36-37) Shi et al. (32) reported that maternal injection of influenza virus causes deficits in social interaction, prepulse inhibition (PPI), and explor- atory behavior in the adult offsrpings. Smith et al have shown that maternal injection of poly(I:C), which evokes an antiviral-like immune response, causes behavioural deficits in the adult offspring. This indicates that the maternal immune response is sufficient to cause changes in the behavior of the adult offspring. Additionally, Smith et al. (33) demonstrated that IL-6 was elevated in poly(I:C) model mice, and co-administration of an anti-IL-6 antibody, but not an anti-TNF-antibody, prevents the social interaction and PPI deficits caused by poly(I:C) in adult offspring. Similarly, maternal injection of poly(1:C) into IL-6 knockout mice does not effect all of the expected behavioral changes. Although Borna disease virus (BDV) has not been implicated in the pathogenesis of autism, neonatal BDV infection profoundly affected social behaviors and stereotypic behaviors in adult rats. (34) In these rats, glial actication is prominent throughout the brain and persists for several weeks in concert with increased levels of proinflammatory cytokine mRNAs including IL-6 mRNA. (38)

Bolte (39) has reported an association between subacute and chronic tetanus infection of the intestinal trac with the development of autism. This researcher has hypothesized that Clostridium tetani produces a potent neurotoxin (TeNT) that has been well demonstrated in laboratory animals as well, that is transported by the vagus nerve to the CNS. This route bypasses TeNT’s normal preferential binding sites in the spinal cord, and therefore the symptoms of a typical tetanus infection are not evident. Once in the brain, TeNT disrupts the release of neurotransmitters by the proteolytic cleavage of synaptobrevin, a synaptic vesicle membrane protein. This inhibition of neurotransmitter release would explain a wide variety of behavioral deficits apparent in autism. Lab animals injected in the brain with TeNT have exhibited many of these behaviors. Some children with autism have also shown a significant reduction in stereotyped behaviors when treated with antimicrobials effective against intestinal clostridia.

Second: The Maternal and Fetal Immune Reactions

Higher titers of autoantibodies to brain proteins/antigens have been reported in autistic subjects. Also cytokines, such as interleukin (IL)-6, tumor necrosis factor (TNF)-and IL-1has been found in the brain and blood of autistic patients. Many viruses or immunostimulants, including influenza virus, cytomegalovirus, polyinosinic: poly- cytidylic acid [poly(I:C)], and bacterial lipopolysac- charide have been used to established animal models of psychiatric and neurodevelopmental disorders such as schizophrenia and autism. Shi et al reported that maternal injection of influenza virus causes deficits in social interaction, prepulse inhibition (PPI), and explor- atory behavior in the adult offsrpings. Smith et al have shown that maternal injection of poly(I:C), which evokes an antiviral-like immune response, causes behavioural deficits in the adult offspring. This indicates that the maternal immune response is sufficient to cause changes in the behavior of the adult offspring. Additionally, Smith et al demon- strated that IL-6 was elevated in poly(I:C) model mice, and co-administration of an anti-IL-6 antibody, but not an anti-TNF-antibody, prevents the social interac- tion and PPI deficits caused by poly(I:C) in adult offspring. Similarly, maternal injection of poly(1:C) into IL-6 knockout mice does not effect all of the expected behavioral changes.

Although Borna disease virus (BDV) has not been implicated in the pathogenesis of autism, neonatal BDV infection profoundly affected social behaviors and stereotypic behaviors in adult rats. In these rats, glial actication is prominent throughout the brain and persists for several weeks in concert with increased levels of proinflammatory cytokine mRNAs including IL-6 mRNA. These findings suggest that IL-6 as a key intermediary should aid in the molecular dissection of the pathways whereby maternal immune activation alters brain development. IL-6 induces Janus tyrosine kinase-2/ signal transducer and activator of transcription-3 (JAK2/STAT3) phosphorylation, and induction of neu- ronal JAK2/STAT3 phosphorylation following IL-6 challenge led to significant deficits in social interaction behaviors in mice. Another potential mechanism is related to the inhibitory effect of IL-6 on DNA methylation. It has been reported that IL-6 exerts many epigenetic changes in cells via increasing expression of the DNA (cytosine-5-)-methyltransferase 1 gene (DNMT1). DNMT1 transfers a methyl group to the cytosine portion of the CpG dinucleotide, and permits or enables the binding of methyl-specific DNA-binding proteins to the methylated CpG site. Methylcytosine DNA-binding proteins can attract histone deacetylases.

