Conversion of intestinal smooth muscle cells in Hirschsprung's disease into enteric neuron-like cells using small...

The enteric nervous system (ENS), often referred to as the "second brain," is a complex network of neurons and glial cells in the gut wall, crucial for regulating its functions. Defects in the ENS can lead to Hirschsprung's disease (HSCR), a serious congenital disorder affecting approximately 1 in 5,000 live births. A promising approach is ENS transplantation. However, obtaining sufficient non-gene edited ENS remains a significant challenge.

In this study, we isolated intestinal smooth muscle cells (ISMCs) from pediatric patients diagnosed with Hirschsprung's disease (HSCR). Subsequently, we converted these ISMCs into enteric neuron-like cells using a set of six small molecules. The expression levels of neural cell markers were assessed through Immunofluorescence (IF) staining and quantitative Real-Time Polymerase Chain Reaction (qRT-PCR).

We demonstrated that ISMCs could be directly reprogrammed into enteric neurons-like cells through a chemical cocktail comprising six small molecules: VCRFYS-where V represents valproic acid, C denotes CHIR99021, R stands for Repsox, F indicates forskolin, Y refers to Y27632, and S signifies SP600125-achieving over 80 % TUJ1-positive yield after five days of neural conversion. Following further maturation, these chemically induced neuronal cells exhibited neuron-specific morphology and gene expression profiles.

Our findings collectively present a transgene-free and chemical-only approach for the direct reprogramming of ISMCs into neuron-like cells. This method offers an alternative strategy for replenishing the ENS in cases of HSCR.