Immune transcriptome alterations in the temporal cortex of subjects with autism

Abstract

Autism is a severe disorder that involves both genetic and environmental factors. Expression profiling of the superior temporal gyrus of six autistic subjects and matched controls revealed increased transcript levels of many immune system-related genes. We also noticed changes in transcripts related to cell communication, differentiation, cell cycle regulation and chaperone systems. Critical expression changes were confirmed by qPCR (BCL6, CHI3L1, CYR61, IFI16, IFITM3, MAP2K3, PTDSR, RFX4, SPP1, RELN, NOTCH2, RIT1, SFN, GADD45B, HSPA6, HSPB8 and SERPINH1). Overall, these expression patterns appear to be more associated with the late recovery phase of autoimmune brain disorders, than with the innate immune response characteristic of neurodegenerative diseases. Moreover, a variance-based analysis revealed much greater transcript variability in brains from autistic subjects compared to the control group, suggesting that these genes may represent autism susceptibility genes and should be assessed in follow-up genetic studies.

Figure 1. Two-way hierarchical clustering of expression data

Hierarchical clustering was performed on log2-transformed expression level of 152 differentially expressed genes which demonstrated |ALR|>1 and p<0.05 in three distinct statistical analyses (groupwise, pairwise and rank-based). Samples were clustered vertically, gene probes were clustered horizontally. Genes are denoted by Affymetrix probes and NCBI gene symbols. Each colored square represents a normalized gene expression level, color coded for increase (red) or decrease (blue). Color intensity is proportional to magnitude of change. The clustering resulted in a near-perfect separation of samples into two discrete groups corresponding to diagnosis (vertical dendrogram, red for AUT and blue for CONT).

Figure 2. Data validation by qPCR

Differential expression of 22 genes was validated by qPCR. X-axis represents -ΔΔCt measured by qPCR, Y-axis denotes DNA microarray reported ALR. Each symbol represents a single gene. Note that the qPCR and microarray data were highly correlated (r=0.89; p<0.001), with agreement in the directionality of change for all investigated transcripts.

Figure 3. Autistic samples show altered expression of transcripts involved in cell communication and differentiation

Differentially expressed genes were functionally classified based on literature search (for references, see Supplemental Material 2). We observed a strong overrepresentation of differentially expressed transcripts mediating cell communication and motility, cell fate and differentiation, and chaperones. The magnitude of the gene expression change is coded by cell shading, and checkmarks denote successful qPCR validation of differential expression. Note that the transcript inductions (grey boxes) greatly outnumbered transcript repressions (white boxes).