Abstract:

Chronic kidney disease (CKD) represents a growing global health challenge due to its high prevalence, multifactorial pathogenesis, and limited therapeutic options. Phytosterols, a class of naturally occurring bioactive compounds widely present in plants, have garnered increasing attention for their anti-inflammatory, antioxidant, and immunomodulatory properties. Among these, stigmasterol (Stig) and β-sitosterol (β-Sito) are the most abundant and biologically active representatives, previously reported to confer protective effects against cardiovascular, metabolic, and renal disorders. In the present study, we explored the potential molecular mechanisms of these phytosterols in a folic acid (FA)-induced CKD mouse model. Comprehensive transcriptomic analysis revealed that Sting1, a gene critically involved in innate immune responses, was markedly upregulated in CKD and significantly downregulated following phytosterol treatment. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses further implicated immune- and inflammation-related pathways, such as cytokine-cytokine receptor interaction and chemokine signaling, as potential mediators of the therapeutic effects. Molecular docking and molecular dynamics (MD) simulations demonstrated stable binding of both phytosterols to the Sting1 protein, with stigmasterol exhibiting higher binding affinity and greater conformational stability. Detailed analyses of root-mean-square deviation (RMSD), root-mean-square fluctuation (RMSF), and radius of gyration (Rg) supported the structural integrity of the phytosterol-Sting1 complexes. MM-PBSA energy decomposition identified PHE-268 and PRO-208 as key residues mediating the interactions. Moreover, cellular thermal shift assays (CETSA) in HK-2 cells confirmed that both compounds enhanced the thermal stability of Sting1 under moderate heat stress (40–49 °C), providing additional evidence for a direct interaction. Taken together, these findings indicated that phytosterols might exert nephroprotective effects through direct modulation of Sting1, thereby attenuating immune and inflammatory dysregulation in CKD. This study offered novel mechanistic insights and established a theoretical foundation for the development of phytosterol-based therapeutic interventions for CKD.

Key words: Chronic kidney disease, Phytosterols, Molecular docking, Molecular dynamics simulations, Sting1