Littérature scientifique sur le sujet « Blood Brain Barriers »

The blood-brain barrier (BBB), blood-cerebrospinal fluid (CSF) barrier (BCSFB), and CSF-brain barriers (CSFBB) are highly regulated barriers in the central nervous system comprising complex multicellular structures that separate nerves and glia from blood and CSF, respectively. Barrier damage has been implicated in the pathophysiology of diverse hypoxia-related neurological conditions, including stroke, multiple sclerosis, hydrocephalus, and high-altitude cerebral edema. Much is known about the damage to the BBB in response to hypoxia, but much less is known about the BCSFB and CSFBB. Yet, it is known that these other barriers are implicated in damage after hypoxia or inflammation. In the 1950s, it was shown that the rate of radionucleated human serum albumin passage from plasma to CSF was five times higher during hypoxic than normoxic conditions in dogs, due to BCSFB disruption. Severe hypoxia due to administration of the bacterial toxin lipopolysaccharide is associated with disruption of the CSFBB. This review discusses the anatomy of the BBB, BCSFB, and CSFBB and the impact of hypoxia and associated inflammation on the regulation of those barriers.

Glioblastoma (GBM) is the most common primary brain tumor and carries a grave prognosis. Despite years of research investigating potentially new therapies for GBM, the median survival rate of individuals with this disease has remained fairly stagnant. Delivery of drugs to the tumor site is hampered by various barriers posed by the GBM pathological process and by the complex physiology of the blood-brain and blood–cerebrospinal fluid barriers. These anatomical and physiological barriers serve as a natural protection for the brain and preserve brain homeostasis, but they also have significantly limited the reach of intraparenchymal treatments in patients with GBM. In this article, the authors review the functional capabilities of the physical and physiological barriers that impede chemotherapy for GBM, with a specific focus on the pathological alterations of the blood-brain barrier (BBB) in this disease. They also provide an overview of current and future methods for circumventing these barriers in therapeutic interventions. Although ongoing research has yielded some potential options for future GBM therapies, delivery of chemotherapy medications across the BBB remains elusive and has limited the efficacy of these medications.

The present report encompasses a thorough review of drug delivery to the brain with a particular focus on using drug carriers such as liposomes and nanoparticles. Challenges in brain drug delivery arise from the presence of one of the strictest barriers in vivo—the blood-brain barrier (BBB). This barrier exists at the level of endothelial cells of brain vasculature and its role is to maintain brain homeostasis. To better understand the principles of brain drug delivery, relevant knowledge of the blood-brain barrier anatomy and physiology is briefly reviewed. Several approaches to overcome the BBB have been reviewed including the use of carrier systems. In addition, strategies to enhance brain drug delivery by specific brain targeting are discussed.

Infections of the central nervous system (CNS) are still a major cause of morbidity and mortality worldwide. Traversal of the barriers protecting the brain by pathogens is a prerequisite for the development of meningitis. Bacteria have developed a variety of different strategies to cross these barriers and reach the CNS. To this end, they use a variety of different virulence factors that enable them to attach to and traverse these barriers. These virulence factors mediate adhesion to and invasion into host cells, intracellular survival, induction of host cell signaling and inflammatory response, and affect barrier function. While some of these mechanisms differ, others are shared by multiple pathogens. Further understanding of these processes, with special emphasis on the difference between the blood–brain barrier and the blood–cerebrospinal fluid barrier, as well as virulence factors used by the pathogens, is still needed.

Manganese (Mn) is a trace nutrient necessary for life but becomes neurotoxic at high concentrations in the brain. The brain is a “privileged” organ that is separated from systemic blood circulation mainly by two barriers. Endothelial cells within the brain form tight junctions and act as the blood–brain barrier (BBB), which physically separates circulating blood from the brain parenchyma. Between the blood and the cerebrospinal fluid (CSF) is the choroid plexus (CP), which is a tissue that acts as the blood–CSF barrier (BCB). Pharmaceuticals, proteins, and metals in the systemic circulation are unable to reach the brain and spinal cord unless transported through either of the two brain barriers. The BBB and the BCB consist of tightly connected cells that fulfill the critical role of neuroprotection and control the exchange of materials between the brain environment and blood circulation. Many recent publications provide insights into Mn transport in vivo or in cell models. In this review, we will focus on the current research regarding Mn metabolism in the brain and discuss the potential roles of the BBB and BCB in maintaining brain Mn homeostasis.