Hornig et al. (40) describe a model for linking infections such as Borna virus with the development of neuro-developmental disorders such as ASD in rats, showing abnormal righting reflexes, hyperactivity, inhibition of open-field exploration, and stereotypic behaviors. These abnormal behaviours corresponding some with ASD in humans, are the results of marked disruption in hippocampus and cerebellum, with reduction in granule and Purkinje cell numbers, and neuronal loss predominantly by apoptosis. Although these inflammatory infiltrates are observed transiently in frontal cortex, glial activation (microgliosis > astrocytosis) is prominent throughout the brain and persists for several weeks in concert with increased levels of proinflammatory cytokine mRNAs (interleukins 1alpha, 1beta, and 6 and tumor necrosis factor alpha) and progressive hippocampal and cerebellar damage. Singh et al. (41-42) have conducted a serologic study of measles, mumps, and rubella viruses in the serum of autistic children in comparison with normal children, and siblings of autistic children. These researchers reported that the level of measles antibody, but not mumps or rubella antibodies, was significantly higher in autistic children as compared with normal children or siblings of autistic children. The antibody to the measles virus antigen was found in 83% of autistic children but not in normal children or siblings of autistic children. This research lab replicated their findings in another study recently. (43)

 The human immune reaction to infections in autism has been demonstrated in many researches. (e.g.44-55) The immunopathogenesis of autism has been associated with the presence of pro-inflammatory cytokines and autoimmunity involving MHC region and HLA, as immune reactions to infectious agents such as measles, herpesvirus-6, etc. The cell-mediated immunity in autism has been shown to be impaired as evidenced by low numbers of CD4 cells and a concomitant T-cell polarity with an imbalance of Th1/Th2 subsets toward Th2. Impaired humoral immunity on the other hand is evidenced by decreased IgA causing poor gut protection. Studies showing elevated brain specific antibodies in autism also support an autoimmune mechanism. Viruses may initiate the process but the subsequent activation of cytokines and other pro-inflammatory agents are the damaging factor causing autism. Inflammatory mediators in autism involve activation of astrocytes and microglial cells. Proinflammatory chemokines (MCP-1 and TARC), and an anti-inflammatory and modulatory cytokine, TGF-beta1, are consistently elevated in autistic brains. In measles virus infection, it has been postulated that there is immune suppression by inhibiting T-cell proliferation and maturation and downregulation MHC class II expression. Cytokine alteration of TNF-alpha is increased in autistic populations. Maternal antibodies may also trigger autism as a mechanism of autoimmunity. MMR antibodies are significantly higher in autistic children as compared to normal children, supporting a role of MMR in autism. Autoantibodies (IgG isotype) to neuron-axon filament protein (NAFP) and glial fibrillary acidic protein (GFAP) are significantly increased in autistic patients (Singh et al., 1997). Increase in Th2 may explain the increased autoimmunity, such as the findings of antibodies to MBP and neuronal axonal filaments in the brain. (44-46)

Blaylock and Strunecka (47) have suggested that most heterogeneous symptoms of ASD have a common immunopathological pathway associated with dysregulation of glutamatergic neurotransmission in the brain with enhancement of excitatory receptor function by pro-inflammatory immune cytokines as the underlying mechanism. Maternal immune activation (MIA) to infections in second and third trimesters have been proposed in etiology of neuro-developmental disorders such as schizophrenia and autism by researchers, e.g. Meyer et al. (48) and Smith et al. (49), but with no specific differentiation between these disorders, the infectious agents, specific immunological reactions and the specific timing of pregnancy. The footprint of an early prenatal infection has been evidenced by a longer, ongoing hyper-responsive inflammatory-like state in many young as well as adult autism subjects, years after the first impact in the cerebral spinal fluid, blood and postmortem brains of autistic subject! (50) This impact is not only limited to immune-pathogenesis but lead to strong and common gene expression changes in the embryonic brain. Most notably, there is an acute and transient upregulation of the α, β and γ crystallin gene family. Furthermore, levels of crystallin gene expression are correlated with the severity of MIA as assessed by placental weight. (51)

