Homogeneous antibody–angiopep 2 conjugates for effective brain targeting

Antibody-based therapy has shown great success in the treatment of many diseases, including cancers. While antibodies and antibody–drug conjugates (ADCs) have also been evaluated for central nervous system (CNS) disorders as well as brain tumors, their therapeutic efficacy can be substantially limited due to low permeability across the blood–brain barrier (BBB). Thus, improving BBB permeability of therapeutic antibodies is critical in establishing this drug class as a reliable clinical option for CNS diseases. Here, we report that, compared with a conventional heterogeneous conjugation, homogeneous conjugation of the synthetic BBB shuttle peptide angiopep-2 (Ang2) to a monoclonal antibody (mAb) provides improved binding affinity for brain microvascular endothelial cells in vitro and accumulation into normal brain tissues in vivo. In a mouse model, we also demonstrate that the homogeneous anti-EGFR mAb–Ang2 conjugate administered intravenously efficiently accumulates in intracranial tumors. These findings suggest that homogeneous conjugation of BBB shuttle peptides such as Ang2 is a promising approach to enhancing the therapeutic efficacy of antibody agents for CNS diseases.

Homogeneous conjugation of angiopep-2 to a monoclonal antibody improves binding affinity for brain microvascular endothelial cells and accumulation into brain tissues and tumors across the BBB.     

Intranasal Delivery of Mesenchymal Stem Cells Significantly Extends Survival of Irradiated Mice with Experimental Brain Tumors

Treatment options of glioblastoma multiforme are limited due to the blood–brain barrier (BBB). In this study, we investigated the utility of intranasal (IN) delivery as a means of transporting stem cell–based antiglioma therapeutics. We hypothesized that mesenchymal stem cells (MSCs) delivered via nasal application could impart therapeutic efficacy when expressing TNF-related apoptosis-inducing ligand (TRAIL) in a model of human glioma. ¹¹¹In-oxine, histology and magnetic resonance imaging (MRI) were utilized to track MSCs within the brain and associated tumor. We demonstrate that MSCs can penetrate the brain from nasal cavity and infiltrate intracranial glioma xenografts in a mouse model. Furthermore, irradiation of tumor-bearing mice tripled the penetration of ^111In-oxine–labeled MSCs in the brain with a fivefold increase in cerebellum. Significant increase in CXCL12 expression was observed in irradiated xenograft tissue, implicating a CXCL12-dependent mechanism of MSCs migration towards irradiated glioma xenografts. Finally, MSCs expressing TRAIL improved the median survival of irradiated mice bearing intracranial U87 glioma xenografts in comparison with nonirradiated and irradiated control mice. Cumulatively, our data suggest that IN delivery of stem cell–based therapeutics is a feasible and highly efficacious treatment modality, allowing for repeated application of modified stem cells to target malignant glioma.     

Plasmodium falciparum erythrocyte membrane protein 1 variants induce cell swelling and disrupt the blood–brain barrier in cerebral malaria

Cerebral malaria (CM) is caused by the binding of Plasmodium falciparum–infected erythrocytes (IEs) to the brain microvasculature, leading to inflammation, vessel occlusion, and cerebral swelling. We have previously linked dual intercellular adhesion molecule-1 (ICAM-1)– and endothelial protein C receptor (EPCR)–binding P. falciparum parasites to these symptoms, but the mechanism driving the pathogenesis has not been identified. Here, we used a 3D spheroid model of the blood–brain barrier (BBB) to determine unexpected new features of IEs expressing the dual-receptor binding PfEMP1 parasite proteins. Analysis of multiple parasite lines shows that IEs are taken up by brain endothelial cells in an ICAM-1–dependent manner, resulting in breakdown of the BBB and swelling of the endothelial cells. Via ex vivo analysis of postmortem tissue samples from CM patients, we confirmed the presence of parasites within brain endothelial cells. Importantly, this discovery points to parasite ingress into the brain endothelium as a contributing factor to the pathology of human CM.     

