The aims of this thesis were firstly to investigate gene expression profiles in human
colonic and terminal ileal biopsies using microarray technology in a well phenotyped
cohort of patients with Crohn's disease, ulcerative colitis and a control cohort. The
role in disease pathogenesis of differentially expressed genes was investigated along
with the expression of candidate genes identified by genome wide association study
and cell lineage analysis. Parallel studies attempted to replicate the Nature Genetics
publications of Peltekova and Stoll and colleagues who investigated the role the IBD5
locus and the DLG5 gene respectively in patients with inflammatory bowel disease.
In the healthy adult colon cluster analysis showed differences in gene expression
between the right and left colon. (x²=25.1, p<0.0001). Developmental genes
HOXA13, (p=2.3x10⁻¹⁶, HOXB13 (p <1xl0⁻⁴⁵), GL11 (p=4.0xl0⁻²⁴), and GLI3
(p=2.1x10⁻²⁸) primarily drove this separation.
Upregulated genes in the Crohn's disease biopsies compared to the controls included
SAA1 (Fold change (FC) +7.5, p=1.47xl0⁻⁴¹) and REGL (FC +7.3, p=2.3xl0⁻¹⁶).
Cellular detoxification genes including-SLC14A2 (FC -2.49, p= 0.00002) were
downregulated. In the Crohn's disease terminal ileal biopsies diubiquitin (FC+11.3,
p<1x10⁻⁴⁵), MMP3 (FC +7.4, p=1.3x10⁻¹¹) and IRTA1 (FC -11.4, p= 4.7xl0⁻¹²) were
differentially expressed compared to controls. In the colon SAA1 (FC +6.3, p=
5.3xl0⁻⁸) was upregulated and TSLP (FC -2.3, p= 2.7xl0⁻⁶) was downregulated
comparing non-inflamed Crohn's disease and control biopsies.
Of the Crohn's disease susceptibility genes identified by genome wide association
scan IL-23A, JAK2 and STAT3 were upregulated in Crohn's disease confirming
dysregulation of Thl7 signalling. Modest differential expression was also observed in
a number of the autophagy genes, notably ATG16L1. When clustering analysis was
undertaken, terminal ileal Crohn's disease and terminal ileal control biopsies
separated from colonic Crohn's disease and colonic control biopsies. Further
clustering analysis of the terminal ileal biopsies showed separation between the
terminal ileal Crohn's disease and control biopsies.
When the ulcerative colitis biopsies and control biopsies were compared,
differentially upregulated genes in ulcerative colitis included SAA1 (p<10⁻⁴⁵) the
alpha defensins, DEFA5&6 (p=0.00003 and p=6.95xl0⁻⁷ respectively), MMP3
(p=5.6x10⁻¹⁰) and MMP7 (p=2.3x10⁻⁷). Increased DEFA5&6 expression was further
characterized to Paneth cell metaplasia by immunohistochemistry and in-situ
hybridization.
Variants in all the examined IBD5 SNPs were associated with Crohn's disease
(p<0.003). The IBD5 locus was also associated more severe Crohn's disease
behaviour. In the absence of the IBD5 risk haplotype, no association of OCTN1/2
variants with Crohn's disease was detected. The analysis of the DLG5 variant 113A
showed there were no associations with inflammatory bowel disease, Crohn's disease
or ulcerative colitis.
Overall these data emphasise the key role of a number of inflammatory molecules and
pathways in pathogenesis of Crohn's disease and ulcerative colitis, and their potential
for translation to therapeutic targets. The results also add considerably to the recent
genome wide association studies in providing complimentary human colonic and ileal
expression data along with detailed analysis of the IL-23 and autophagy pathways.
