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In the era of the advanced nanomaterials, use of nanoparticles has been highlighted in biomedical research. However, the demonstration of DNA plasmid delivery with nanoparticles for in vivo gene delivery experiments must be carefully tested due to many possible issues, including toxicity. The purpose of the current study was to deliver a Notch Intracellular Domain (NICD)-encoded plasmid via poly(lactic- co -glycolic acid) (PLGA) nanoparticles and to investigate the toxic environmental side effects for an in vivo experiment. In addition, we demonstrated the target delivery to the endothelium, including the endocardial layer, which is challenging to manipulate gene expression for cardiac functions due to the beating heart and rapid blood pumping. For this study, we used a zebrafish animal model and exposed it to nanoparticles at varying concentrations to observe for specific malformations over time for toxic effects of PLGA nanoparticles as a delivery vehicle. Our nanoparticles caused significantly less malformations than the positive control, ZnO nanoparticles. Additionally, the NICD plasmid was successfully delivered by PLGA nanoparticles and significantly increased Notch signaling related genes. Furthermore, our image based deep-learning analysis approach evaluated that the antibody conjugated nanoparticles were successfully bound to the endocardium to overexpress Notch related genes and improve cardiac function such as ejection fraction, fractional shortening, and cardiac output. This research demonstrates that PLGA nanoparticle-mediated target delivery to upregulate Notch related genes which can be a potential therapeutic approach with minimum toxic effects. 

Notch signaling is a highly conserved signaling system that is required for embryonic development and regeneration of organs. When the signal is lost, maldevelopment occurs and leads to a lethal state. Delivering exogenous genetic materials encoding Notch into cells can reestablish downstream signaling and rescue cellular functions. In this study, we utilized the negatively charged and FDA approved polymer poly(lactic-co-glycolic acid) to encapsulate Notch Intracellular Domain-containing plasmid in nanoparticles. We show that primary human umbilical vein endothelial cells (HUVECs) readily uptake the nanoparticles with and without specific antibody targets. We demonstrated that our nanoparticles are non-toxic, stable over time, and compatible with blood. We further demonstrated that HUVECs could be successfully transfected with these nanoparticles in static and dynamic environments. Lastly, we elucidated that these nanoparticles could upregulate the downstream genes of Notch signaling, indicating that the payload was viable and successfully altered the genetic downstream effects.