Enteric Neuron Imbalance and Proximal Dysmotility in Ganglionated Intestine of the Sox10Dom/+ Hirschsprung Mouse Model

Abstract

Background & Aims In Hirschsprung disease (HSCR), neural crest-derived progenitors (NCPs) fail to completely colonize the intestine so that the enteric nervous system (ENS) is absent from distal bowel. Despite removal of the aganglionic region, many HSCR patients suffer from residual intestinal dysmotility. To test the hypothesis that inappropriate lineage segregation of NCPs in proximal ganglionated regions of the bowel could contribute to such postoperative disease, we investigated neural crest (NC)-derived lineages and motility in ganglionated, postnatal intestine of the Sox10Dom/+ HSCR mouse model. Methods Cre-mediated fate-mapping was applied to evaluate relative proportions of NC-derived cell types. Motility assays were performed to assess gastric emptying and small intestine motility while colonic inflammation was assessed by histopathology for Sox10Dom/+ mutants relative to wildtype controls. Results Sox10Dom/+ mice showed regional alterations in neuron and glia proportions as well as Calretinin+ and nNOS+ neuronal subtypes. In the colon, imbalance of enteric NC derivatives correlated with the extent of aganglionosis. All Sox10Dom/+ mice exhibited reduced small intestinal transit at 4-weeks of age, and at 6-weeks, Sox10Dom/+ males had increased gastric emptying rates. Sox10Dom/+ mice surviving to 6-weeks of age had little or no colonic inflammation when compared to wildtype littermates, suggesting that these changes in GI motility are neurally mediated. Conclusions The Sox10Dom mutation disrupts the balance of NC-derived lineages and affects GI motility in the proximal, ganglionated intestine of adult animals. This is the first report identifying alterations in enteric neuronal classes in Sox10Dom/+ mutants, which suggests a previously unrecognized role for Sox10 in neuronal subtype specification.

Bowel dysfunction following pullthrough surgery is associated with an overabundance of nitrergic neurons in Hirschsprung disease

“…Our study excluded patients with total colonic aganglionosis, but a similar trend was seen toward a higher incidence of constipation in patients with a longer length of aganglionosis. Studies in mice have suggested that the extent of neuronal subtype imbalance also correlates with the length of aganglionosis [8]; however, we found no correlation between the proportion of NOS neurons and the length of bowel resected. In addition, studies have shown that disturbances in NOS expression are greater in older HD mice [9], but we found no similar correlation between NOS and age in our patients.…”

“…NOS+ neurons are one of the first subtypes to appear in embryonic mouse colon [20], and failure of neuronal maturation to progress normally may result in an overabundance of NOS+ neurons. In previous studies in mouse models of HD, neuronal subtype imbalance was found to be limited to the mid and distal colon and was normal in proximal colon and small bowel [8, 9]. The extent of neuronal abnormality in HD patients is not yet known and warrants further study, as this knowledge is essential to determine the appropriate length of resection and to predict functional outcome after pullthrough.…”

“…The overexpression of nitric oxide synthase (NOS) and its related proteins is unique to HD and are not a result of bowel dilatation alone [7]. Similar alterations in NOS expression have been observed in several mouse models of HD and are linked to intestinal dysmotility [8-10]. Intestinal motility relies on a careful balance between inhibitory (e.g.…”

Mouse models of Hirschsprung disease and other developmental disorders of the enteric nervous system: Old and new players

“…Previous studies demonstrated that hedgehog signaling is required for the normal migration of enteric neural progenitors and glial cell generation through Sox10 regulation (Liu et al, 2015; Ngan et al, 2011). In Sufu mutants, the networks of enteric ganglia were disorganized and neuron:glial cell ratios were reduced, a defect reminiscent of that recently described in Sox10 and Raldh1, -2, and -3 mutants (Musser et al, 2015; Wright-Jin et al, 2013). Because mice lacking Sufu in neural crest cells die before the gut is completely colonized by ENCCs, the authors were however not able to determine whether loss of Sufu results in an absence of neurons in the distal bowel and thus a HSCR-like phenotype.…”

“…While transcription factors that participate with Sox10 auto-regulation have been identified (Wahlbuhl et al, 2012), cofactors that participate with Sox10 in regulation of Ednrb are not yet known. Analysis of Sox10 Dom mouse mutants revealed that alterations in this transcription factor not only lead to distal aganglionosis, but can also disrupt normal proportions of neuron subtypes in ganglionated small intestine that lead to deficits in intestinal transit (Musser et al, 2015). Similarly, Zeb2 plays essential roles in enteric neurogenesis by promoting ENCC proliferation and early migration within the fetal bowel (Stanchina et al, 2010; Van de Putte et al, 2003) via interactions with Sox10.…”