Design of a biomimetic and neuroprotective nanocapsule for the treatment of post-ischemic stroke effects

Christos Tapeinos¹, Matteo Battaglini^1,2, Attilio Marino¹, Ivana Cavaliere³, Gianni Ciofani^1,3

¹ Smart Bio-Interfaces, Istituto Italiano di Tecnologia, Pontedera (PI), Italy

²The Biorobotics Institute, Scuola Superiore Sant’Anna, Pontedera (PI), Italy

³ Department of Mechanical and Aerospace Engineering, Politecnico di Torino, Torino, Italy

Presenting author’s e-mail: This email address is being protected from spambots. You need JavaScript enabled to view it.; This email address is being protected from spambots. You need JavaScript enabled to view it.

INTRODUCTION

Ischemic stroke occurs due to reduced perfusion to a brain region, because of the blockage of a blood vessel, resulting in death or permanent neurological deficits. Although, many treatments that make use of therapeutic nanoparticles have been presented, unfortunately to date, no effective treatment has been found to prevent damage to the ischemic brain after stroke. Oxidative stress is related to the pathogenesis of stroke, and overproduction of reactive oxygen (ROS) and reactive nitrogen species (RNS) are thought to be the main cause of this. In view of this, we hypothesized that the targeted delivery of antioxidant (CeO₂) nanoparticles (NPs) and iNOS inhibitors (i.e. L-NIL) across the blood brain barrier (BBB) will reduce the overproduced ROS and RNS, resulting to the amelioration of the neurological deficits caused by oxidative stress.

METHODS

The fabrication of the biomimetic and neuroprotective nanocapsules (BIONICS) was achieved using a hot-emulsion ultra-sonication method, while their characterization was achieved using various material characterization techniques (SEM, TEM, EDX, ICP, FT-IR, TGA). The stability of the nanoparticles was studied using DLS in various biologically relevant media and under conditions that simulate oxidative stress at different time points. The antioxidant ability of CeO₂ NPs was studied using EPR as well as antioxidant assay kits. A nitric oxide assay kit was also used to assess the ability of CeO₂ NPs and L-NIL to inhibit the overproduction of RNS. The internalization of the LMNVs was studied at various time points using confocal microscopy and their ability to inhibit oxidative stress in vitro was tested on human astrocytes and endothelial cells using flow cytometry.

RESULTS AND DISCUSSION

The synthesized nanosystem that had an average size of 150 nm, comprised of 70% of a lipid matrix and 30% of CeO₂ NPs. BIONICS presented excellent short-term and long-term stability in aqueous media, while it’s stability was affected under conditions simulating oxidative stress. EPR results showed that the CeO₂ NP-loaded nanocapsules demonstrated excellent antioxidant capacity which was similar to the antioxidant capacity of the corresponding plain CeO₂ NPs. In addition, CeO₂ NPs and L-NIL were found able to scavenge overexpressed RNS. Confocal imaging demonstrated a time-dependent internalization of BIONICS while flow cytometry results demonstrated that BIONICS has a protective effect against oxidative stress by inhibiting ROS-mediated apoptosis.

CONCLUSION

BIONICS demonstrated excellent short-term and long-term stability as well as excellent antioxidant capacity, resulting in the inhibition of ROS and RNS mediated oxidative stress, and in a subsequent neuroprotection of astrocytes and endothelial cells.

ACKNOWLEDGMENTS

“This project has received funding from the European Union’s Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement No 793644”.