The presence of barriers, such as the blood–brain barrier (BBB) and brain–tumor barrier (BTB), limits the penetration of antineoplastic drugs into the brain, resulting in poor response to treatments. Many techniques have been developed to overcome the presence of these barriers, including direct injections of substances by intranasal or intrathecal routes, chemical modification of drugs or constituents of BBB, inhibition of efflux pumps, physical disruption of BBB by radiofrequency electromagnetic radiation (EMP), laser-induced thermal therapy (LITT), focused ultrasounds (FUS) combined with microbubbles and convection enhanced delivery (CED). However, most of these strategies have been tested only in preclinical models or in phase 1–2 trials, and none of them have been approved for treatment of brain tumors yet. Concerning the treatment of brain metastases, many molecules have been developed in the last years with a better penetration across BBB (new generation tyrosine kinase inhibitors like osimertinib for non-small-cell lung carcinoma and neratinib/tucatinib for breast cancer), resulting in better progression-free survival and overall survival compared to older molecules. Promising studies concerning neural stem cells, CAR-T (chimeric antigen receptors) strategies and immunotherapy with checkpoint inhibitors are ongoing.

The homeostasis of the central nervous system (CNS) is ensured by the endothelial, epithelial, mesothelial and glial brain barriers, which strictly control the passage of molecules, solutes and immune cells. While the endothelial blood-brain barrier (BBB) and the epithelial blood-cerebrospinal fluid barrier (BCSFB) have been extensively investigated, less is known about the epithelial and mesothelial arachnoid barrier and the glia limitans. Here, we summarize current knowledge of the cellular composition of the brain barriers with a specific focus on describing the molecular constituents of their junctional complexes. We propose that the brain barriers maintain CNS immune privilege by dividing the CNS into compartments that differ with regard to their role in immune surveillance of the CNS. We close by providing a brief overview on experimental tools allowing for reliable in vivo visualization of the brain barriers and their junctional complexes and thus the respective CNS compartments.

The passage of peptides across the blood-brain or blood-cerebrospinal fluid barrier is extremely limited. Peptides can be hindered from entering the central nervous system due to the hydrophilic nature of peptides and their susceptibility to enzymatic degradation by various peptidases. This limitation can be overcome through chemical modifications of opioid peptides with the goal of increasing biological stability and blood-central nervous system permeation. In the present studies, an in vitro bovine brain microvessel endothelial cell model of the blood-brain barrier was characterized both functionally and enzymatically. This primary culture model was found to be reflective of the in vivo blood-brain barrier in reference to predicting a peptides relative lipophilicity. Bovine brain microvessel endothelial cells were also found to be quite active enzymatically as far as the peptidases known to be involved in the degradation of methionine enkephalin. The conformationally stable analog of methionine enkephalin, DPDPE, was also characterized for its ability to enter the CNS using the in situ brain perfusion technique. DPDPE was found to enter the brain by both saturable and non-saturable uptake mechanisms. Chlorohalogenation was also found to significantly improve the central nervous system entry as well as biological stability of a potent opioid agonist, biphalin. In addition, the mu-opioid receptor selective antagonist, CTAP, was also evaluated for its ability to enter the CNS. The amount of CTAP that crossed both the blood-brain and blood-cerebrospinal fluid barrier was quantitatively comparable to the mu-selective agonist, morphine. Biphalin was found to enter both spinal and supra-spinal sites that have been shown previously to express mu and delta opioid receptors. In situ brain perfusion experiments identified a saturable component that contributes to the brain entry of [¹²⁵125I-Tyr¹]biphalin. Further experiments revealed that [¹²⁵125I-Tyr¹]biphalin was entering the CNS by the large neutral amino acid transporter and not by the leucine enkephalin uptake system or DPDPE transport system. This research has provided important preliminary work for the characterization of peptide transport into the brain. The importance of using neuropharmaceutical drug delivery vectors in modern medicine needs attention for the evolution of successful drug design targeted for CNS entry.