Impact of gut–brain interaction in emerging neurological disorders

The central nervous system (CNS) is a reservoir of immune privilege. Specialized immune glial cells are responsible for maintenance and defense against foreign invaders. The blood–brain barrier (BBB) prevents detrimental pathogens and potentially overreactive immune cells from entering the periphery. When the double-edged neuroinflammatory response is overloaded, it no longer has the protective function of promoting neuroregeneration. Notably, microbiota and its derivatives may emerge as pathogen-associated molecular patterns of brain pathology, causing microbiome–gut–brain axis dysregulation from the bottom-up. When dysbiosis of the gastrointestinal flora leads to subsequent alterations in BBB permeability, peripheral immune cells are recruited to the brain. This results in amplification of neuroinflammatory circuits in the brain, which eventually leads to specific neurological disorders. Aggressive treatment strategies for gastrointestinal disorders may protect against specific immune responses to gastrointestinal disorders, which can lead to potential protective effects in the CNS. Accordingly, this study investigated the mutual effects of microbiota and the gut–brain axis, which may provide targeting strategies for future disease treatment.     

Endothelial LRP1 protects against neurodegeneration by blocking cyclophilin A

The low-density lipoprotein receptor–related protein 1 (LRP1) is an endocytic and cell signaling transmembrane protein. Endothelial LRP1 clears proteinaceous toxins at the blood–brain barrier (BBB), regulates angiogenesis, and is increasingly reduced in Alzheimer’s disease associated with BBB breakdown and neurodegeneration. Whether loss of endothelial LRP1 plays a direct causative role in BBB breakdown and neurodegenerative changes remains elusive. Here, we show that LRP1 inactivation from the mouse endothelium results in progressive BBB breakdown, followed by neuron loss and cognitive deficits, which is reversible by endothelial-specific LRP1 gene therapy. LRP1 endothelial knockout led to a self-autonomous activation of the cyclophilin A–matrix metalloproteinase-9 pathway in the endothelium, causing loss of tight junctions underlying structural BBB impairment. Cyclophilin A inhibition in mice with endothelial-specific LRP1 knockout restored BBB integrity and reversed and prevented neuronal loss and behavioral deficits. Thus, endothelial LRP1 protects against neurodegeneration by inhibiting cyclophilin A, which has implications for the pathophysiology and treatment of neurodegeneration linked to vascular dysfunction.     

Sonodynamic therapy induces oxidative stress,DNA damage and apoptosis in glioma cells

Malignant glioma remains one of the most challenging diseases to treat because of the invasive growth of glioma cells and the existence of the blood–brain barrier (BBB), which blocks drug delivery to the brain. New strategies are urgently needed to overcome these shortcomings and improve the outcomes. Ultrasound represents a promising noninvasive and reversible BBB opening approach and the related sonodynamic therapy (SDT) is rapidly emerging. This study aims to explore the ultrasound parameters for BBB opening and the cell killing effect of SDT in human glioma U373 cells by using a recently reported sonosensitizer, sinoporphyrin sodium (DVDMS). The in vitro BBB model indicated that SDT caused a time-dependent permeability increase, which peaked at 2 h post treatment and then recovered gradually. The results of toxicology tests showed significant U373 cell viability loss and apoptosis increase after DVDMS-SDT, accompanied by enhanced cleaved-caspase-3 level and DNA fragmentation, in which reactive oxygen species (ROS) were a major triggering intermediate during DVDMS-SDT. Furthermore, DVDMS-SDT produced DNA damage and the underlying mechanisms were evaluated, in order to provide a fundamental basis for DVDMS-SDT application in glioma therapy. The findings indicated that the DNA molecules could be temporarily regulated by SDT and DNA double-strand breaks (DSBs), which increased the difficulty of cellular self-repair, thus aggravating cell apoptosis and inhibiting glioma cell invasive growth. Therefore, this study supports the use of SDT as an alternative approach for glioma therapy.

Malignant glioma remains one of the most challenging diseases to treat because of the invasive growth of glioma cells and the existence of the blood–brain barrier (BBB), which blocks drug delivery to the brain.     

