Herein we present a detailed analysis of maternal antibodies to human fetal brain in a large cohort of mothers for whom detailed familial information is available. The presence of autoantibodies to proteins at 37kDa and 73kDa occurred significantly more often in AU mothers when compared with two distinct control populations. Bands that were shown to be different between autism and controls were seen in more than a quarter of mothers who had children with autism. The fact that these bands were not found in all mothers of children with autism further emphasizes the heterogeneity that is widely reported in autism, and the variety of etiologic mechanisms that likely exist (Hertz-Picciotto et al., 2006
). Furthermore, while none of the individual bands were noted exclusively in mothers of children with AU, the simultaneous presence of the 37kDa and 73kDa protein bands was unique to the AU group.
Previous studies have also suggested a role for maternal antibodies in the etiology of some cases of autism (Dalton et al., 2003
). Moreover, Zimmerman et al. (2006)
recently reported differing patterns of serum immunoreactivity to pre-natal rat brain between mothers of children with autism and mothers of control children. Furthermore, the authors demonstrated that immunoreactivity persisted in maternal circulation for up to 18 years post-delivery (Zimmerman et al., 2006
). Interestingly, the group differences in brain reactivity patterns were observed only with pre-natal rat brain protein and not post-natal (day 8) rat brain protein. The patterns described in the Zimmerman study differ from those presented in the current report, which is possibly due to disparities between rat and human brain proteins, or differences in sample processing. However, the presence of maternal antibody reactivity against neuronal protein associated with an outcome of autism in the child, is consistent across the two studies.
The transplacental passage of maternal IgG isotype antibodies has long been known as a mechanism for fetal immune instruction (Garty et al., 1994
) and protection (Harris et al., 2006
; Simister, 2003
). A recently described organelle in the placental epithelium that expresses the low affinity IgG receptor, FcγRIIb, as well as the IgG receptor and transport protein FcRn, appears to provide a dedicated transport mechanism for maternal IgG to enter fetal circulation (Mishima et al., 2006
). Detectable levels of maternal IgG are present in fetal circulation as early as 18 weeks gestation, and by 38 weeks gestation, fetal levels are comparable with maternal levels. Interestingly, neonatal IgG, which is overwhelmingly maternal in origin, is seen at levels exceeding the maternal concentration at delivery and persists at detectable levels up to 6 months post-delivery (Garty et al., 1994
). Further, the presence of IgG heavy- and light-chain bands in the fetal brain immunoblots supports the transport of maternal IgG into the fetal brain during gestation. Thus the window of exposure to maternal IgG coincides substantially with critical periods of early neurodevelopment.
Despite the beneficial nature of the majority of maternal IgG received by the fetus, a number of neonatal autoimmune diseases have been demonstrated to result from pathogenic maternal IgG. Notably, the presence of maternal anti-Ro/SS-A and anti-La/SS-B antibodies cause neonatal lupus syndrome, often leading to congenital heart block (Tincani et al., 2006
) In addition, cases of neonatal anti-phospholipid syndrome (APS), mediated through maternal autoantibodies, have been observed in the newborn infants of mothers with primary APS (Soares Rolim et al., 2006
). Finally, abnormal thyroid function is often noted in infants born to mother with Hashimoto's thyroiditis or Graves' disease, caused by placental transfer of maternal anti-thyroid antibodies(Fu et al., 2005
). Typically, symptoms of neonatal thyroiditis resolve as maternal antibodies are cleared from the circulation of the infant.
Our data suggest that the presence of maternal autoantibodies to fetal brain proteins of approximately 37kDa and 73kDa molecular weight confers an elevated risk for autism. We performed western blots using several other protein sources in order to determine the tissue specificity of these autoantibodies. Interestingly, the reactivity observed towards fetal brain protein was not seen among kidney, duodenum or adult brain proteins. This finding suggests the possibility that the autoreactivity described herein is targeted towards proteins expressed exclusively, or at substantially higher levels, during fetal development.
Maternal plasma collection for the current retrospective study occurred on average 3.5 years after the birth of the affected child, and approximately 18 months after a diagnosis of autism. As circulating antibody titers are known to vary over time based on the immunological state of the individual (Toptygina et al., 2005
), the maternal antibody profile observed at the time of the registration of her child into the CHARGE study may be slightly different than during gestation. However, it has been demonstrated that antibodies, such as those generated in response to vaccination, can persist for many years due to the maintenance and subsequent polyclonal reactivation of memory B cells (Shinefield et al., 2002
). It is currently unknown whether or not successive children from those mothers with reactivity to fetal brain will have autism. A longitudinal analysis of subsequent offspring from these control mothers will allow us to resolve this issue.
Increasing attention has been given to the notion that autism, as a spectrum of disorders, likely encompasses numerous, etiologically distinct behavioral phenotypes. Our observation of maternal reactivity to two protein bands at 37kDa and 73kDa more frequently in mothers of AU children exhibiting behavioral regression than in those with early onset AU may help to elucidate biologic mechanisms contributing to phenotypic variance in ASD. Assuming that the observed maternal autoantibody reactivity was also present during the prenatal and/or early postnatal period, the association of autoantibodies to neural antigens with delayed onset autism appears paradoxical. While beyond the scope of the present study, this could be explained by a pathogenic mechanism involving the interference of maternal autoantibodies with neurodevelopmental pathways for which compensatory mechanisms exist, but are ultimately overwhelmed, leading to disease symptoms. Such a pathogenesis is noted in Rett syndrome, where mutations in the gene Mecp2
manifest in behavioral regression around 18 months of age (Williamson and Christodoulou, 2006
). Finally, it is important to note that the presence of maternal autoantibodies to both the 37 kDa and 73 kDa proteins does not provide an etiologic mechanism for all cases of regressive autism, and their presence is strongly associated with the regressive phenotype only in a sub-population of individuals.
These data provide evidence for an association between the presence of maternal immune system biomarkers and a diagnosis of autism in a subset of children. The presence of specific anti-fetal brain antibodies in the circulation of mothers during pregnancy may be a potential trigger that, when paired with genetic susceptibility, is sufficient to induce a downstream effect on neurodevelopment leading to autism. At present, we are investigating maternal plasma reactivity against fetal brain in a prospective cohort to determine the effect of the gestational autoantibody profile as it relates to an outcome of autism. Furthermore, work is currently under way to determine the protein targets of these antibodies, the identification of which will allow us to better understand potential pathogenic mechanisms as well as create specific screening assays.