Inhalable Plant Exosome-like Nanoparticles from Houttuynia cordata Ameliorate Acute Lung Injury via Redox Modulation and Immune Reprogramming

Background: Acute lung injury (ALI) features oxidative stress, macrophage dysfunction, and barrier disruption, with limited safe therapies. Plant-derived exosome-like nanovesicles (PENs) delivered noninvasively to the lung are promising but understudied. Methods: In vitro, HELNs were evaluated for their ability to scavenge ROS, restore mitochondrial membrane potential (MMP) modulate M1/M2 polarization, and suppress pro-inflammatory cytokines. RNA-seq was also used to evaluate the TNF/NOD-like pathway and mitochondrial-related gene expression. In vivo, the evaluated parameters included biodistribution, therapeutic outcomes (lung wet/dry weight ratio, BALF protein concentration, MPO and MDA levels, histology), immune cell profiles, and cytokines. Biosafety was assessed via hematology, serum biochemistry, and histology. Results: In vitro, HELNs acted dose-dependently by scavenging ROS, recovering MMP, promoting M2 and suppressing M1 polarization, and reducing TNF-α, IL-6 release. Additionally, RNA-seq confirmed downregulation of TNF/NOD-like signals and regulated mitochondrial gene expression. In vivo, nebulized HELNs preferentially accumulated in inflamed lungs, with significantly higher pulmonary retention than intravenous delivery. HELNs reduced lung wet/dry weight ratio, BALF protein, MPO and MDA levels, and histological injury scores; decreased neutrophils and CD4+/CD8+ T cells; increased alveolar macrophages and Tregs; and lowered TNF-α, IL-1β, IL-6. No significant systemic toxicity was observed. Conclusions: Nebulized HELNs offer a safe, effective, noninvasive ALI therapy by disrupting the oxidative stress-inflammation vicious cycles, reprogramming pulmonary immunity, and restoring barrier integrity, with strong translational potential.