Fronto-insular (FI) cortex is part of the neural system involved in self-awareness and social reciprocity and contains von Economo neurons (VENs) that selectively degenerate in fronto-temporal dementia (FTD) and may also be involved in autism (Seeley, 2006; Allman, 2005). Deficits in self-awareness and social reciprocity are characteristic of both conditions. The activity of FI is reduced in autistic subjects relative to controls when they introspect their feelings (Silani, 2008).
Objectives:
Determine the networks of genes that are abnormally expressed in FI of autistic brains compared to controls. Quantify the number of quiescent, activated, and phagocytic microglia in autistic brains compared to age matched controls.
Methods:
We used RNA-Seq to quantify expression across the entire set of genes in FI obtained from well phenotyped autistic cases and controls, then determined which genes are most informative in discriminating the different groups using an information theoretic approach. The most informative genes were subjected to IPA and GO analysis. We used IBA1 immunocytochemistry and stereology to quantify the number of microglia in the brain.
Results:
Autism brains classified as group A display a greater number of activated microglia, the key cellular participants in the inflammatory response in the brain, compared to autism group B brains and controls. Including additional autistic and control subjects for microglia staining (Iba1) and classification we found autistic (n=11) subjects compared to controls (n=9) have significantly more activated and phagocytic microglia while the control subjects have significantly more quiescent microglia. An IPA analysis based on the 100 most informative genes revealed that autism-A brains exhibit a network of potentially interacting genes related to immune function and inflammation. IL-6 is the major hub in this network. The proteins for several of the genes in this network, including IL-6 receptor and ATF3, are preferentially immuno-stained in VENs. Previous studies have found significantly increased amounts of IL-6 protein in autistic subjects relative to controls in frontal cortex (Li, 2009) and anterior cingulate cortex (Vargas, 2005). There are many studies implicating IL-6 in social functioning (Ader, 2007). For the 1000 genes with the highest NMI values, we performed a GO analysis comparing autism-A, -B and controls, which revealed a set of significantly enriched GO terms related to stress, inflammation and apoptosis, reflecting the network of genes centered on IL-6 in autism-A. The comparison of the three groups also reveals a subset of genes related to synaptic vesicles and neurotransmitter release that is enriched in the autism-B brains.
Conclusions:
Autism-A exhibits a gene network involved in inflammation, and Autism-A subjects have a greater number of activated and phagocytic microglia compared to controls. We counted the microglia in an additional group of autistic subjects for which we did not have RNA-Seq data, and all of these subjects had increased numbers of activated and phagocytic microglia compared to controls. Autism-B has increased expression of genes which are involved in the presynaptic active zone where a change in vesicle fusion can alter neurotransmitter release and may perturb cortical function.