Abstract
The delivery of therapeutics to the central nervous system (CNS) remains a major challenge in part due to the presence of the blood-brain barrier (BBB). Recently, cell-derived vesicles, particularly exosomes, have emerged as an attractive vehicle for targeting drugs to the brain, but whether or how they cross the BBB remains unclear. Here, we investigated the interactions between exosomes and brain microvascular endothelial cells (BMECs) in vitro under conditions that mimic the healthy and inflamed BBB in vivo. Transwell assays revealed that luciferase-carrying exosomes can cross a BMEC monolayer under stroke-like, inflamed conditions (TNF-α activated) but not under normal conditions. Confocal microscopy showed that exosomes are internalized by BMECs through endocytosis, co-localize with endosomes, in effect primarily utilizing the transcellular route of crossing. Together, these results indicate that cell-derived exosomes can cross the BBB model under stroke-like conditions in vitro. This study encourages further development of engineered exosomes as drug delivery vehicles or tracking tools for treating or monitoring neurological diseases.
Highlights
Despite significant advances in drug delivery, a major challenge remains in delivering therapeutics effectively to the brain for the treatment of central nervous system (CNS) diseases, including trauma, stroke, autoimmune diseases, neurodegenerative diseases and tumors[1, 4, 35, 48, 67]
In order to monitor their distribution across brain microvascular endothelial cells (BMECs) monolayers in vitro, the exosomes were labeled with humanized Gaussia luciferase (hGluc). hGluc was fused with lactadherin, which can be bound to cell membrane phophatidylserines (PS) and is highly enriched on the outer leaflet of exosomal membrane20, 56. 293T cells were transduced with lentivirus expressing each free hGluc, hGluc-Lactadherin without and with a Green Fluorescent Protein (GFP) tag used to monitor transduction efficiency (Fig. S1a)
As adherens junctions (AJs) and TJs play a major role in preventing passage through the blood-brain barrier (BBB) and maintaining intercellular tight junctions, we examined if TNF-α alters the function of intercellular tight junctions
Summary
Despite significant advances in drug delivery, a major challenge remains in delivering therapeutics effectively to the brain for the treatment of central nervous system (CNS) diseases, including trauma, stroke, autoimmune diseases, neurodegenerative diseases and tumors[1, 4, 35, 48, 67]. The brain endothelial cells have a complex arrangement of tight junctions (TJs) and adherens junctions (AJs), which play key roles in regulating paracellular permeability[72] These junctions prevent transport of most molecules except those normally used for homeostasis, including for nutrition or bidirectional hormonal communication and reflecting the changing properties of the BBB depending on conditions[4]. While this complex interface protects the brain from harmful chemicals or toxins that may be present in systemic circulation, it results in the inability of therapeutics to cross the BBB, with approximately 98% of small molecule pharmaceuticals and almost all of large molecule biologic drugs, including recombinant proteins, monoclonal antibodies, or genebased medicines, failing to cross the BBB40, 41. Particulate drug carriers such as dendrimers[27], nanoparticles[43] and liposomes[64] have been tested for drug delivery across the BBB, they remain not widely used in the clinic considering their immunogenicity, limited half-life in vivo, and, importantly, relative lack of specificity and efficacy in crossing the BBB17, 67
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