Objectives: The objective of this study was to survey host gene expression and microflora in intestinal biopsies from children with autistic disorder and gastrointestinal complaints (AUT-GI) vs children with gastrointestinal complaints alone (Control-GI).
Methods: Transcription profiling was pursued by cDNA microarray using RNA extracted from ileal biopsies (4 per patient) of 15 male AUT-GI and 7 age-matched, male Control-GI patients. Pathway analysis was performed using Ingenuity Pathway Analysis and GO Ontology. Changes in gene expression were confirmed by quantitative real-time PCR. Intestinal microbiota were investigated in ileal and cecal biopsies from AUT-GI and Control-GI children using amplicon-based, bar-coded pyrosequencing of the V2 region of bacterial 16S rRNA gene. Taxonomic classification of 525,519 bacterial sequences was accomplished using the Ribosomal Database Project classifier tool. Differences in microbiota between the two groups were further evaluated and confirmed using Bacteroidete-, Firmicute-, and Sutterella-specific real-time PCR.
Results: Microarray and pathway analysis revealed significant changes in genes involved in carbohydrate metabolism and transport and inflammation in ileal biopsies from AUT-GI as compared to Control-GI subjects. Real-time PCR confirmed significant decreases in the AUT-GI group in the primary brush border disaccharidases, sucrase isomaltase (p=0.0013), maltase glucoamylase (p=0.0027), and lactase (p=0.0316) as well as in two enterocyte hexose transporters, sodium glucose co-transporter 1 (p=0.0082) and glucose transporter 2 (p=0.0101). In contrast, increases were confirmed for inflammation-related genes in AUT-GI subjects: complement component 1, q subcomponent, A chain (p=0.0022), resistin (p=0.0316), CD163 (p=0.0150), tumor necrosis factor-like weak inducer of apoptosis (p=0.015), and interleukin 17F (p=0.0220). No significant group differences were observed for the enterocyte-specific marker, villin. In conjunction with changes in intestinal gene expression, bacterial content differed between the AUT-GI and Control-GI groups: pyrosequencing and real-time PCR revealed lower levels of Bacteroidetes (ileum: 50% reduction, p=0.0027; cecum: 25% reduction, p=0.0220, and higher Firmicute/Bacteroidete ratios in AUT-GI children (ileum: p=0.0006; cecum: p=0.0220). High levels of Sutterella species were found in 47% of AUT-GI biopsies (7/15), whereas Sutterella was not detected in any Control-GI biopsies (0/7; ileum: p = 0.0220; cecum: p = 0.0368).
Conclusions: We describe a distinctive syndrome in autistic children wherein gastrointestinal dysfunction is associated with altered gene expression reflecting intestinal inflammation, impaired carbohydrate metabolism and transport, and dysbiosis. These findings may provide insights into pathogenesis and enable new strategies for therapeutic intervention.