Aerosol flow-driven instability and droplet-interfacial stabilization of mRNA lipid nanoparticles during mesh...

Messenger RNA (mRNA)-loaded lipid nanoparticles (LNPs) represent a promising platform for pulmonary gene delivery; however, their structural integrity is susceptible to mechanical and interfacial stresses during mesh nebulization. Device-dependent aerosol dynamics, including velocity gradients, turbulence intensity, and droplet breakup behavior, may induce LNP deformation, aggregation, and mRNA leakage. However, the mechanistic links between aerosol physics and nanoparticle destabilization remain insufficiently defined. Here, we systematically investigated device- and formulation-dependent determinants of mRNA-LNP stability during mesh nebulization. Particle image velocimetry and laser diffraction analyses revealed substantial differences in flow symmetry, turbulence intensity, and droplet size distributions across commercial mesh nebulizers. Devices exhibiting laminar-dominant, high-velocity flow profiles generated broader droplet distributions and greater post-nebulization loss of encapsulation efficiency, whereas moderate flow fields reduced the droplet collision frequency and improved the preservation of LNP integrity. Computational fluid dynamics simulations incorporating Weber number-guided droplet collision modeling further indicated regime-dependent transitions between coalescence and rebound interactions, accompanied by distinct pressure redistribution patterns during droplet impact. Excipient screening identified polyvinyl alcohol (PVA) as an effective interfacial stabilizer. PVA supplementation improved post-nebulization particle size, polydispersity index, and encapsulation efficiency in a concentration-dependent manner without compromising aerodynamic performance. Optimized PVA-containing formulations restored in vitro transfection efficiency and significantly enhanced pulmonary mRNA expression in vivo, accompanied by improved nanoparticle delivery to the distal lung regions. Collectively, this study supports a device-formulation co-optimization strategy and provides mechanistic insights into the influence of aerosol-phase hydrodynamics, droplet interaction dynamics, and interfacial modulation on the stability of inhalable mRNA-LNPs.