Inhibition of metal-induced amyloid β-peptide aggregation by a blood–brain barrier permeable silica–cyclen nanochelator

Alzheimer''s disease (AD) is a neurodegenerative malady associated with amyloid β-peptide (Aβ) aggregation in the brain. Metal ions play important roles in Aβ aggregation and neurotoxicity. Metal chelators are potential therapeutic agents for AD because they could sequester metal ions from the Aβ aggregates and reverse the aggregation. The blood–brain barrier (BBB) is a major obstacle for drug delivery to AD patients. Herein, a nanoscale silica–cyclen composite combining cyclen as the metal chelator and silica nanoparticles as a carrier was reported. Silica–cyclen was characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM), Fourier transform infrared (FT-IR) and dynamic light scattering (DLS). The inhibitory effect of the silica–cyclen nanochelator on Zn²⁺- or Cu²⁺-induced Aβ aggregation was investigated by using a BCA protein assay and TEM. Similar to cyclen, silica–cyclen can effectively inhibit the Aβ aggregation and reduce the generation of reactive oxygen species induced by the Cu–Aβ₄₀ complex, thereby lessening the metal-induced Aβ toxicity against PC12 cells. In vivo studies indicate that the silica–cyclen nanochelator can cross the BBB, which may provide inspiration for the construction of novel Aβ inhibitors.

A BBB-passable nanoscale silica–cyclen chelator effectively reduces the metal-induced Aβ aggregates and related ROS, thereby decreasing the neurotoxicity of Aβ.     

Quantitative protein expression of blood‐brain barrier transporters in the vasculature of brain metastases of patients with lung and breast cancer

This study determined absolute transporter protein abundances in isolated microvessels of human noncancerous cerebral cortex as well as brain metastases of patients with lung and breast cancer, using a validated targeted proteomics approach. As compared with those in microvessels of noncancerous cerebral cortex, the median protein abundances of glucose transporter 1 (a brain endothelial marker) and sodium‐potassium pump (Na/K ATPase, a plasma membrane marker) were decreased by ~ 80% in brain metastasis microvessels. The major efflux transporters (ABCB1 and ABCG2) fell to undetectable in microvessels of more than 80% of brain metastasis specimens. Monocarboxylate transporter 1 was undetectable in microvessels of more than 80% of brain metastases, whereas large neutral amino acid transporter 1 levels remained unchanged. In conclusion, the integrity of the physical and biochemical barrier with respect to transporter protein expression is largely disrupted in brain metastasis tumor vasculatures. Differential transporter protein abundances at the blood‐brain barrier and blood‐brain tumor barrier provided mechanistic and quantitative basis for prediction of heterogeneous drug penetration into human brain and brain tumors, which is critical not only to the understanding of the success or failure of systemic chemotherapy in the treatment of brain tumors but also to the development of more effective therapeutic strategies.

Study Highlights

• WHAT IS THE CURRENT KNOWLEDGE ON THE TOPIC?

Systemic chemotherapy for brain metastases is controversial, at least partly due to the incomplete understanding of the blood‐brain barrier (BBB) and blood‐brain tumor barrier (BBTB) with respect to transporter protein expression and function.

• WHAT QUESTION DID THIS STUDY ADDRESS?

This study determined the absolute protein abundances of major BBB transporters in isolated microvessels of brain metastases from lung and breast cancer patients as well as non‐cancerous brain cortex from primary or metastatic brain cancer patients.

• WHAT DOES THIS STUDY ADD TO OUR KNOWLEDGE?

This study complemented our previous knowledge on transporter expression in microvessels of human normal brain cortex and glioblastoma, which collectively provided a comprehensive dataset on the quantitative protein expression of BBB transporters in the vasculatures of human normal brain, primary and metastatic brain tumors.

• HOW MIGHT THIS CHANGE CLINICAL PHARMACOLOGY OR TRANSLATIONAL SCIENCE?

Differential transporter protein abundances at the BBB and BBTB provided mechanistic and quantitative basis for prediction of heterogeneous drug penetration into human brain and brain tumors, which is critical not only to the understanding of the success or failure of systemic chemotherapy in the treatment of brain tumors but also to the development of more effective therapeutic strategies.     

