Propriétés de surface des nanoparticules et interactions avec les cellules endothéliales vasculaires

To date, imaging and treatment of brain tumors are proved to be very inefficient due to the presence of the blood-brain barrier (BBB). The (BBB) is a semipermeable barrier which prevents or restrains most therapeutic and diagnostic molecules reach the central nervous system (CNS). Polymeric nanoparticles (NPs) loaded by therapeutics molecules and diagnostic agents could represent a promising solution to help active ingredients to cross the BBB and as a consequence, their biodistribution to the brain could be improved.  Polymeric NPs composed of di-block copolymers, such as poly (ethylene glycol) blocks (PEG) that bind to polyester hydrophobic chains, are considered one of the most versatile nanocarriers for transporting therapeutic molecules across the BBB. PEG on the surface of NPs improves the NPs colloidal stability. Furthermore, PEG surface coating decreases the non-specific adsorption of proteins on the surface of NPs (also called opsonization); therefore, the clearance rate of the NPs is slowed down and NPs circulation times in blood is extended. Despite the beneficial effect of the PEG coating on the surface of NPs, the exact role of the surface properties related to the PEG chain length on NPs interactions with the vascular endothelial cells is poorly understood.  In first article, the role of PEG chain length on brain vascular endothelial cells endocytosis and transcytosis is investigated on monolayers of bend.3 cells as an in vitro BBB model. The NPs transport mechanisms were then investigated by using different endocytosis inhibitory processes. Our results revealed that NPs endocytosis and transcytosis rates increased with PEG chain length. Higher endocytosis and transcytosis rates were observed for PLA-PEG5000 and PLA-PEG10000 NPs. Moreover, the mechanistic studies demonstrated that the PEG chain length influenced the endocytosis pathway taken by PEGylated NPs through an in vitro model of BBB. In second article, the effect of PEG length on NPs cytotoxicity and inflammatory processes has been investigated in two vascular endothelial cell lines (bEnd.3 and HUVEC). The effect of PEG chain length coating on gene expression that are involved in the inflammation response was investigated by qPCR. Moreover, the potential Reactive Oxygen Species (ROS) generation was evaluated with DCFH-DA probe. The results showed that PEGylated NPs induce a mild and transient activation of inflammatory cytokine and chemokine genes. However, the length of the PEG chains did not modulate significantly gene expression of inflammatory cytokines and chemokines. Furthermore, our results showed that PEGylated NPs did not induce ROS generation.   In summary, the chain length of PEG influences the endocytosis and transcytosis rate. The pathway of endocytosis involved in internalization and transcytosis is influenced by the length of PEG chains. In contrast, differences in the length of PEG chains did not significantly modulate the expression of cytokines and inflammatory chemokines. These results contribute to develop nanoformulations that cross the BBB more efficiently while keeping the toxic and inflammatory effects minimal, particularly on the vascular endothelial cells of the BBB. Nevertheless, these perspectives have to be confirmed on in vivo models.

Keyphrases: Cytokines, blood-brain barrier, Nanoparticles, diblock PLA-b-PEG, Endocytosis, Transcytosis, Reactive Oxygen Species, Cytokines, Chemokines