A novel in vivo model of permanent intestinal aganglionosis

Abstract

BackgroundEnteric neuromuscular disease is a characteristic of several disease states, including Hirschsprung disease, esophageal achalasia, Chagas disease, and gastroparesis. Medical therapy for these conditions is limited, and surgical intervention may incur significant morbidity. Alternatively, transplantation of neural progenitor cells may regenerate enteric ganglia. Existing aganglionosis model systems are limited by swift animal demise or by spontaneous regeneration of native ganglia. We propose a novel protocol to induce permanent aganglionosis in a segment of rat jejunum, which may serve as an experimental transplantation target for cellular therapy. Materials and methodsThis protocol was performed in 17 adult female Sprague–Dawley rats. A laparotomy was performed and a 1-cm segment of jejunum was isolated from continuity. Among 14 rats, the isolated segments were treated with benzalkonium chloride (BAC) for 20 min to induce aganglionosis. Jejunal segment isolation was performed without BAC treatment in three rats. The animals were euthanized at posttreatment days 21–166. Muscle layer diameter was compared among normal, isolated, and BAC-treated isolated jejunal segments. The presence of jejunal ganglia was documented by immunohistochemical staining (IHC) for beta-III tubulin (TUJ1) and S100, markers of neuronal and glial cell lineages, respectively. ResultsGanglia were identified by IHC in normal and isolated jejunal segments. Isolated segments had significantly hypertrophied smooth muscle layers compared with normal jejunum (diameter 343 ± 53 μm versus 211 ± 37 μm, P < 0.0001). BAC-treated jejunal segments had no IHC evidence of ganglionic structures. Aganglionosis was persistent in all specimens up to 166 days after treatment. ConclusionsThe exclusion of a jejunal segment from continuity and concurrent treatment with BAC results in an effective, reproducible, and permanent model of aganglionosis. Muscular hypertrophy and aganglionosis in the isolated jejunal segment make it an ideal recipient site for transplantation of neuroglial precursor cells.