Oral Delivery of Bioencapsulated Proteins Across Blood–Brain and Blood–Retinal Barriers

Delivering neurotherapeutics to target brain-associated diseases is a major challenge. Therefore, we investigated oral delivery of green fluorescence protein (GFP) or myelin basic protein (MBP) fused with the transmucosal carrier cholera toxin B subunit (CTB), expressed in chloroplasts (bioencapsulated within plant cells) to the brain and retinae of triple transgenic Alzheimer''s disease (3×TgAD) mice, across the blood–brain barriers (BBB) and blood–retinal barriers (BRB). Human neuroblastoma cells internalized GFP when incubated with CTB-GFP but not with GFP alone. Oral delivery of CTB-MBP in healthy and 3×TgAD mice shows increased MBP levels in different regions of the brain, crossing intact BBB. Thioflavin S–stained amyloid plaque intensity was reduced up to 60% by CTB-MBP incubation with human AD and 3×TgAD mice brain sections ex vivo. Amyloid loads were reduced in vivo by 70% in hippocampus and cortex brain regions of 3×TgAD mice fed with bioencapsulated CTB-MBP, along with reduction in the ratio of insoluble amyloid β 42 (Aβ₄₂) to soluble fractions. CTB-MBP oral delivery reduced Aβ₄₂ accumulation in retinae and prevented loss of retinal ganglion cells in 3×TgAD mice. Lyophilization of leaves increased CTB-MBP concentration by 17-fold and stabilized it during long-term storage in capsules, facilitating low-cost oral delivery of therapeutic proteins across the BBB and BRB.     

Effective Intravenous Therapy for Neurodegenerative Disease With a Therapeutic Enzyme and a Peptide That Mediates Delivery to the Brain

The blood–brain barrier (BBB) presents a major challenge to effective treatment of neurological disorders, including lysosomal storage diseases (LSDs), which frequently present with life-shortening and untreatable neurodegeneration. There is considerable interest in methods for intravenous delivery of lysosomal proteins across the BBB but for the most part, levels achievable in the brain of mouse models are modest and increased lifespan remains to be demonstrated. In this study, we have investigated delivery across the BBB using a mouse model of late-infantile neuronal ceroid lipofuscinosis (LINCL), a neurodegenerative LSD caused by loss of tripeptidyl peptidase I (TPP1). We have achieved supraphysiological levels of TPP1 throughout the brain of LINCL mice by intravenous (IV) coadministration of recombinant TPP1 with a 36-residue peptide that contains polylysine and a low-density lipoprotein receptor binding sequence from apolipoprotein E. Importantly, IV administration of TPP1 with the peptide significantly reduces brain lysosomal storage, increases lifespan and improves neurological function. This simple “mix and inject” method is immediately applicable towards evaluation of enzyme replacement therapy to the brain in preclinical models and further exploration of its clinical potential is warranted.     

Molecular architecture determines brain delivery of a transferrin receptor–targeted lysosomal enzyme

Delivery of biotherapeutics across the blood–brain barrier (BBB) is a challenge. Many approaches fuse biotherapeutics to platforms that bind the transferrin receptor (TfR), a brain endothelial cell target, to facilitate receptor-mediated transcytosis across the BBB. Here, we characterized the pharmacological behavior of two distinct TfR-targeted platforms fused to iduronate 2-sulfatase (IDS), a lysosomal enzyme deficient in mucopolysaccharidosis type II (MPS II), and compared the relative brain exposures and functional activities of both approaches in mouse models. IDS fused to a moderate-affinity, monovalent TfR-binding enzyme transport vehicle (ETV:IDS) resulted in widespread brain exposure, internalization by parenchymal cells, and significant substrate reduction in the CNS of an MPS II mouse model. In contrast, IDS fused to a standard high-affinity bivalent antibody (IgG:IDS) resulted in lower brain uptake, limited biodistribution beyond brain endothelial cells, and reduced brain substrate reduction. These results highlight important features likely to impact the clinical development of TfR-targeting platforms in MPS II and potentially other CNS diseases.     

Surface Characteristics of Nanoparticles Determine Their Intracellular Fate in and Processing by Human Blood–Brain Barrier Endothelial Cells In Vitro

A polarized layer of endothelial cells that comprises the blood–brain barrier (BBB) precludes access of systemically administered medicines to brain tissue. Consequently, there is a need for drug delivery vehicles that mediate transendothelial transport of such medicines. Endothelial cells use a variety of endocytotic pathways for the internalization of exogenous materials, including clathrin-mediated endocytosis, caveolar endocytosis, and macropinocytosis. The different modes of endocytosis result in the delivery of endocytosed material to distinctive intracellular compartments and therewith correlated differential processing. To obtain insight into the properties of drug delivery vehicles that direct their intracellular processing in brain endothelial cells, we investigated the intracellular processing of fixed-size nanoparticles in an in vitro BBB model as a function of distinct nanoparticle surface modifications. Caveolar endocytosis, adsorptive-mediated endocytosis, and receptor-mediated endocytosis were promoted by the use of uncoated 500-nm particles, attachment of the cationic polymer polyethyleneimine (PEI), and attachment of prion proteins, respectively. We demonstrate that surface modifications of nanoparticles, including charge and protein ligands, affect their mode of internalization by brain endothelial cells and thereby their subcellular fate and transcytotic potential.     

