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Updated: Apr 20, 2026
Author Spotlight: Collecting Neural Stem and Progenitor Cells from Live Animals Using a Novel Brain Milking Protocol
Published on: February 9, 2024
Prospective identification and culture of rat enteric neural stem cells (ENSCs)
Tingting Gao1, Haijiao Chen2, Mei Liu2
1Department of Pediatric Surgery, Affiliated Hospital of Nantong University, Nantong, 226001, Jiangsu, China.
Insights
Researchers developed a new method to culture enteric neural stem cells (ENSCs) from fetal rat guts. This advance offers a potential source of cells for future Hirschsprung
Area of Science:
- Developmental biology
- Stem cell research
- Gastroenterology
Background:
- Hirschsprung disease (HD) is a congenital disorder caused by defective enteric nervous system development.
- Neural crest cell colonization failure in the gut leads to aganglionosis, characteristic of HD.
- Cell replacement therapy is a potential treatment for HD, but requires a reliable source of enteric neural stem cells (ENSCs).
Purpose of the Study:
- To establish a method for culturing ENSCs from DsRed transgenic fetal rat gut.
- To provide a sufficient cell supply for future Hirschsprung disease research and potential transplantation therapies.
Main Methods:
- Culturing ENSCs from embryonic day 18 (E18) DsRed transgenic rat fetuses.
- Utilizing a specific culture medium containing chicken embryo extract, bFGF, and EGF.
- Observing cell proliferation, neurosphere formation, and differentiation using microscopy and fluorescence staining.
Main Results:
- Proliferation and formation of cell clusters observed within 2 days of culture.
- Neurosphere-like structures formed by day 8.
- Differentiated cells exhibited network-like connections and tested positive for neurofilament and GFAP.
Conclusions:
- A successful method for isolating and culturing ENSCs from E18 DsRed transgenic rats was established.
- This method provides a viable approach for obtaining ample cells for future research on Hirschsprung disease transplantation.
Abstract:
Hirschprung's disease (HD), a very common congenital abnormality in children, occurs mainly due to the congenital developmental defect of the enteric nervous system. The absence of enteric ganglia from the distal gut due to deletion in gut colonization by neural crest progenitor cells may lead to HD. The capacity to identify and isolate the enteric neuronal precursor cells from developing and mature tissues would enable the development of cell replacement therapies for HD. However, a mature method to culture these cells is a challenge. The present study aimed to propose a method to culture enteric neural stem cells (ENSCs) from the DsRed transgenic fetal rat gut. The culture medium used contained 15 % chicken embryo extract, basic fibroblast growth factor, and epidermal growth factor. ENSCs were cultured from embryonic day 18 in DsRed transgenic rat. Under inverted microscope and fluorescence staining, ENSCs proliferated to form small cell clusters on the second day of culture. The neurospheres-like structure were suspended in the medium, and there were some filaments between the adherent cells from day 3 to day 6 of the culture. The neurospheres were formed by ENSCs on day 8 of the culture. Network-like connections were formed between the adherent cells and differentiated cells after adding 10 % FBS. The differentiated cells were positive for neurofilament and glial fibrillary acidic protein antibodies. The present study established a method to isolate and culture ENSCs from E18 DsRed transgenic rats in the terminal stage of embryonic development. This study would offer a way to obtain plenty of cells for the future research on the transplantation of HD.
