Decoding nattokinase efficacy: From digestion and absorption to lipid pathway modulation in high-fat diet-induced atherosclerosis

Nattokinase (NK), an alkaline serine protease, is recognized for its cardiovascular benefits through the modulation of blood coagulation and fibrinolytic pathways. Atherosclerosis (AS), a primary cause of cardiovascular disease (CVD) worldwide, is responsive to dietary interventions. However, studies investigating NK’s digestion, absorption, and mechanisms in mitigating AS remain limited. This study evaluated the effects of high-dose NK treatment (H-NK, 900 FU/kg body weight) administered over six weeks in ApoE -/- mice, which resulted in significant reductions in atherosclerotic lesions. In vitro digestion simulations showed that 25.8% of NK’s enzymatic activity was retained post-digestion, and thrombolysis assays demonstrated a thrombolytic efficacy of 30%-40% at concentrations ≥ 5,000 μg/mL, regardless of clot age. Absorption experiments using a rat everted intestinal sac model revealed a small intestine absorption rate of 37.0%, with the absorbed peptides being predominantly hydrophobic, uncharged, and shorter than 10 amino acids. Proteomic analysis of mouse liver tissues identified that high-dose NK down-regulated PXDN (peroxidasin) and PNLIP (pancreatic lipase), proteins involved in promoting lipid oxidation, and intestinal lipid absorption. These proteomic changes enhanced lipid metabolism, reduced lipid peroxidation, and alleviated AS through the "Lipid and Atherosclerosis" pathway. Overall, this study provides a deeper understanding of NK’s biological activity and supports its potential as a novel dietary supplement for the management of CVD.