Dynamic in vivo imaging of cerebral blood flow and blood–brain barrier permeability

The brain is characterized by an extremely rich blood supply, regulated by changes in blood vessel diameter and blood flow, depending on metabolic demands. The blood–brain barrier (BBB)—a functional and structural barrier separating the intravascular and neuropil compartments—characterizes the brain's vascular bed and is essential for normal brain functions. Disruptions to the regional cerebral blood supply, to blood drainage and to BBB properties have been described in most common neurological disorders, but there is a lack of quantitative methods for assessing blood flow dynamics and BBB permeability in small blood vessels under both physiological and pathological conditions. Here, we present a quantitative image analysis approach that allows the characterization of relative changes in the regional cerebral blood flow (rCBF) and BBB properties in small surface cortical vessels. In experiments conducted using the open window technique in rats, a fluorescent tracer was injected into the tail vein, and images of the small vessels at the surface of the cortex were taken using a fast CCD camera. Pixel-based image analysis included registration and characterization of the changes in fluorescent intensity, followed by cluster analysis. This analysis enabled the characterization of rCBF in small arterioles and venules and changes in BBB permeability. The method was implemented successfully under experimental conditions, including increased rCBF induced by neural stimulation, bile salt-induced BBB breakdown, and photothrombosis-mediated local ischemia. The new approach may be used to study changes in rCBF, neurovascular coupling and BBB permeability under normal and pathological brain conditions.     

Complement C3a receptor–mediated vascular dysfunction: a complex interplay between aging and neurodegeneration

Vascular dysfunction resulting in compromised blood-brain barrier (BBB) integrity is evident in aging and disease. Although the complement C3a/C3a receptor (C3a/C3aR) axis influences normal brain aging and disease progression, the mechanisms governing endothelial C3aR–mediated neurovascular inflammation and BBB permeability remain unexplored. In this issue of the JCI, Propson et al. investigated endothelial C3a/C3aR signaling in normal, aged, and neurodegenerative mouse models. Endothelial C3aR signaling modulated age-dependent increases in VCAM1, initiated peripheral lymphocyte infiltration, and enhanced microglial activity. Increased calcium release downstream of C3aR signaling disrupted the vascular endothelial cadherin (VE-cadherin) junctions, increased BBB permeability, and degraded vascular structure and function. Mice lacking C3aR (C3ar1^–/–) and mice treated with a C3aR antagonist showed attenuated age-related microglial reactivity and neurodegeneration. These results confirm that complement-mediated signaling impacts vascular health and BBB function in normal aging and neurodegenerative disease, suggesting that complement inhibitors represent a therapeutic option for cerebral microvascular dysfunction.     

Endothelial LRP1 transports amyloid-β1–42 across the blood-brain barrier

According to the neurovascular hypothesis, impairment of low-density lipoprotein receptor–related protein-1 (LRP1) in brain capillaries of the blood-brain barrier (BBB) contributes to neurotoxic amyloid-β (Aβ) brain accumulation and drives Alzheimer’s disease (AD) pathology. However, due to conflicting reports on the involvement of LRP1 in Aβ transport and the expression of LRP1 in brain endothelium, the role of LRP1 at the BBB is uncertain. As global Lrp1 deletion in mice is lethal, appropriate models to study the function of LRP1 are lacking. Moreover, the relevance of systemic Aβ clearance to AD pathology remains unclear, as no BBB-specific knockout models have been available. Here, we developed transgenic mouse strains that allow for tamoxifen-inducible deletion of Lrp1 specifically within brain endothelial cells (Slco1c1-CreER^T2 Lrp1^fl/fl mice) and used these mice to accurately evaluate LRP1-mediated Aβ BBB clearance in vivo. Selective deletion of Lrp1 in the brain endothelium of C57BL/6 mice strongly reduced brain efflux of injected [¹²⁵I] Aβ_1–42. Additionally, in the 5xFAD mouse model of AD, brain endothelial–specific Lrp1 deletion reduced plasma Aβ levels and elevated soluble brain Aβ, leading to aggravated spatial learning and memory deficits, thus emphasizing the importance of systemic Aβ elimination via the BBB. Together, our results suggest that receptor-mediated Aβ BBB clearance may be a potential target for treatment and prevention of Aβ brain accumulation in AD.     

