Hepatic Cholesterol Regulation Through Multi-Botanical Extract Targeting of the PCSK9–LDLr–SREBP-2 Axis in HepG2 Cells

<b>Background/Objectives</b>: Botanical and nutraceutical approaches have increasingly been considered as alternatives or complements to conventional lipid-lowering therapies, particularly in individuals with mild-to-moderate dyslipidemia or statin intolerance. This study aimed to evaluate a multi-botanical formulation, combining black garlic, sesame, <i>Gastrodia elata</i>, and <i>Primula veris</i> extracts, for its effects on hepatic cholesterol regulation and the PCSK9-LDLr-SREBP-2 axis in vitro. <b>Methods</b>: Each extract was chemically characterised for its polysaccharide, polyphenol, flavonoid, and sesamin content. HepG2 cells were exposed to normal (5 mM) or high-glucose (30 mM) conditions to mimic metabolic stress. Dose-response studies identified optimal concentrations for cell viability. Hepatic safety was assessed via MTT and ROS assays, while cholesterol metabolism was evaluated by measuring HMG-CoA reductase levels, total cholesterol, LDL levels, bile acid production, free cholesterol levels, and the expression of PCSK9, LDLr, and SREBP-2 using ELISA and Western blot. <b>Results</b>: All individual extracts improved cell viability, reduced oxidative stress, and moderately modulated cholesterol metabolism. The multi-botanical combination exhibited synergistic effects, enhancing cell viability (+47.5% vs. untreated), suppressing ROS, reducing HMGR levels, and lowering total intracellular cholesterol more effectively than single extracts or the statin-like reference RYRF. Importantly, the combination strongly downregulated PCSK9 and inhibited SREBP-2 proteolytic activation while upregulating LDLr, indicating coordinated transcriptional and post-translational regulation. Bile acid production and free cholesterol excretion were also significantly increased, supporting improved cholesterol clearance. <b>Conclusions</b>: This four-botanical formulation effectively modulates hepatic cholesterol homeostasis via a multifactorial, synergistic mechanism distinct from statin-like agents. The results suggest its potential as a safe, non-statin strategy to support cardiometabolic health. Future studies are warranted to confirm long-term efficacy and clinical relevance.