 Many of the proteins encoded by the major histocompatibility complex (MHC) play a vital role in the formation, refinement, maintenance, and plasticity of the brain. Manipulations of levels of MHC molecules by prenatal infections have been shown to significantly alter brain connectivity and function, evidenced in ASD (Autism Spectrum Disorders). (52) Elevation of cerebrospinal fluid levels of tumor necrosis factor-alpha has been shown to be significantly higher than controls in autistic patients. (53) Active neuroinflammatory process in the cerebral cortex, white matter, and notably in cerebellum have been also shown in autistic patients. Immunocytochemical studies have also demonstrated marked activation of microglia and astroglia, and macrophage chemoattractant protein (MCP)-1 and tumor growth factor-beta1, derived from neuroglia, as the most prevalent cytokines in brain tissues of autistic subjects. (54) Elevated different cytokines have also been shown in many studies of autistic subjects, as early as age 2,  as signs of neuro-inflammation of their brains but with no link to any specific one. In addition, increasing cytokine levels are associated with more impaired communication and aberrant behaviors in autistic children. (55-56) Perhaps the most recent evidence of neuro-inflammation in autism as a result of an infection invasion, has been the evidence of increased serum levels of high mobility group box 1 protein in patients with autistic disorders. High-mobility group box 1 protein (HMGB1) is a nuclear protein that is released passively when cells such as neurons die after an insult such as microbial invasions. HMGB1 behaves as a trigger of inflammation, attracting inflammatory cells, and of tissue repair, recruiting stem cells and promoting their proliferation. Moreover, HMGB1 activates dendritic cells (DCs) and promotes their functional maturation and their response to lymph node chemokines. Activated leukocytes actively secrete HMGB1 in the microenvironment. Thus, HMGB1 acts in an autocrine/paracrine fashion and sustains long-term repair and defense programs. These immune responses will also be directed against self-antigens that DCs process at the time of injury and can lead to autoimmunity.

 Despite the hardly debatable scientific evidence in the literature of the link between prenatal infections and causation of ASD and other neuro-developmental disorders, through an neuro-inflammatory reaction of the body, there is yet a lack of direct evidence between any specific infection, any specific inflammatory reaction and any specific neuro-developmental disorder. Therefore and thus far it seems that any prenatal infections, viral or bacterial, could cause an inflammatory reaction in defense in the pregnant mother and fetus through cytokines, MHC and HLA systems that disrupt the fetal developing brain and causing any of the neurodevelopmental disorders. Now lets see if there is any specific neuro-developmental time window in causing any of the neuro-developmental disorders specifically. Knowing what specific infection and what specific inflammatory reaction or at least what specific neuro-developmental time window causing what specific neuro-developmental disorder is essential for the future in early prediction and even prevention of any of these disorders, as the current treatment of these disorders have never led to acceptable symptomatic reliefs, lest recovery!

Third: The Sensitive Neurodevelopmental Time Window

Meyer et al. (60), while believe that there could be “specific gestational Windows” associated with a differing vulnerability to infection-mediated disturbances in normal brain development, and admit to the debate and uncertainty of these specific time windows, presume that “the earlier the worse” would be such risks. These researchers’ explanation is “infections in early gestation may not only interfere with fundamental neurodevelopmental events such as cell proliferation and differentiation, but it may also predispose the developing nervous system to additional failures in subsequent cell migration, target selection, and synapse maturation, eventually leading to multiple brain and behavioral abnormalities in the adult offspring.” Marco et al. (61) also stress on the critical time windows of brain development that upon disruption by stress or infections, causing neuro-developmental disorders. These researchers, even through their animal models, could not delineate any specific prenatal time windows linking to any specific disorder, but assert on the general specific time windows, “i.e. prenatal stage, early infancy and adolescence”.