<Emphasis Type="Italic">In Vitro</Emphasis> and Initial <Emphasis Type="Italic">In Vivo</Emphasis> Evaluation of <Superscript>68</Superscript>Ga-Labeled Transferrin Receptor (TfR) Binding Peptides as Potential Carriers for Enhanced Drug Transport into TfR Expressing Cells

Purpose  

The transferrin receptor (TfR) is one of the most attractive targets to overcome the blood–brain barrier (BBB). It has recently been shown that THRPPMWSPVWP binds to the TfR and is subsequently internalized into TfR-expressing cells. Here, we evaluated the ability of THRPPMWSPVWP to become internalized into human TfR-expressing cells via endocytosis to determine its potential to act as a carrier system for the transport of small molecules across the BBB.     

Bench-to-bedside review: Burn-induced cerebral inflammation – a neglected entity?

Severe burn injury remains a major burden on patients and healthcare systems. Following severe burns, the injured tissues mount a local inflammatory response aiming to restore homeostasis. With excessive burn load, the immune response becomes disproportionate and patients may develop an overshooting systemic inflammatory response, compromising multiple physiological barriers in the lung, kidney, liver, and brain. If the blood–brain barrier is breached, systemic inflammatory molecules and phagocytes readily enter the brain and activate sessile cells of the central nervous system. Copious amounts of reactive oxygen species, reactive nitrogen species, proteases, cytokines/chemokines, and complement proteins are being released by these inflammatory cells, resulting in additional neuronal damage and life-threatening cerebral edema. Despite the correlation between cerebral complications in severe burn victims with mortality, burn-induced neuroinflammation continues to fly under the radar as an underestimated entity in the critically ill burn patient. In this paper, we illustrate the molecular events leading to blood–brain barrier breakdown, with a focus on the subsequent neuroinflammatory changes leading to cerebral edema in patients with severe burns.     

IL-17 crosses the blood–brain barrier to trigger neuroinflammation: a novel mechanism in nitroglycerin-induced chronic migraine

BackgroundChronic migraine places a disabling burden on patients, which is extensively modeled by the nitroglycerin (NTG)-treated animal model. Although the NF-κB pathway is involved in an increase in CGRP levels and activation of the trigeminal system in the NTG model, the relationship between NTG and neuroinflammation remains unclear. This study aimed to optimize a chronic NTG rat model with hyperalgesia and the ethological capacity for estimating migraine therapies and to further explore the underlying mechanism of NTG-induced migraine.MethodsRats were administered different doses of NTG s.c. daily or every 2 d; 30 min and 2 h later, the mechanical threshold was tested. After 9 d, the rats were injected with EB or Cy5.5 for the permeability assay. The other animals were sacrificed, and then, brainstem and caudal trigeminal ganglion were removed to test CGRP, c-Fos and NOS activity; Cytokines levels in the tissue and serum were measured by ELISA; and NF-κB pathway and blood–brain barrier (BBB)-related indicators were analyzed using western blotting. Immunohistochemistry was performed to observe microglial polarization and IL-17A⁺ T cell migration in the medulla oblongata.ResultsNTG (10 mg/kg, s.c., every 2 d for a total of 5 injections) was the optimal condition, resulting in progressive hyperalgesia and migraine behavior. TNC neuroinflammation with increases in cytokines, CGRP and c-Fos and activation of the NF-κB pathway was observed, and these changes were alleviated by ibuprofen. Furthermore, NTG administration increased BBB permeability by altering the levels functional proteins (RAGE, LRP1, AQP4 and MFSD2A) and structural proteins (ZO-1, Occludin and VE-cadherin-2) to increase peripheral IL-17A permeation into the medulla oblongata, activating microglia and neuroinflammation, and eventually causing hyperalgesia and migraine attack.ConclusionsThis study confirmed that NTG (10 mg/kg, s.c., every 2 d for a total of 5 injections) was the optimal condition to provoke migraine, resulting in mechanical hyperalgesia and observable migraine-like behavior. Furthermore, IL-17A crossed the blood–brain barrier into the medulla oblongata, triggering TNC activation through microglia-mediated neuroinflammation. This process was a novel mechanism in NTG-induced chronic migraine, suggesting that IL-17A might be a novel target in the treatment of migraine.     