Thesis advisor: Cyril P. Opeil
The physiology of the vasculature in the central nervous system (CNS) which includes the blood-brain-barrier (BBB) and other factors, prevents the transport of most anticancer agents to the brain and restricts delivery to infiltrating brain tumors. The heterogeneous vascular permeability in tumor vessels (blood-tumor barrier; BTB), along with several other factors, creates additional hurdles for drug treatment of brain tumors. Different methods have been used to bypass the BBB/BTB, but they have their own limitations such as being invasive, non-targeted or requiring the formulation of new drugs. Magnetic Resonance Imaging guided Focused Ultrasound (MRIgFUS), when combined with circulating microbubbles, is an emerging noninvasive method to temporarily permeabilize the BBB and BTB. The purpose of this thesis was to use this alternative approach to deliver chemotherapeutic agents through the BBB/BTB for brain tumor treatment in a rodent model to overcome the hinderances encountered in prior approaches tested for drug delivery in the CNS. The results presented in thesis demonstrate that MRIgFUS can be used to achieve consistent and reproducible BBB/BTB disruption in rats. It enabled us to achieve clinically-relevant concentrations of doxorubicin (~ 4.8±0.5 µg/g) delivered to the brain with the sonication parameters (0.69 MHz; 0.55 MPa; 10 ms bursts; 1 Hz PRF; 60 s duration), microbubble concentration (Definity, 10 µl/kg), and liposomoal doxorubicin (Lipo-DOX) dose (5.67 mg/kg) used. The resulting doxorubicin concentration was reduced by 32% when the agent was injected 10 minute after the last sonication. Three weekly sessions of FUS and Lipo-DOX appeared to be safe in the rat brain, despite some minor tissue damage. Importantly, the severe neurotoxicity seen in earlier works using other approaches does not appear to occur with delivery via FUS-BBB disruption. The resuls from three weekly treatments of FUS and Lipo-DOX in a rat glioma model are highly promising since they demonstrated that the method significantly inhibits tumor growth and improves survival. Animals that received three weekly sessions of FUS + Lipo-DOX (N = 8) had a median survival time that was increased significantly (P<0.001) compared to animals who received Lipo-DOX only (N = 6), FUS only (N = 8), or no treatment (N = 7). Median survival for animals that received FUS + Lipo-DOX was increased by 100% relative to untreated controls, whereas animals who received Lipo-DOX alone had only a 16% improvement. Animals who received only FUS showed no improvement. No tumor cells were found in histology in 4/8 animals in the FUS + Lipo-DOX group, and only a few tumor cells were detected in two animals. Tumor doxorubicin concentrations increased monotonically (823±600, 1817±732 and 2432±448 ng/g) in the control tumors at 9, 14 and 17 days respectively after administration of Lipo-DOX. With FUS-induced BTB disruption, the doxorubicin concentrations were enhanced significantly (P<0.05, P<0.01, and P<0.0001 at days 9, 14, and 17, respectively) and were greater than the control tumors by a factor of two or more (2222±784, 3687±796 and 5658±821 ng/g) regardless of the stage of tumor growth. The transfer coefficient Ktrans was significantly (p<0.05) enhanced compared to control tumors only at day 9 but not at day 14 or 17. These results suggest that FUS-induced enhancements in tumor drug delivery for Lipo-DOX are relatively consistent over time, at least in this tumor model. These results are encouraging for the use of large drug carriers, as they suggest that even large/late-stage tumors can benefit from FUS-induced drug enhancement. Corresponding enhancements in Ktrans were found variable in large/late-stage tumors and not significantly different than controls, perhaps reflecting the size mismatch between the liposomal drug (~100 nm) and Gd-DTPA (molecular weight: 938 Da). Overall, this thesis research provides pre-clinical data toward the development of MRIgFUS as a noninvasive method for the delivery of agents such as Lipo-DOX across the BBB/BTB to treat patients with diseases of the central nervous system
