Low serum myeloperoxidase in autistic children with gastrointestinal disease

Anthony J Russo1, Arthur Krigsman2, Bryan Jepson2, Andy Wakefield21Research Director, Health Research Institute/Pfeiffer Treatment Center, Warrenville, IL, USA; 2Thoughtful House Center for Children, Austin, TX, USAAim: To assess serum myeloperoxidase (MPO) levels in autistic children with severe gastrointestinal (GI) disease and to test the hypothesis that there is an association between serum MPO concentration and inflammatory GI disease, including antineutrophil cytoplasmic antibodies (ANCA), previously seen in a subgroup of autistic children.Subjects and methods: Serum from 40 autistic children with chronic digestive disease (most with ileo-colonic lymphoid nodular hyperplasia (LNH) and inflammation of the colorectum, small bowel and/or stomach), and 48 controls (12 age-matched autistic children with no GI disease, 20 age-matched children without autism or GI disease, and 16 nonautistic individuals with no family history of autism) were tested using enzyme-linked immunosorbent assays designed to quantitate serum MPO levels. MPO serum concentration of autistic children with GI disease was compared to GI disease severity (including LNH and erythema) and presence of ANCA.Results: We found that a significant number of autistic children with chronic digestive disease had low serum levels of MPO. However, there was no significant relationship between these levels and severity of GI disease, including the presence of ANCA.Discussion: These results suggest a relationship between low MPO levels and GI disease seen in a subpopulation of autism spectrum disorders individuals. MPO concentration may therefore be a useful biomarker for GI disease in this group of autistic children.Keywords: autism spectrum disorders, autism, myeloperoxidase, GI disease, oxidative stres

A molecular biomarker for prediction of clinical outcome in children with ASD, constipation, and intestinal inflammation

Abstract In children with autism spectrum disorder (ASD) who present to the gastroenterologist with chronic constipation on a background of colonic inflammation, we have identified two distinct clinical subtypes: (1) patients who experience a sustained state of GI symptomatic remission while on maintenance anti-inflammatory therapy (fast responders) and, (2) those with recurrent right-sided fecal loading requiring regular colon cleanouts during treatment for enterocolitis (slow responders). We hypothesized that a detailed molecular analysis of tissue from the affected region of the colon would provide mechanistic insights regarding the fast versus slow response to anti-inflammatory therapy. To test this, ascending colon biopsy tissues from 35 children with ASD (20 slow responders and 15 fast responders) were analyzed by RNAseq. Hierarchical cluster analysis was performed to assign samples to clusters and gene expression analysis was performed to identify differentially expressed transcripts (DETs) between samples within the clusters. Significant differences were found between the two clusters with fast responder-predominant cluster showing an upregulation of transcripts involved in the activation of immune and inflammatory response and the slow responder-predominant cluster showing significant over-representation of pathways impacting colonic motility (e.g. genes involved in tryptophan and serotonin degradation and mitochondrial dysfunction). Regression analysis identified a single long non-coding RNA that could predict cluster assignment with a high specificity (0.88), sensitivity (0.89) and accuracy (0.89). Comparison of gene expression profiles in the ascending colon from a subset of patients with ASD, chronic right-sided fecal loading constipation and a slow versus fast response to therapy has identified molecular mechanisms that likely contribute to this differential response following the primary therapeutic intervention (i.e. treatment for colonic inflammation with brief induction immunosuppression followed by maintenance non-steroidal anti-inflammatory therapy). Importantly, we have identified a transcript that, if validated, may provide a biomarker that can predict from the outset which patients will be slow responders who would benefit from an alternate therapeutic strategy in treating their constipation

Identification of Unique Gene Expression Profile in Children with Regressive Autism Spectrum Disorder (ASD) and Ileocolitis

<div><p>Gastrointestinal symptoms are common in children with autism spectrum disorder (ASD) and are often associated with mucosal inflammatory infiltrates of the small and large intestine. Although distinct histologic and immunohistochemical properties of this inflammatory infiltrate have been previously described in this ASD^GI group, molecular characterization of these lesions has not been reported. In this study we utilize transcriptome profiling of gastrointestinal mucosal biopsy tissue from ASD^GI children and three non-ASD control groups (Crohn's disease, ulcerative colitis, and histologically normal) in an effort to determine if there is a gene expression profile unique to the ASD^GI group. Comparison of differentially expressed transcripts between the groups demonstrated that non-pathologic (normal) tissue segregated almost completely from inflamed tissue in all cases. Gene expression profiles in intestinal biopsy tissue from patients with Crohn's disease, ulcerative colitis, and ASD^GI, while having significant overlap with each other, also showed distinctive features for each group. Taken together, these results demonstrate that ASD^GI children have a gastrointestinal mucosal molecular profile that overlaps significantly with known inflammatory bowel disease (IBD), yet has distinctive features that further supports the presence of an ASD-associated IBD variant, or, alternatively, a prodromal phase of typical inflammatory bowel disease. Although we report qPCR confirmation of representative differentially expressed transcripts determined initially by microarray, these findings may be considered preliminary to the extent that they require further confirmation in a validation cohort.</p> </div

Comparison of microarray results with qPCR results from 12 representative transcripts differentially-regulated in both terminal ileum and colon, exclusively in ASD^GI samples.

<p>Comparison of microarray results with qPCR results from 12 representative transcripts differentially-regulated in both terminal ileum and colon, exclusively in ASD^GI samples.</p

Characteristics of Study Population.

<p>Characteristics of Study Population.</p

Summary for IPA Diseases and Disorders Category.

<p>Summary for IPA Diseases and Disorders Category.</p

Summary for IPA Canonical Pathways Category.

<p>Summary for IPA Canonical Pathways Category.</p

Gastrointestinal Symptoms in ASD Study Population.

<p>Gastrointestinal Symptoms in ASD Study Population.</p

Overlapping unique ASD^GI gene expression from TI and colon.

<p>Pair-wise comparisons were performed between each of the disease groups (ASD^GI, CD and UC) and the control (non-histopathologic tissue) samples. <b>A.</b> There were 1409 unique DETs (differentially-expressed transcripts) in the ASD^GI versus control comparison in TI mucosa. <b>B</b>. There were 1189 unique DETs in the ASD^GI versus control comparison in colonic mucosa. <b>C.</b> The overlap between those two lists is displayed in this Venn diagram. There are a total of 178 DETs shared in ASD^GI tissues (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0058058#pone.0058058.s004" target="_blank">Table <b>S4</b></a>). This list of 178 DETs was imported into Ingenuity Pathway Analysis software for further analysis.</p

Principal Component Analysis (PCA) scatterplot representing 53 individual microarray datasets from terminal ileum tissues.

<p>Each circle represents the cumulative gene expression profile for an individual sample. Samples with similar profiles cluster together in the three-dimensional space.</p