A first-principles study on potential chelation agents and indicators of Alzheimer's disease

Human-serum transferrin is involved in the transportation of aluminum across the blood–brain barrier. Aluminum accumulation within the neuron causes the cell to degrade. In our research, we considered 12 potential chelators of aluminum from the aluminum–human serum transferrin complex and 3 potential indicators of Alzheimer''s. We performed Density Functional Theory calculations comparing the binding energies of aluminum–chelator complexes and the binding energy of the aluminum–human serum transferrin complex and determined the charge transfer of the aluminum–chelator complex. Our results showed that CDTA is the only one that has direct chelation potential, but 1-ethyl-3-hydroxypyridin-2-one, citric acid, DTPA, oxalic acid, and salicylhydroxamic acid also had a strong and stable bond with aluminum and still have the ability to be potential chelators. The charge transfer calculation further enforces that these 6 chelators have strong and stable bonds with aluminum. Furthermore, we evaluated potential indicators of Alzheimer''s disease. Metals that have a similar binding affinity to human serum transferrin as that of iron prove to be potential indicators of Alzheimer''s disease. Due to the minimal difference in binding energies of the gallium–human serum transferrin complex and the indium–human serum transferrin complex to the iron–human serum transferrin complex, we determined that gallium and indium could be potential indicators of Alzheimer''s disease.

Human-serum transferrin is involved in the transportation of aluminum across the blood–brain barrier.     

The surface-anchored NanA protein promotes pneumococcal brain endothelial cell invasion

In humans, Streptococcus pneumoniae (SPN) is the leading cause of bacterial meningitis, a disease with high attributable mortality and frequent permanent neurological sequelae. The molecular mechanisms underlying the central nervous system tropism of SPN are incompletely understood, but include a primary interaction of the pathogen with the blood–brain barrier (BBB) endothelium. All SPN strains possess a gene encoding the surface-anchored sialidase (neuraminidase) NanA, which cleaves sialic acid on host cells and proteins. Here, we use an isogenic SPN NanA-deficient mutant and heterologous expression of the protein to show that NanA is both necessary and sufficient to promote SPN adherence to and invasion of human brain microvascular endothelial cells (hBMECs). NanA-mediated hBMEC invasion depends only partially on sialidase activity, whereas the N-terminal lectinlike domain of the protein plays a critical role. NanA promotes SPN–BBB interaction in a murine infection model, identifying the protein as proximal mediator of CNS entry by the pathogen.Streptococcus pneumoniae (SPN; pneumococcus) accounts for 50% of bacterial meningitis in humans. This often devastating infection carries a 30% mortality rate, and up to half of survivors experience neurological sequelae reflecting a wide spectrum of brain injuries, including cortical neuronal necrosis and hippocampal neuronal apoptosis (Koedel et al., 2002; van de Beek et al., 2004; Weisfelt et al., 2006; Weber and Tuomanen, 2007). To cause meningitis, bloodborne bacteria must first attach to and penetrate human brain microvascular endothelial cells (hBMECs), which is the single-cell layer comprising the majority of the blood–brain barrier (BBB; Tuomanen, 1996; Kim, 2003). The molecular mechanisms underlying the distinct central nervous system (CNS) tropism of SPN are poorly understood. All SPN clinical isolates express the surface-anchored sialidase (neuraminidase) NanA (Cámara et al., 1994; Pettigrew et al., 2006) that targets sialic acid residues on host cells and bacterial flora to promote SPN mucosal colonization (Shakhnovich et al., 2002; Tong et al., 2002). Here, we use an isogenic SPN NanA-deficient mutant and heterologous expression of the NanA constructs to probe the contribution of this protein to SPN-hBMEC interactions in vitro and BBB penetration in vivo.