Nanoparticulate carriers, often referred to as nanoparticles (NPs), represent an important pharmacological advance for drug protection and tissue-specific drug delivery. Accessing the central nervous ...system (CNS), however, is a complex process regulated by mainly three brain barriers. While some leukocyte (i.e., immune cell) subsets are equipped with the adequate molecular machinery to infiltrate the CNS in physiological and/or pathological contexts, the successful delivery of NPs into the CNS remains hindered by the tightness of the brain barriers. Here, we present an overview of the three major brain barriers and the mechanisms allowing leukocytes to migrate across each of them. We subsequently review different immune-inspired and -mediated strategies to deliver NPs into the CNS. Finally, we discuss the prospect of exploiting leukocyte trafficking mechanisms for further progress.
Most drug delivery strategies perform suboptimally in the CNS due to the presence of three major brain barriers: the blood–brain barrier, the blood–leptomeningeal barrier, and the blood–cerebrospinal fluid barrier.Immune cells use a plethora of molecules to bypass the different barriers and infiltrate the CNS under homeostasis and in diverse pathological contexts.Nanoparticulate carriers, often shortened to nanoparticles (NPs), have recently emerged as superior vehicles capable of protecting drugs while also promoting tissue-specific delivery, but remain ineffective in transporting pharmaceutical agents into the CNS.Exploiting the molecular and cellular pathways used by immune cells to overcome the protective brain barriers may prove useful for enhancing NP delivery into the CNS.
Plasma cells (PC) are found in the CNS of multiple sclerosis (MS) patients, yet their source and role in MS remains unclear. We find that some PC in the CNS of mice with experimental autoimmune ...encephalomyelitis (EAE) originate in the gut and produce immunoglobulin A (IgA). Moreover, we show that IgA+ PC are dramatically reduced in the gut during EAE, and likewise, a reduction in IgA-bound fecal bacteria is seen in MS patients during disease relapse. Removal of plasmablast (PB) plus PC resulted in exacerbated EAE that was normalized by the introduction of gut-derived IgA+ PC. Furthermore, mice with an over-abundance of IgA+ PB and/or PC were specifically resistant to the effector stage of EAE, and expression of interleukin (IL)-10 by PB plus PC was necessary and sufficient to confer resistance. Our data show that IgA+ PB and/or PC mobilized from the gut play an unexpected role in suppressing neuroinflammation.
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•Gut-derived IgA+ plasma cells (PC) access extra-intestinal tissues in the steady state•Gut-derived IgA+ PC access the CNS during EAE and attenuate disease•IgA+ PC attenuate EAE in an IL-10-dependent manner•An IgA-promoting protist and BAFF overexpression both confer resistance to EAE
Immune cells can travel from the gut to the brain to suppress neuroinflammation in a mouse model of multiple sclerosis.
Helminthic worms are ancestral members of the intestinal ecosystem that have been largely eradicated from the general population in industrialized countries. Immunomodulatory mechanisms induced by ...some helminths mediate a “truce” between the mammalian host and the colonizing worm, thus allowing for long-term persistence in the absence of immune-mediated collateral tissue damage. This concept and the geographic discrepancy between global burdens of chronic inflammatory diseases and helminth infection have sparked interest in the potential of using helminthic worms as a therapeutic intervention to limit the progression of autoimmune diseases such as multiple sclerosis (MS). Here, we present and evaluate the evidence for this hypothesis in the pre-clinical animal model of MS, experimental autoimmune encephalitis, in helminth-infected MS patients and in clinical trials of administered helminth immunotherapy (HIT).
The blood-brain barrier (BBB) has a major role in maintaining brain homeostasis through the specialized function of brain endothelial cells (BECs). Inflammation of the BECs and loss of their ...neuroprotective properties is associated with several neurological disorders, including the chronic neuro-inflammatory disorder multiple sclerosis (MS). Yet, the underlying mechanisms of a defective BBB in MS remain largely unknown. Endothelial to mesenchymal transition (EndoMT) is a pathophysiological process in which endothelial cells lose their specialized function and de-differentiate into mesenchymal cells. This transition is characterized by an increase in EndoMT-related transcription factors (TFs), a downregulation of brain endothelial markers, and an upregulation of mesenchymal markers accompanied by morphological changes associated with cytoskeleton reorganization. Here, we postulate that EndoMT drives BEC de-differentiation, mediates inflammation-induced human BECs dysfunction, and may play a role in MS pathophysiology. We provide evidence that stimulation of human BECs with transforming growth factor (TGF)-β1 and interleukin (IL)-1β promotes EndoMT, a process in which the TF SNAI1, a master regulator of EndoMT, plays a crucial role. We demonstrate the involvement of TGF-β activated kinase 1 (TAK1) in EndoMT induction in BECs. Finally, immunohistochemical analysis revealed EndoMT-associated alterations in the brain vasculature of human post-mortem MS brain tissues. Taken together, our novel findings provide a better understanding of the molecular mechanisms underlying BECs dysfunction during MS pathology and can be used to develop new potential therapeutic strategies to restore BBB function.
