Sodium butyrate promotes enteric glial cells neurogenesis by inhibiting Kdm2a and inducing Klf4 expression

To explore the potential molecular targets and mechanisms of sodium butyrate (NaB) in the treatment of Hirschsprung disease (HSCR), with a focus on analyzing its regulatory effects on enteric glial cell function and neurogenesis.

A HSCR rat model was established, and targeted metabolomics was used to detect differences in short-chain fatty acids (SCFAs) between sham and model groups. Rats were treated with a mixture of SCFAs and immunofluorescence was used to assess the expression of neurogenesis markers in colon tissues. Network pharmacology identified potential targets of NaB, a major SCFA component. Molecular docking was performed to evaluate the binding affinity between NaB and selected target proteins. Sodium butyrate was applied to the enteric glial cell line in vitro. The expression levels of glial cell and neural stem cell markers were measured using qRT-PCR, Western blot, and immunofluorescence assays. Cell migration was assessed by Transwell assays, apoptosis and proliferation were analyzed by flow cytometry. siRNA transfection was conducted to knock down candidate genes in vitro.

Targeted metabolomics showed significantly reduced SCFAs in HSCR rats; exogenous SCFAs treatment increased Nestin and p75^NTR in colon tissue. Network pharmacology showed significant overlap between NaB and HSCR targets, with Klf4 as a core node. After NaB treatment, the expression of the glial cell marker Gfap decreased, while the expression levels of stem cell markers Nes and p75^NTR were upregulated. Correspondingly, low concentrations of NaB significantly enhanced the migratory ability of EGCs, with no obvious effect on proliferation and apoptosis. Mechanistically, NaB inhibits demethylase Kdm2a, increases H3K36me2 levels, and activates Klf4 expression.

This study reveals that NaB promotes the transformation of enteric glial cells into an undifferentiated stem cell-like state and reshapes the neurogenic microenvironment through epigenetic reprogramming (Kdm2a/H3K36me2/Klf4 axis), providing new potential targets and therapeutic strategies for the treatment of HSCR.