Thesis (PhD) — Boston College, 2014
Submitted to: Boston College. Graduate School of Arts and Sciences
Discipline: Physics

2006/2007
INTRODUZIONE A basse concentrazioni la bilirubina non coniugata (unconjugated bilirubin, UCB) prodotta dalla degradazione dell’emoglobina, sembra essere un potente anti-ossidante, mentre è estremamente dannosa ad alte concentrazioni, causando encefalopatia nei neonati con severo ittero. Il 70% dei bambini che presentano kernittero muoiono entro sette giorni di vita, mentre il 30% dei sopravvissuti manifesta irreversibili conseguenze come sordità, ritardo mentale e danni cerebrali permanenti. L’encefalopatia dovuta ad alti livelli di bilirubina rappresenta oggi la maggior causa di riammissione ospedaliera nei neonati entro il primo mese di vita. Storicamente gli studi riguardanti le modalità di ingresso della bilirubina nel sistema nervoso centrale si sono concentrati sulla barriera emato-encefalica (blood brain barrier, BBB), costituita dai microvasi e dai capillari del cervello (micro vessels, MV). Tali studi hanno dimostrato come solamente la bilirubina non coniugata e non legata all’albumina del sangue, definita “bilirubina libera” (free bilirubin, Bf) sia capace di attraversare le membrane cellulari e diffondere nel tessuto. Tuttavia i microvasi non sono l’unica interfaccia sangue-tessuto presente nel cervello. Una seconda barriera è costituita dai plessi coroidei (CP). Questi, collocati nei ventricoli del cervello, mediano il passaggio delle molecole dal sangue al liquido cefalorachidiano e viceversa, posseggono una ampia superficie di scambio, il più alto flusso sanguigno del sistema nervoso centrale ed un fenotipo barriera meno restrittivo rispetto ai microvasi del parenchima. L’ingresso della bilirubina nel cervello sembra essere attivamente controllato da due trasportatori appartenenti alla famiglia delle “ATP dependent transporters”, Mrp1 e Pgp. Tali trasportatori potrebbero mantenere bassa la concentrazione della bilirubina limitandone l’ ingresso a livello di barriere o agendo direttamente a livello delle cellule del parenchima. Nonostante l’ impatto di questi trasportatori sulla disponibilità nel sistema nervoso centrale non solo della bilirubina ma egualmente di atre molecole potenzialmente tossiche, così come dei principi attivi, la loro espressione e localizzazione nelle interfacce sangue-cervello non sono del tutto chiare. Per tali motivi il lavoro di questi tre anni di tersi è stato incentrato a: Ia) chiarire il livello di espressione proteica relativa di Mrp1 e Pgp nelle due principali barriere cerebrali, la BBB (blood brain barrier, barriera emano encefalica) e la BCSFB (blood CerebroSpinal Fluid barrier, barriera emato liquorale); Ib) Definire l’andamento della loro espressione nel corso dello sviluppo post-natale in situazione fisiologica. II) Valutare l’effetto di elevati livelli serici di bilirubina sull’espressione di Mrp1 e Pgp nelle barriere emato encefaliche, come prima linea di difesa verso la bilirubina nel kernittero. Per raggiungere questo secondo obiettivo abbiamo utilizzato il ratto Gunn, considerato il modello in vivo per la sindrome di Crigler-Najjar e il kernittero. I ratti Gunn presentano elevati livelli di bilirubina serica ed un quadro clinico simile a quanto si riscontra nell’uomo. L’iperbilirubinemia, nel ratto, è dovuta ad una mutazione nell’enzima responsabile della coniugazione del pigmento, passaggio fondamentale per la sua successiva eliminazione. Nell’omozigote (jj) la bilirubina totale nel sangue (TBS) è molte volte più alta che nell’eterozigote (Jj) in cui l’allele non mutato codifica per l’enzima nella sua forma attiva, sufficiente a mantenere livelli di bilirubina normali. RISULTATI Ia) Attraverso una quantificazione relativa dell’ espressione proteica, ottenuta tramite Western blot, abbiamo dimostrato una espressione speculare dei due trasportatori nelle interacce sangue cervello. Mentre i microvasi sono caratterizzati dalla