Primary cell isolation from the central nervous system (CNS) has allowed fundamental understanding of blood-brain barrier (BBB) properties. However, poorly described isolation techniques or ...suboptimal cellular purity has been a weak point of some published scientific articles. Here, we describe in detail how to isolate and enrich, using a common approach, endothelial cells (ECs) from adult mouse brains, as well as pericytes (PCs) and astrocytes (ACs) from newborn mouse brains. Our approach allowed the isolation of these three brain cell types with purities of around 90%. Furthermore, using our protocols, around 3 times more PCs and 2 times more ACs could be grown in culture, as compared to previously published protocols. The cells were identified and characterized using flow cytometry and confocal microscopy. The ability of ECs to form a tight monolayer was assessed for passages 0 to 3. The expression of claudin-5, occludin, zonula occludens-1, P-glycoprotein-1 and breast cancer resistance protein by ECs, as well as the ability of the cells to respond to cytokine stimuli (TNF-α, IFN-γ) was also investigated by q-PCR. The transcellular permeability of ECs was evaluated in the presence of pericytes or astrocytes in a Transwell® model by measuring the transendothelial electrical resistance (TEER), dextran-FITC and sodium fluorescein permeability. Overall, ECs at passages 0 and 1 featured the best properties valued in a BBB model. Furthermore, pericytes did not increase tightness of EC monolayers, whereas astrocytes did regardless of their seeding location. Finally, ECs resuspended in fetal bovine serum (FBS) and dimethyl sulfoxide (DMSO) could be cryopreserved in liquid nitrogen without affecting their phenotype nor their capacity to form a tight monolayer, thus allowing these primary cells to be used for various longitudinal in vitro studies of the blood-brain barrier.
Resolution of inflammation is an active and regulated process that leads to the clearance of cell debris and immune cells from the challenged tissue, facilitating the recovery of homeostasis. This ...physiological response is coordinated by endogenous bioactive lipids known as specialized pro-resolving mediators (SPMs). When resolution fails, inflammation becomes uncontrolled leading chronic inflammation and tissue damage, as occurs in multiple sclerosis (MS).
SPMs and the key biosynthetic enzymes involved in SPM production were analysed by metabololipidomics and qPCR in active brain lesions, serum and peripheral blood mononuclear cells (PBMC) of MS patients as well as in the spinal cord of mice with experimental autoimmune encephalomyelitis (EAE). We also tested the therapeutic actions of the SPM coined Maresin-1 (MaR1) in EAE mice and studied its impact on inflammation by doing luminex and flow cytometry analysis.
We show that levels of MaR1 and other SPMs were below the limit of detection or not increased in the spinal cord of EAE mice, whereas the production of pro-inflammatory eicosanoids was induced during disease progression. Similarly, we reveal that SPMs were undetected in serum and active brain lesion samples of MS patients, which was linked to impaired expression of the enzymes involved in the biosynthetic pathways of SPMs. We demonstrate that exogenous administration of MaR1 in EAE mice suppressed the protein levels of various pro-inflammatory cytokines and reduced immune cells counts in the spinal cord and blood. MaR1 also decreased the numbers of Th1 cells but increased the accumulation of regulatory T cells and drove macrophage polarization towards an anti-inflammatory phenotype. Importantly, we provide clear evidence that administration of MaR1 in mice with clinical signs of EAE enhanced neurological outcomes and protected from demyelination.
This study reveals that there is an imbalance in the production of SPMs in MS patients and in EAE mice, and that increasing the bioavailability of SPMs, such as MaR1, minimizes inflammation and mediates therapeutic actions. Thus, these data suggest that immunoresolvent therapies, such as MaR1, could be a novel avenue for the treatment of MS.
Activated leukocyte cell adhesion molecule (ALCAM) is a cell adhesion molecule found on blood–brain barrier endothelial cells (BBB-ECs) that was previously shown to be involved in leukocyte ...transmigration across the endothelium. In the present study, we found that ALCAM knockout (KO) mice developed a more severe myelin oligodendrocyte glycoprotein (MOG)35–55–induced experimental autoimmune encephalomyelitis (EAE). The exacerbated disease was associated with a significant increase in the number of CNS-infiltrating proinflammatory leukocytes compared with WT controls. Passive EAE transfer experiments suggested that the pathophysiology observed in active EAE was linked to the absence of ALCAM on BBB-ECs. In addition, phenotypic characterization of unimmunized ALCAM KO mice revealed a reduced expression of BBB junctional proteins. Further in vivo, in vitro, and molecular analysis confirmed that ALCAM is associated with tight junction molecule assembly at the BBB, explaining the increased permeability of CNS blood vessels in ALCAM KO animals. Collectively, our data point to a biologically important function of ALCAM in maintaining BBB integrity.
Multiple sclerosis (MS) is an autoimmune and neurodegenerative disease driven by inflammation and demyelination in the brain, spinal cord, and optic nerve. Optic neuritis, characterized by ...inflammation and demyelination of the optic nerve, is a symptom in many patients with MS. The optic nerve is the highway for visual information transmitted from the retina to the brain. It contains axons from the retinal ganglion cells (RGCs) that reside in the retina, myelin forming oligodendrocytes and resident microglia and astrocytes. Inflammation, demyelination, and axonal degeneration are also present in the optic nerve of mice subjected to experimental autoimmune encephalomyelitis (EAE), a preclinical mouse model of MS. Monitoring the optic nerve in EAE is a useful strategy to study the presentation and progression of pathology in the visual system; however, current approaches have relied on sectioning, staining and manual quantification. Further, information regarding the spatial load of lesions and inflammation is dependent on the area of sectioning. To better characterize cellular pathology in the EAE model, we employed a tissue clearing and 3D immunolabelling and imaging protocol to observe patterns of immune cell infiltration and activation throughout the optic nerve. Increased density of TOPRO staining for nuclei captured immune cell infiltration and Iba1 immunostaining was employed to monitor microglia and macrophages. Axonal degeneration was monitored by neurofilament immunolabelling to reveal axonal swellings throughout the optic nerve. In parallel, we developed a convolutional neural network with a UNet architecture (CNN-UNet) called BlebNet for automated identification and quantification of axonal swellings in whole mount optic nerves. Together this constitutes a toolkit for 3-dimensional immunostaining to monitor general optic nerve pathology and fast automated quantification of axonal defects that could also be adapted to monitor axonal degeneration and inflammation in other neurodegenerative disease models.
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