ABSTRACT: The objectives of this study were to measure effects of an aspirin intervention on gene expression in normal colonic epithelial and stromal tissue in healthy humans and to determine whether response differed by UGT1A6*2 genotype. We also sought to characterize gene expression differences within colonic tissue microenvironments by identifying genes that were differentially expressed between epithelial and stromal tissue. No statistically significant differences in gene expression were observed in response to aspirin or UGT1A6 genotype, but tissue PGE2 levels were lower with aspirin compared to placebo (p <0.001). Transcripts differentially expressed between epithelium and stroma (N = 4916, P <0.01, false discovery rate <0.001), included a high proportion of genes involved in cell signaling, cellular movement, and cancer. Genes preferentially expressed in epithelium were involved in drug and xenobiotic metabolism, fatty acid and lipid metabolism, apoptosis signaling, and ion transport. Genes preferentially expressed in stroma included those involved in inflammation, cellular adhesion, and extracellular matrix production. Wnt-Tcf4 pathway genes were expressed in both epithelium and stroma but differed by subcellular location. Our results suggest that, in healthy individuals, subtle effects of aspirin on gene expression in normal colon tissue are likely overwhelmed by inter-individual variability in microarray analyses. Differential expression of critical genes between colonic epithelium and stroma suggest that these tissue types need to be considered separately. In a randomized, cross-over, placebo-controlled trial of 44 healthy men and women, homozygous for UGT1A6*1 or UGT1A6*2, we explored differences between global epithelial and stromal expression, using Affymetrix U133 + 2.0 microarrays and tested effects of 60-day aspirin supplementation (325 mg/d) on epithelial and stromal gene expression and colon prostaglandin E2 (PGE2) levels Participants were randomly assigned, blocked on sex and genotype, to the order in which they received treatment A or treatment B. (Note: The type of treatment i.e., pill type active or placebo is masked. Please contact the study authors directly if you would like access to this information). The washout period between intervention periods was three months. A sigmoidoscopy was performed at least 60 days after day 1 of each intervention period. Therefore, two biopsies were obtained from each participant and then processed, where by, epithelial and stromal fractions were separated and analyzed using Affymetrix U133+2.0 microarrays. This resulted in obtaining total RNA from 88 epithelial and and 88 matched stromal samples. A total of 176 gene arrays were measured. Participants were labeled consecutively. There are 4 samples from each person designated A1-A4 (participant 1); A5-A6 (participant 2); B1-B4 (participant 3) etc. to W1-W4 (participant 44). Within each person, odd numbers (1 & 3) are epithelial tissue and even numbers (2 & 4) are stromal tissue. The epithelial and stroma from each intervention period are paired together (e.g., A1 & A2 are from the first treatment Participant 1 received; A3 & A4 are from the second treatment Participant 1 received).
