Objective Epigenetic mechanisms, including DNA methylation (DNAm), have been proposed
to play a key role in Crohn's disease (CD) pathogenesis. However, the specific cell
types and pathways affected as well as their potential impact on disease phenotype
and outcome remain unknown. We set out to investigate the role of intestinal epithelial
DNAm in CD pathogenesis. Design We generated 312 intestinal epithelial organoids (IEOs)
from mucosal biopsies of 168 patients with CD (n=72), UC (n=23) and healthy controls
(n=73). We performed genome-wide molecular profiling including DNAm, bulk as well
as single-cell RNA sequencing. Organoids were subjected to gene editing and the functional
consequences of DNAm changes evaluated using an organoid-lymphocyte coculture and
a nucleotide-binding oligomerisation domain, leucine-rich repeat and CARD domain containing
5 (NLRC5) dextran sulphate sodium (DSS) colitis knock-out mouse model. Results We
identified highly stable, CD-associated loss of DNAm at major histocompatibility complex
(MHC) class 1 loci including NLRC5 and cognate gene upregulation. Single-cell RNA
sequencing of primary mucosal tissue and IEOs confirmed the role of NLRC5 as transcriptional
transactivator in the intestinal epithelium. Increased mucosal MHC-I and NLRC5 expression
in adult and paediatric patients with CD was validated in additional cohorts and the
functional role of MHC-I highlighted by demonstrating a relative protection from DSS-mediated
mucosal inflammation in NLRC5-deficient mice. MHC-I DNAm in IEOs showed a significant
correlation with CD disease phenotype and outcomes. Application of machine learning
approaches enabled the development of a disease prognostic epigenetic molecular signature.
Conclusions Our study has identified epigenetically regulated intestinal epithelial
MHC-I as a novel mechanism in CD pathogenesis.
