Bixin and Montelukast Combination Mitigates the Progression of MASLD to Cytokine-Driven Inflammatory Stress in an In Vitro Fluid-Dynamic Model.

Metabolic-dysfunction-associated steatotic liver disease (MASLD) begins with simple steatosis and progresses to metabolic dysfunction-associated steatohepatitis (MASH), fibrosis, cirrhosis, and liver cancer, primarily driven by increased adiposity, oxidative stress, pro-inflammatory cytokines (such as IL-17), and an imbalance between immune mediators. The multifactorial nature of the disease necessitates combination therapies targeting both inflammatory and lipogenic pathways. Here, the edible pigment Bixin, together with the anti-inflammatory drug Montelukast, was evaluated in an in vitro fluid-dynamic cell culture system of steatotic HepG2 cells induced with the IL-17 cytokine. This model mimicked the progression from hepatic steatosis to cytokine-driven inflammatory stress, marked by elevated levels of pro-inflammatory cytokines, IL-6, and TNF-α. Bixin-Montelukast combination decreased intracellular neutral-lipid accumulation by 34.88%, diminished oxidative stress (29.67% and 41.55% decrease in ROS and malondialdehyde levels, respectively), and decreased liver injury (18.91% decrease in ALT activity). The combination significantly (p < 0.05) downregulated key inflammatory mediators (IL-6, TNF-α) and gene/protein expressions of the lipogenic marker SCD1. Bixin, when combined with Montelukast, ameliorated hepatic lipid accumulation, oxidative stress, lipotoxicity, and inflammation. Such nutraceutical-drug treatment may be a promising strategy to limit disease progression from simple steatosis to cytokine-driven inflammatory sequel of MASLD.