Network-guided Evaluation of β-sitosterol in Inflammatory and Profibrotic Mesangial-Cell Models Relevant to Chronic Glomerulonephritis

Chronic glomerulonephritis (CGN) is characterized by persistent inflammatory injury and progressive fibrotic remodeling, yet compound-target relationships underlying natural-product-based interventions remain incompletely defined. This study integrated network pharmacology, validated molecular docking, and in vitro experiments to evaluate candidate constituents of Cordyceps sinensis (C. sinensis) in CGN-related pathological processes. Candidate constituents of C. sinensis were screened, and the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform-derived compound-associated targets were intersected with CGN-associated genes. Functional enrichment and protein-protein interaction analyses were used to prioritize biological processes and hub targets. PTGS2 was identified as a key overlapping target. Molecular docking was performed using a celecoxib-bound COX-2 structure. Re-docking of the co-crystallized celecoxib ligand reproduced the crystallographic pose with an RMSD of 0.876 Å, supporting the docking protocol. β-Sitosterol and linoleyl acetate showed predicted compatibility with the COX-2 docking region, with binding affinities of -7.2 and -7.5 kcal/mol, respectively. β-Sitosterol was selected for compound-level validation in HBZY-1 rat glomerular mesangial cells. At 0.5-10 µM, β-sitosterol did not markedly reduce cell viability. In Lipopolysaccharide (LPS)-stimulated cells, β-sitosterol reduced Tnf, Il6, and Ptgs2 expression, decreased prostaglandin E2 (PGE2) production, and reduced cyclooxygenase-2 (COX-2) protein abundance. In transforming growth factor β1 (TGF-β1)-treated cells, β-sitosterol reduced Col1a1 and Acta2 expression and decreased alpha-smooth muscle actin (α-SMA) protein abundance. These findings provide hypothesis-generating evidence that β-sitosterol modulates inflammatory and profibrotic activation in mesangial-cell models, but they do not establish direct COX-2 enzymatic inhibition or therapeutic efficacy in CGN.