Prenatal exposure to antibodies from mothers of children with autism produces neurobehavioral alterations: A pregnant dam mouse model
Introduction
Autism is a neurodevelopmental disorder characterized by core clinical deficits in the areas of socialization, verbal and nonverbal communication, and restricted and repetitive patterns of behavior. The disorder is complex, encompasses a broad range of pervasive developmental disabilities, and in the majority of cases there is no defined etiology. Suggestions of an immune abnormality as a contributing factor in autism were initially based on the presence of a family history of autoimmune dysfunction (Comi et al., 1999) and the identification of serum antibodies targeting brain epitopes in affected children (Plioplys et al., 1989, Singer et al., 2006, Singh et al., 1997, Singh et al., 1993, Todd and Ciaranello, 1985, Zimmermann, 1993). Subsequent studies, however, have shifted the hypothesis from a primary autoimmune abnormality to one of potential transplacental antibody exposure in utero. More specifically, several investigators have identified the presence of unique serum antibodies against human fetal brain tissue in mothers with offspring having the diagnosis of autistic spectrum disorders (Braunschweig et al., 2008, Croen et al., 2008, Singer et al., 2008). A correlation between maternal antibodies against fetal brain proteins in the 36–39 kDa range and the presence of developmental regression in affected offspring has been used to support a functional association (Braunschweig et al., 2008, Singer et al., 2008).
Although antibody detection in mothers of children with autistic disorders (MCAD) raises the possibility that placentally transferred antibodies interfere with fetal brain development, their presence alone is insufficient to define causality. For example, autoantibodies are present in healthy individuals as well as mothers of typically developing offspring and may even have a regulatory role (Archelos and Hartung, 2000). Hence, the confirmation of a pathogenic role for maternal antibodies in neurodevelopmental disorders or autism requires: demonstrating that the passive transfer of maternal IgG can induce a behavioral disorder in offspring; showing that immunoglobulins are present at the pathological site; and defining the mechanism(s) by which passively transferred IgG alters brain development.
The pregnant dam (mouse) model provides a valuable system for investigating the IgG placental transfer hypothesis. The putative pathological antibody(ies) can be injected during the gestational period and offspring evaluated for changes in behavior. Similar approaches have identified behavioral abnormalities in offspring following exposure of pregnant dams to serum antibodies from mothers of dyslexic children and from a mother with anti-Purkinje antibodies who had an autistic child (Dalton et al., 2003, Vincent et al., 2003). In addition, mouse behavioral paradigms are available to evaluate overactivity, anxiety, and sociability, clinical deficits that occur in children with neurodevelopmental disorders (Crawley, 2007, Moy et al., 2007). While studies in more advanced species, e.g., monkeys, (Martin et al., 2008) might provide a broader social repertoire, the mouse model is more economical, permits testing in larger numbers, and provides the ability to analyze embryonic brain for inflammation and structural alterations.
The goal of this exploratory study was to use the pregnant mouse model to establish whether the intraperitoneal injection of IgG during gestation, obtained from mothers of children with autistic disorder, produces postnatal behavioral effects in offspring. Behavior was compared to that in mice whose dams received IgG from mothers of typically developing children (IgG-controls) and simple-controls. Basic observations of neurobehavioral and neuromotor development and weight gain were recorded during the first 21 days of life. Behavioral paradigms, used in adolescent and adult mice, included measures of open field behavior and activity, anxiety-like behavior, acoustic startle response, pre-pulse inhibition, and sociability. Pilot investigations in embryonic mouse brains were designed to confirm a maternal-to-fetal transfer of human IgG, to assess for gross neuroanatomical alterations, and to identify an immune reaction by use of cytokine measurement, immunohistochemical staining for microglial reactivity, and ELISA assay of brain derived neurotrophic factor (BDNF). It was hypothesized that behavioral differences would be identified in adolescent and adult mice prenatally exposed to MCAD-IgG and that evidence of immune reactivity would be found in the embryonic brain.
Section snippets
Animals
C57BL/J6 male and female mice were purchased from Jackson Laboratory. Mice were maintained and mated in the certified animal facility of the Johns Hopkins Broadway Research Building. All investigations in offspring were performed without regard to gender.
Isolation of IgG from human sera
IgG was isolated from pooled serum samples of 63 mothers of children with autistic disorder (MCAD) and 63 mothers of unaffected children (IgG-controls), participants in a previously published study (Singer et al., 2008) (see Table 1). Autistic
Animal health, numbers, and gender
Following intraperitoneal injections, no pregnant dam experienced any evidence of illness, decreased activity, or change in dietary intake. Litter sizes in MCAD, IgG-control, and saline-control groups were equivalent. All pups were in good general health and body weights from P1-21 days (Fig. 1) remained equivalent.
The number of animals participating in each behavioral task analysis is shown in Table 3. Gender ratios for the adolescent and adult behavioral tests were as follows: MCAD 45% male,
Discussion
The goal of this study was to investigate the hypothesis that placentally-transferred antibodies are pathogenic in neurodevelopmental disorders. More specifically, we sought to determine whether, in a pregnant mouse model, the transfer of MCAD IgG could induce a behavioral disorder that mimics some symptoms seen in autism. Although many human behavioral signs and symptoms cannot be replicated in mice, evaluations of activity, social interaction, anxiety, response to sensory stimuli, and motor
Acknowledgement
Research was supported by a grant from the Hussman Foundation.
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