GNPs-pIL-4 reprograms macrophage polarization and activates the OSM/GSNOR/ENG axis to improve angiogenesis in ischemic limbs

Therapeutic angiogenesis based on gene therapies is a potential peripheral artery disease (PAD) treatment yet needs a more stable, efficient, and high-affinity delivery vector and an optimized delivery strategy. Here, we engineered three-dimensional graphene nanoparticles modified with folic acid and polyethyleneimine for macrophage-specific delivery of interleukin-4 plasmids (pIL-4), forming GNPs-pIL-4 for local intramuscular injection. GNPs-pIL-4 had uniform size, positive surface charge, and strong nucleic acid loading capacity ( K _d : 25 nanomolar). In vivo, GNPs-pIL-4 reshaped the ischemic microenvironment by inducing reparative M2 macrophage polarization. Mediated by macrophages, GNPs-pIL-4 improved muscle contractility, delayed strength loss, and enhanced blood perfusion and oxygen saturation. Mechanistically, GNPs-pIL-4 specifically turned on the oncostatin M–mediated macrophage-endothelium communication and then activated the angiogenesis, with increased endothelial sprouting, migration, and tube formation. These effects attributed to the down-regulated S -nitrosoglutathione reductase expression, thereby increasing S-nitrosylation at the C209 site of endoglin. GNPs-pIL-4 represents a promising gene therapy strategy for PAD.