One-step sonochemical preparation of redox-responsive nanocapsules for glutathione mediated RNA release

Efficient RNA delivery to targeted cells requires the use of stable interactive carriers that provide RNA protection during the extracellular transit and trigger release once internalised. One strategy to avoid the premature extracellular RNA drain coupled to sufficient intracellular release is the use of stimuli-responsive delivery materials exploiting as a triggering mechanism the redox gradient between the extra- and intracellular compartments. This work describes a facile route for the preparation of redox-active nanocarriers containing disulphides that combine RNA protection and delivery on demand based on intracellular glutathione (GSH) levels. A one-step sonochemical technology was employed to generate thiolated chitosan (TC) nanocapsules with a diameter between 250 and 570 nm and simultaneously load them with RNA. Their size and physiological stability were directly proportional to the extent of disulphide cross-linking, which in turn could be ruled by adjusting the processing pH and degree of chitosan thiolation. TC processing into nanocapsules showed to be advantageous in terms of RNA condensation and protection compared to the typically employed nanocomplexation. Fluorescence microscopy imaging revealed that: (i) the nanocapsules enter the human fibroblasts and migrate to the perinuclear regions within 1 h, and (ii) the cargo release may occur after the internalisation. These redox-responsive and biocompatible drug carriers demonstrated an effective (∼60%) and sustained (up to 72 h) RNA release at intracellular GSH concentrations (10 mM) in vitro, based on disulphide reduction and consequent capsule disassembly.