Keck_2021_Cell.Mol.Gastroenterol.Hepatol__

BACKGROUND AND AIMS: Hirschsprung's disease (HSCR) is a congenital intestinal motility disorder defined by the absence of enteric neuronal cells (ganglia) in the distal gut. The development of HSCR-associated enterocolitis remains a life-threatening complication. Absence of enteric ganglia implicates innervation of acetylcholine-secreting (cholinergic) nerve fibers. Cholinergic signals have been reported to control excessive inflammation, but the impact on HSCR-associated enterocolitis is unknown. METHODS: We enrolled 44 HSCR patients in a prospective multicenter study and grouped them according to their degree of colonic mucosal acetylcholinesterase-positive innervation into fiber-low and fiber-high patient groups. The fiber phenotype was correlated with the tissue cytokine profile as well as immune cell frequencies using Luminex analysis and fluorescence-activated cell sorting (FACS) analysis of colonic tissue and immune cells. Using confocal immunofluorescence microscopy, macrophages were identified in close proximity to nerve fibers and characterized by RNA-seq analysis. Microbial dysbiosis was analyzed in colonic tissue using 16S-rDNA gene sequencing. Finally, the fiber phenotype was correlated with postoperative enterocolitis manifestation. RESULTS: The presence of mucosal nerve fiber innervation correlated with reduced T-helper-17 (Th17) cytokines and cell frequencies. In fiber-high tissue, macrophages colocalized with nerve fibers and expressed significantly less interleukin-23 than macrophages from fiber-low tissue. HSCR patients lacking mucosal nerve fibers showed microbial dysbiosis and had a higher incidence of postoperative enterocolitis. CONCLUSION: The mucosal fiber phenotype might serve as a prognostic marker for enterocolitis development in HSCR patients and may offer an approach to personalized patient care and new therapeutic options.