forte espressione di Pgp, ed i livelli di Mrp1 sono 15-20 volte inferiori rispetto ai plessi, questi ultimi presentano una elevata espressione di Mrp1 ed una quasi completa assenza di Pgp. Per quanto riguarda l’espressione di Mrp1 nei plessi coroidei (CP), abbiamo potuto evidenziare una differenza, con la massima espressione nel plesso del 4° ventricolo rispetto ai ventricoli laterali. Tramite immunofluorescenza abbiamo poi evidenziato per entrambe i trasportatori una localizzazione lato sangue, con Pgp luminale nei vasi e Mrp1 baso-laterale nel plessi coroidei. Ib) Anche l’andamento dell’espressione durane lo sviluppo post-natale differisce. Mentre Mrp1 è sin dalla nascita (2 giorni di vita) altamente espresso in entrambe le barriere, Pgp è inizialmente espresso a livelli più bassi (4,6 volte meno) rispetto all’ adulto (60 giorni). Contemporaneamente anche la densità dei vasi nel parenchima aumenta. II) Nel modello iperbilirubinemico rappresentato dal ratto Gunn, la TBS (jj) e molte volte più alta che nell’ eterozigote (Jj) e tale differenza permane per tutto l’arco di tempo esaminato (0-60 giorni dalla nascita). Al contrario la bilirubina libera (calcolata) è elevata solo nelle prime due settimane di vita, quando il rapporto bilirubina-albumina nel sangue è superiore all’unità. Poi, il rapido aumento della concentrazione ematica di albumina determina un significativo calo della Bf. Mentre l’analisi degli effetti (macroscopici) dell’iperbilirubinemia sullo sviluppo degli emisferi cerebrali non evidenzia differenze tra Jj e jj; in questi ultimi la crescita del cervelletto è severamente inibita. Già a 17 giorni di vita l’ipoplasia del cervelletto si manifesta con una differenza nel peso del 50% nei jj rispetto agli animali normo bilirubinemici di pari età. Durante tale periodo anche l’espressione dei due trasportatori nelle barriere è modificata. L’espressione proteica di Pgp nella BBB degli animali iperbilirubinemici è aumentata ad ogni età presa in esame. Tuttavia tale incremento non modifica in maniera importante la quantità del trasportatore nei MV durante lo sviluppo post natale, rimanendo quindi poco espresso (5 volte meno rispetto all’adulto) almeno fino ai 17 giorni di vita. Contemporaneamente la presenza Mrp1 nella BCSFB è inibita. Già a 9 giorni nel plesso del 4° ventricolo Mrp1 è il 50% rispetto al controllo (pari età, Jj). Anche se nei plessi dei ventricoli laterali l’inibizione dell’espressione è inferiore, nell’insieme la quantità di Mrp1 è fortemente ridotta negli animali iperbilirubinemici. Contrariamente, nei ratti Jj, Mrp1 ha un andamento simile a quello descritto nella sezione (Ib). CONCLUSIONI I risultati da noi ottenuti sottolineano importanti differenze tra le due barriere. La barriera emato-encefalica si sviluppa durante il primo periodo post-natale, in un ambiente caratterizzato dalla forte presenza di membrane cellulari. Similarmente l’espressione di Pgp è inizialmente bassa ed incrementa molto durante lo sviluppo post-natale. Al contrario I plessi coroidei appaiono precocemente in età embrionale, contribuiscono allo sviluppo del cervello e posseggono il più alta espressione di enzimi di fase II, coinvolti nel metabolismo di potenziali sostanze tossiche, del cervello. Un alto livello di Mrp1 sin dalla nascita suggerisce un suo coinvolgimento nel trasporto di qualche sostanza importante nello sviluppo del cervello o in un suo precoce coinvolgimento nel mantenimento dello stato ossido riduttivo, o nell’eliminazione di metabolici dal sistema nervoso centrale. Elevati livelli di bilirubina, come nel modello Gunn, modulano sia l’espressione di Pgp nella BBB, che di Mrp1 nella BCSFB. Tuttavia l’incremento nell’espressione di Pgp nei microvasi non sembra essere sufficiente a contrastare efficacemente l’ingresso della bilirubina libera, molto elevata fino al 17 giorno di vita. La simultanea riduzione di Mrp1 nei plessi coroidei, può facilitare l’ingresso o ridurre l’efflusso della bilirubina nel liquido cefalo rachidiano, consentendo l’accumulo e conseguente danno dei tessuti esposti.
