Dual Modulation of Canonical and Non-canonical TGF-β/ROS/Erk1/2 Pathways: Synergistic Activation of Nrf-2 and Antioxidant Enzymes (SOD1, GPx, HO-1) by Quercetin Loaded in Solid Lipid Nanoparticles and Curcumin in Atherosclerosis Therapy.

Background: Atherosclerosis remains the leading cause of mortality worldwide, highlighting the urgent need for innovative treatments targeting chronic inflammation. Recent research indicates that quercetin (QCT) and curcumin, two naturally occurring compounds, have potential therapeutic benefits in cardiovascular diseases. Objectives: This study focuses on the novel synthesis of nano-quercetin (N-QCT) encapsulated in solid lipid nanoparticles (SLNs) and investigates the synergistic cardioprotective effects of N-QCT and curcumin on human vascular smooth muscle cells (VSMCs). The underlying molecular mechanisms, particularly the involvement of the TGF-β signaling pathway in VSMCs, are explored. Methods: The VSMCs, including TGF-β-stimulated VSMCs, were treated with N-QCT, curcumin, or a combination of both. The MTT assay was performed to evaluate the cytotoxic effects of these treatments. The cytotoxicity of various concentrations of curcumin and QCT was used to calculate the Combination Index (CI), with CI analysis quantifying synergy or antagonism. Furthermore, following TGF-β stimulation, antioxidant enzyme activity, nuclear transcription factor erythroid 2-related factor (Nrf2) mRNA expression, reactive oxygen species (ROS) production, NADPH oxidases (NOX) expression, and extracellular signal-regulated kinase (Erk)1/2 phosphorylation were measured in the treated VSMCs. Results: The N-QCT and curcumin significantly influenced Nrf2 mRNA expression and upregulated downstream antioxidant enzymes, including HO-1, GPx, and SOD1. The combination treatment further enhanced Nrf2 protein expression and modulated Erk1/2 phosphorylation. Notably, the synergistic effect of the combination produced pronounced cardioprotective outcomes, characterized by reduced ROS production and decreased phosphorylation of Erk1/2 via the TGF-β/NOX/Erk1/2 and ROS/Nrf2 signaling pathways. Conclusions: The findings demonstrate that the combination of QCT encapsulated in SLNs and curcumin synergistically reduces oxidative stress and inflammation in TGF-β-stimulated VSMCs. This effect is achieved through the inhibition of ROS/Erk1/2 signaling and the activation of Nrf2 and antioxidant enzymes. These natural compounds, when used together, represent a promising therapeutic approach for mitigating the inflammatory processes associated with atherosclerosis.