Bacillus subtilis extracellular vesicles surface-displaying superoxide dismutase attenuate alcoholic liver injury by activating the Nrf2/HO-1 axis

Abstract

Alcoholic liver disease (ALD) represents a globally prevalent progressive hepatic disorder with continuously rising incidence. Its pathological mechanisms mainly stem from the direct toxic effects of alcohol metabolites, accompanied by aggravated oxidative stress, lipid overaccumulation and inflammatory infiltration.. While current therapies that act on these pathogenic mechanisms alleviate symptoms, they are often limited by gastrointestinal adverse effects and long-term hepatic metabolic burden. Bacterial extracellular vesicles (BEVs), emerging as natural carriers of bioactive molecules and mediators of intercellular communication, offer a novel hepatoprotective strategy against alcoholic liver injury. Superoxide dismutase (SOD), as a crucial antioxidant, has been extensively verified for its capacity to eliminate free radicals. Here, we leverage the advantages of BEVs and the multiple effects of SOD, utilizing surface display technology to locate SOD in Bacillus subtilis 168-derived EVs (termed SEVs). In vitro simulated digestive fluids experiment confirms the stability and digestive resistance of SEVs, while in vivo biodistribution assays demonstrate the liver-targeting capability. In ethanol-exposed hepatocytes, SEVs significantly attenuate reactive oxygen species (ROS) overproduction and lipid deposition. In a murine ALD model, SEVs administration reduces hepatic steatosis, serum transaminase levels, and inflammatory infiltration. Mechanistically, SEVs activate the Nrf2/HO-1 antioxidant pathway, a key regulator of cellular redox balance and inflammation, thereby counteracting oxidative damage and inflammatory reaction caused by alcohol stimulation. Notably, SEVs exhibit superior biocompatibility without inducing secondary hepatic burden. Our findings emphasize the dual advantages of SEVs as liver-targeted delivery vehicles and multifunctional liver-protective agents, highlighting their translational potential for ALD management.

Graphical abstract

Keywords

Alcoholic liver disease / Bacterial extracellular vesicles / Surface display / Superoxide dismutase / Nrf2/HO-1

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Funding

the National Natural Science Foundation of China (42277279)

the Zhejiang Provincial Natural Science Foundation of China (LQN25C010005)

the financial support from ‘Pioneer’ and‘Leading Goose’R&D Program of Zhejiang Province(2025C01100)

Zhejiang Provincial Traditional Chinese medicine science and technology project(2026ZL0262)

Medical and Health Science Program of Zhejiang Province(2025HY0276)

Zhejiang Science and Technology Plan for Disease Prevention and Control (2026JKY092)

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