Developing and optimizing a biocompatible tauopathy model using extracellular vesicle-mediated gene delivery.

Introduction: Tauopathy models are essential in vitro systems for investigating tau-targeted therapies and advancing Alzheimer's disease research. Extracellular vesicles (EVs), owing to their high biocompatibility, low toxicity, and reduced immunogenicity, represent promising carriers for gene delivery and disease modeling. Methods: We investigated the potential of EVs as a delivery system for the human four-repeat tau isoform lacking N-terminal sequences (4R0N) and enhanced green fluorescent protein (EGFP) into Neuro-2a cells. EV-mediated transfection efficiency was compared with conventional methods, including lentiviral and chemical (lipofectamine and polyethyleneimine, PEI) approaches. Response surface methodology (RSM) was used to optimize EV-mediated delivery parameters. Results: EVs successfully delivered large plasmid DNA into Neuro-2a cells, resulting in detectable tau and EGFP expression. Optimization via RSM further improved gene delivery efficiency and reproducibility compared to unoptimized EV preparations and conventional transfection methods. Discussion: These findings demonstrate that EVs can serve as a robust and biocompatible platform for tau gene delivery, providing a promising alternative to traditional transfection strategies for generating physiologically relevant tauopathy models.