Identification of novel genes in Hirschsprung disease pathway using whole genome expression study

This study aims to identify new genes not described previously that may be relevant in the etiology or pathophysiology of patients with Hirschsprung disease (HD). This was done by identifying differences in gene expression between normal and abnormal segments of bowel in HD patients compared with controls.

Full-thickness colonic tissue samples were taken from HD patients, both from the diseased (Ds) and normal segment of the colon (Nr), and from controls (Ct). Samples were further dissected into mucosa (MUC) and muscle (MUS). RNA was extracted and analyzed on Affymetrix Gene Chip Human Gene 1.0 ST arrays. Statistical analyses using ANOVA with a fold change cut off of 2 was applied to detect a number of differentially expressed genes. Selected genes were revalidated by quantitative real-time reverse transcriptase polymerase chain reaction.

Thirty-four samples (18 MUS and 16 MUC) were analyzed. MUC (1.64 ± 0.46 μg/mg) and MUS (0.83 ± 0.48 μg/mg) showed good RNA extraction yield and quality. Of the 24,987 filtered on expression genes, MUS showed 220 genes with expression difference of 2-fold, out of which 120 genes were significant with P ≤ .05. Similarly, MUC demonstrated 206 genes with 2-fold changes and 9 had P ≤ .05. Some genes showing differential expression between groups and therefore subject to further analysis were RELN, GAL, GAP43, NRSN1, and GABRG2.

Analyzed data showed significant differences in expression of above sets of genes with up- and down-regulation, which has not been described before in HD and could have a role in pathogenesis of this condition.