 Fang et al. (62) from Tawian in a very recent case-control study have associated bacterial and genital tract infections during the third trimester with the risk of ASD. Fatemi et al. (63) investigated the impact of maternal exposure to human influenza virus (H1N1) in lab mice on Day 9 of pregnancy (second trimester) on the brain development. In Day 0 infected mice vs. controls, showed significant increase of 170% in pyramidal cell density and 33% decrease in nonpyramidal cell density, with 29% decrease in the pyramidal cell nuclear size. Fourteen-week-old exposed mice continued to show significant increases in both pyramidal and non pyramidal cell density, and 37-43% decrease in pyramidal cell nuclear size vs. controls. Brain and ventricular area measurements in adult exposed mice also showed significant increases and decreases respectively vs. controls. While the rate of pyramidal cell proliferation per unit area decreased from birth to adulthood in both control and exposed groups, nonpyramidal cell growth rate increased only in the exposed adult mice. Although this animal data show the early experimental prenatal exposure to an infection such as influenza virus has both short-term and long-lasting deleterious effects on developing brain structure with implications for neuro-developmental disorders such as autism and schizophrenia as the authors suggest, yet there is no accurate time window detection to differentiate among these disorders. Winter et al. (64) also demonstrated that influenza viral infection in late first trimester and middle-late second trimester in pregnant mice significantly alters levels of serotonin, 5-hydroxyindoleacetic acid, and taurine, modeling disruptions that occur in patients with schizophrenia and autism. Finally and most recently Lee et al. have reported that timing of prenatal infection does not appear to influence risk of ASD in their Swedish population, as infection in all trimesters could lead to these disorders.

 Four: The Genetic damage or vulnerability

As in schizophrenia (read the post “from infection to schizophrenia”) genetics does not seem to be directly involved, as the concordance rate for twins is only 60% and none for dizygotic twins. Moreover the candidate genes in autism, while may share with other neuro-developmental or other psychiatric disorders, seem to be more associated with the phenotypic surface of the disorder, e.g. speech, language, and social aptitudes (region at 7q31-q33, cytogenetic abnormalities at the 15q11-q13 locus, & chromosome 15) than any direct underlying pathophysiology. (66) Lastly like in schizophrenia, the genetic vulnerability in autism seems to be a byproduct of infections footprint in our genome in the form of mutative CNVs (Copy Number Variants). (67)

 Conclusion

Autism and other autism spectrum disorders (ASD) like any other neuro-developmental disorders such as schizophrenia, seem to be the result of prenatal infections that in turn cause maternal and fetal inflammatory reactions that all disrupt the normal brain development. While we have come so far to know this, we are still at the tip of the discovery iceberg, there is not yet any specificity in the type of infection, inflammatory reactions and the timing of the prenatal damage. If the clinical onset of any of this disorders could lead us to any timing assumptions, then it seems that the earlier the onset of the disorder, e.g. autism would be associated with an earlier infection, e.g. in the first trimester. In the same token, a later onset disorder such as schizophrenia, a later infection, e.g. in the second trimester and if we include disorders such as Bipolar Disorder, it could have been caused by infection insults in the third trimester! But these assumptions, while holding some factual evidence (e.g. 60), are in strong need of validation by the future research aiming specifically to find such timing links. The future research needs to identify if there is any specific associations between the type of prenatal infections or even maternal and fetal inflammatory reactions to any of these disorders, or all are non-specific and only timing of the insults is the crucial determining factor!

For the time being, as far as we are certain of the role of prenatal infections in the causation of autism and other neurodevelopment disorders, the prevention seems to be easier than the treatment!  By educating the public, specially the mother about the role of infections during any stage of pregnancy and the risk of these disorders, use of low risk vaccinations, treating or preventing the infections could be a great step towards the prevention of these life-long disorders. Although we may not be able to prevent or treat any of these maternal infections, or the pregnant women may not wish to end their pregnancy for such a possible not proven risk, identifying the maternal and fetal inflammatory reactions that could be the main sharp edge of the insult, causing these disorders, could be a better feasible preventive approach!  While we cannot recommend abortions upon presence of prenatal infections, we may be able to do so when destructive pathological maternal and fetal inflammatory reactions have been detected and presented to the pregnant mother and her family! 

Dr.Mostafa Showraki, MD, FRCPC                                                               Lecturer, University of Toronto,School of Medicine,Author: “ADHD:Revisited” Book “adhdrevisited.com”/”medicinerevisited.com”

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