Three-Dimensional Imaging of the Enteric Nervous System in Human Pediatric Colon Reveals New Features of Hirschsprung’s Disease

作者：Joshua D. Eisenberg, Rebecca P Bradley, Kahleb D. Graham, Rachel H. Ceron, Amanda M Lemke, Benjamin J. Wilkins, Ali Hadi Naji, Robert O. Heuckeroth · 发表于：Gastroenterology · 年份：2024 · DOI：10.1053/j.gastro.2024.02.045 · 被引用次数：27 · 研究领域：Congenital gastrointestinal and neural anomalies、Gastrointestinal motility and disorders、Intestinal Malrotation and Obstruction Disorders

Background & Aims Hirschsprung's disease is defined by the absence of the enteric nervous system (ENS) from the distal bowel. Primary treatment is "pull-through" surgery to remove bowel that lacks ENS, with reanastomosis of "normal" bowel near the anal verge. Problems after pull-through are common, and some may be due to retained hypoganglionic bowel (ie, low ENS density). Testing this hypothesis has been difficult because counting enteric neurons in tissue sections is unreliable, even for experts. Tissue clearing and 3-dimensional imaging provide better data about ENS structure than sectioning. Methods Regions from 11 human colons and 1 ileal specimen resected during Hirschsprung's disease pull-through surgery were cleared, stained with antibodies to visualize the ENS, and imaged by confocal microscopy. Control distal colon from people with no known bowel problems were similarly cleared, stained, and imaged. Results Quantitative analyses of human colon, ranging from 3 days to 60 years old, suggest age-dependent changes in the myenteric plexus area, ENS ganglion area, percentage of myenteric plexus occupied by ganglia, neurons/mm 2 , and neuron Feret's diameter. Neuron counting using 3-dimensional images was highly reproducible. High ENS density in neonatal colon allowed reliable neuron counts using 500-μm 2 × 500-μm 2 regions (36-fold smaller than in adults). Hirschsprung's samples varied 8-fold in proximal margin enteric neuron density and had diverse ENS architecture in re...