Astragalin attenuates caerulein-induced acute pancreatitis by targeting the NLRP3 signaling pathway and gut...

Acute pancreatitis (AP) has caused great concern worldwide due to its serious threat to human health. Astragalin is a bioactive natural flavonoid compound with several pharmacological activities, but it remains unclear about its effect on AP. The objective of this experiment was to explore the mitigating efficacy of astragalin on caerulein-induced AP model and examine the underlying mechanisms.

Following the assessment of astragalin's direct effects on pancreatic acinar cells using an in vitro AP model, an in vivo mouse model was established to further validate its efficacy and elucidate the underlying mechanisms. Pancreatic histopathology, amylase, and lipase levels of mice were observed to determine the optimal therapeutic dose of astragalin. The network pharmacology and RNA sequencing technology were used to reveal the possible targets and pathways. Subsequent molecular docking and western blot were conducted to validate the association between astragalin and key target molecules, as well as the NLRP3 signaling pathway. Combined with metagenomics and metabolomics analysis, the astragalin effective gut microbiota-metabolite-gene network was constructed. Moreover, fecal microbiota transplantation experiments were performed to clarify the importance of gut microbiota in astragalin-mediated alleviation of AP.

The results showed that astragalin attenuated caerulein-induced injury in AR42J cells in vitro. Consistent with these findings, in vivo experiments revealed that astragalin treatment significantly improved pancreatic pathological injury, cell apoptosis, and systemic inflammatory response in AP mice, particularly at high doses. The integrated analysis of network pharmacology and transcriptomics revealed that the NLRP3 signaling pathway was a key molecular pathway, which was further validated using western blot. Docking analysis showed that 12 target genes had good docking activity with astragalin. More intriguingly, it was found that astragalin could reverse gut microbiota dysbiosis by restoring microbial diversity, altering bacterial community composition, and modulating key metabolic pathways. Specifically, astragalin-effective correlation networks were constructed with Lachnoclostridium sp. YL32, Roseburia intestinalis, Ruminococcus gnavus, Lachnospiraceae bacterium Choco86, Anaerobutyricum hallii, etc. as the core strains, 22 metabolites, including 5-Methoxytryptophan, D-Serine, L-Tryptophan, L-Methionine, etc. as core metabolites, and NLRP3 pathway-related genes as the main regulatory targets. Furthermore, fecal microbiota transplantation experiments confirmed the involvement of gut microbiota in AP remission.

Collectively, these findings identify astragalin as a promising therapeutic agent for AP, targeting both the NLRP3 signaling cascade and gut microbial homeostasis.