Exploring the antidepressant-like effect of stigmasterol using network pharmacology with molecular docking and in vivo experimental validation

With a global prevalence of 3.8% and affecting approximately 300 million people, depression remains a global burden calling for renewed efforts including novel antidepressants to address treatment gaps. Stigmasterol, an unsaturated naturally abundant phytosterol with reported neuropsychiatric activity in preclinical studies, was studied for potential antidepressant-like activity and mechanism of action using computational and experimental approaches. Using network pharmacology, we identified potential key targets of major depressive disorder and stigmasterol (STG) by analysing intersection genes for protein-protein interaction, Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. We subsequently carried out molecular docking of stigmasterol with key targets identified to confirm antidepressant activity and potential mechanism(s) while the forced swim test (FST) and tail suspension test (TST) were used for experimental validation. To experimentally validate the involvement of monoaminergic mechanism(s) in STG's action, mice were pretreated with selective inhibitors of monoamine synthesis and storage after which the antidepressant-like effects of STG was re-evaluated in FST. Forty intersection target genes were obtained with AKT1, TP53 and IL1B as well as MAO_B, MAO_A and SLC6A4 being identified as key targets. GO and KEGG enrichment analysis further reinforced the involvement of monoamine regulation, especially serotonin. Molecular docking showed STG interacts with SLC6A4 (-9.732 kcal mol^-1), MAO_A (-12.471 kcal mol^-1) and MAO_B (-14.614 kcal mol^-1) further buttressing monoaminergic involvement. STG demonstrated significant antidepressant-like effects similar to fluoxetine-treated mice in both FST and TST. This antidepressant action was significantly influenced by serotonin neurotransmission. Overall, we confirmed the antidepressant-like activity of STG and its possible multitarget potential in depression management.