Suppression of SRC protein kinase activity alleviates the severity of aganglionosis by impairing CAV1/FLNA expression.

ObjectiveHirschsprung's disease (HSCR) is a rare congenital disorder attributed to the defects of enteric neural crest cells. We aim to identify characteristic phosphorylation proteins and preliminarily explore underlying related action mechanisms in HSCR.MethodsColon samples from HSCR patients underwent proteomic and phosphoproteomic sequencing to identify differentially expressed phosphoproteins (DEPPs) and proteins (DEPs). Interaction network construction and analysis of correlations with upstream phosphorylating kinases were employed to pinpoint core proteins. HSCR rat models were established through enema administration of Benzalkonium chloride and evaluated by measuring colon cross-sectional area, colon weight, AchE, and PGP9.5 levels. Histopathological damage was assessed via hematoxylin and eosin staining. Protein expression was analyzed using western blotting. Furthermore, the impact of SRC kinase in HSCR was investigated utilizing an SRC-specific inhibitor in HSCR rat models.ResultsA total of 5725 DEPPs were identified, with SRC kinase emerging as a key regulatory protein. In the HSCR rat model, SRC expression was elevated along with increased pCAV1 and FLNA levels. Notably, inhibition of SRC protein kinase activity by 1-(tert-butyl)-3-(4-chlorophenyl)-1H-pyrazolo[3,4-d] pyrimidin-4-amine (PP2) led to reduced colon cross-sectional area and weight, an increase in the number of colonic ganglion cells, heightened AchE levels, enhanced PGP9.5 expression, and slight enlargement of the crypt, thereby alleviating HSCR symptoms in rats. Additionally, SRC kinase inhibition following PP2 treatment decreased the expression of pCAV1 and FLNA.ConclusionsInhibition of SRC kinase activity may potentially reduce CAV1/FLNA expression, ultimately alleviating the severity of HSCR in rats.