Abstract
Identification of the stem cell niche is crucial for understanding the factors that regulate these cells. Rodent enteric neural crest-derived stem cells have previously been isolated by flow cytometry and culture of cell suspensions from the outer smooth muscle layers or the entire gut wall from postnatal and adult animals. Such cell suspensions contain a mixture of cell types, including smooth muscle, fibroblasts and cells associated with the vasculature and extrinsic innervation. Thus these preparations may be contaminated by stem cells associated with extrinsic sensory and autonomic nerves and by other types of stem cell that reside in the gut. Here we describe a different approach, similar to that recently used for infant human gut, to obtain enteric ganglion-derived cells, with properties of neural progenitor cells, using isolated myenteric ganglia from postnatal rat ileum. Myenteric ganglia were separated from the gut wall, dispersed and resulting cell dissociates were plated in non-adherent culture conditions with EGF and FGF-2. Under these conditions neurosphere-like bodies (NLB) developed. Cells in NLB incorporated BrdU and expressed the stem cell marker nestin but not the pan-neuronal marker PGP 9.5. Upon growth factor withdrawal some BrdU-immunopositive cells assumed the morphology of neurons and expressed PGP 9.5; others were flattened and expressed the glial cell marker GFAP. This work therefore provides evidence that neural crest-derived progenitors in the postnatal rat gut are located in the myenteric plexus, and shows that these cells can be expanded and differentiated in NLB in vitro.
Highlights
Enteric neuronal precursors from the foetal gut have been studied by several authors, notably by Gershon and his co-workers, who have used immunoselection with antibodies to the low-affinity nerve growth factor receptor p75 (Chalazonitis et al 1998a; Chalazonitis et al 1998b) and to NC-1 (Pomeranz et al 1993) to isolate neural crest-derived cells from the foetal rat gut
We report that nestin mRNA is expressed in isolated enteric ganglia from postnatal rats, and that neural progenitors can be isolated from a cell suspension of isolated myenteric ganglia of postnatal rodents
In order to study the properties of myenteric ganglion cells in isolation from surrounding non-neuronal tissues, segments of plexus consisting of linked ganglia were separated by a combination of enzymatic digestion and mechanical agitation of the outer layers of the gut wall
Summary
Enteric neuronal precursors from the foetal gut have been studied by several authors, notably by Gershon and his co-workers, who have used immunoselection with antibodies to the low-affinity nerve growth factor receptor p75 (Chalazonitis et al 1998a; Chalazonitis et al 1998b) and to NC-1 (Pomeranz et al 1993) to isolate neural crest-derived cells from the foetal rat gut. Recent studies have provided evidence that a population of neural crest-derived stem cells exists after birth, in the postnatal and adult rodent (Bondurand et al 2003; Kruger et al 2002) and human (Rauch et al 2006a) (Almond et al 2007) gut. The first published method involves prospective flow cytometry of cell suspensions derived from outer smooth muscle layers (muscularis externa), in which myenteric ganglia are embedded; cells that express high levels of the low-affinity nerve growth factor receptor, p75 are selected (Kruger et al 2002). The presence of stem cells in clumps of cells obtained from partial dispersal of the entire gut wall has been inferred after such clumps were grown in adherent culture and gave rise to a range of cell types; no attempt was made to separate or enrich neural progenitors
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