................................................................ ....................... .. .BACKGROUND The unconjugated bilirubin (UCB), a heme degradation product, has been suggested to be a potent antioxidant at low concentration while it seems to be extremely dangerous at higher concentrations, causing encephalopathy in severely jaundiced neonates. Around 70% of children with kernicterus die within seven days, while the 30% survivors usually suffer irreversible sequels, including hearing loss, paralysis of upward gaze, mental retardation, and cerebral palsy with athetosis. Bilirubin encephalopathy is actually the leading cause of hospital readmission of newborns within the first month after birth. Historically the studies concerning the bilirubin entry the central nervous system have focused on the blood brain barrier (BBB), located at the level of the endothelial cells forming the brain micro vessels (MV), leading to the “free bilirubin theory”. It consists in the idea that only the free unconjugated bilirubin, the part of bilirubin exceeding the binding ability of the serum albumin, is able to cross the cell membranes and diffuse in tissue. In brain a second blood brain barrier is present. It is located at the level of the epithelial cells forming the choroids plexuses, between the blood and the cerebrospinal fluid (blood-cerebrospinal fluid barrier, BCSFB). Despite the largest surface area available for the exchanges, the high blood flux, the strategically position between two circulating fluids and the more leaky phenotype, limited studies have been made concerning its role in limiting the bilirubin entry the brain. Two ATP dependent transporters, the Multidrug Resistance-associated Protein 1 (Mrp1) and the MultiDrug resistance Protein (Pgpor MDR1), appear to be actively involved in UCB trafficking. The transporters play an important role in keeping extra cellular bilirubin concentration, such as potentially toxic compounds, below toxic levels by limiting the entry of UCB from blood to brain, or else in controlling intracellular bilirubin levels in parenchyma cells. Despite the importance of Mrp1 and Pgp on BBI their pattern of expression and cellular localization remains still unsettled. Based on these considerations - The first aim of the thesis was clarify the relative protein expression of these transporters at the two major BBI protecting the brain from toxic insults (Ia), and to identify their post-natal developmental profile of expression and cellular localisation (Ib). Similarly, no data about the Mrp1 and Pgp expression on BBI during the bilirubin encephalopathy are available. - The second aim of the thesis was investigate a relation between the high level of blood bilirubin and Mrp1 and Pgp expression in brain barriers in vivo using the Gunn rat (II), in witch the symptoms closely correlate to the human kernicterus and Crigler-Najjar syndrome type I. In this animal model, a mutation in the enzyme responsible for the conjugation and subsequent elimination of bilirubin, leads to the total absence of the enzymatic activity in the homozygous animals (jj), causing a severe life long hyperbilirubinemia. In the heterozygous Gunn rats (Jj), the enzymatic activity, until if reduced, is present and result in normal serum bilirubin levels. RESULTS Ia) By quantitative Western blot, we have demonstrated a mirroring expression of the two transporters at the blood brain interfaces in the adult rat. On the BBB the Pgp is strongly expressed and the Mrp1 amount is 15-20 times lower than in CPs. At the contrary, the CPs are characterized by the high expression of Mrp1, with a difference between the lateral ventricle (LV) and the 4th ventricle (4thV) CP, the former being a lower Mrp1 expression than in the last. In both LV and 4thV CPs, Pgp is virtually absent. By immunofluorescence we revealed that both ABC transporters are located at the blood side, the Pgp luminal on MV, and Mrp1 basal on CPs. Ib) With respect to the post-natal development, the Mrp1 expression is high since the early post-natal age and do not change significantly from birth to adult life in both barriers. By contrast, Pgp expression is weak a P9 and increase 4.6 fold with maturation on MV. Synchronously the density of Pgp stained MV in parenchyma seems increasing. II) In the homozygous Gunn rat (jj) the total bilirubin in serum is several time higher than in the heterozygous (Jj) animals all life long. By contrast the (calculated) free bilirubin is extremely elevated until the first week of life, when the bilirubin-albumin ratio exceed the unit, then drop due to the developmental increasing albumin concentration in blood. While no differences in Cx weight have been found between Jj and jj rat at every postnatal age, the cerebellum development is strongly impaired by the bilirubin toxic effect, displaying a Summarymarked hypoplasia, with about the 50% of weight loss respect the Jj control at 17 days after the birth. Concerning the ABC transporters, the differential pattern of expression between blood brain interfaces is maintained. But, in jj Gunn rats, the Pgp expression at the BBB is up-regulated at every post natal age analysed, also if this increase do not seems to be sufficient to confer protection at list until P17, when the amount of the transporter in the MV is about 5 times lower than in adult (P60). At the same time the Mrp1 expression on the BCSFB is down regulated. Since P9 the amount of Mrp1 in the 4thV CP of jj rats drops around to the 50% respect the amount in the littermates. In the LV CP the decrease is less marked, but in any case the Mrp1expression in both CPs is strongly impaired. This down regulation seems to be post-transcriptional. In the Jj animals, the Mrp1 relative expression is already high in both plexuses at early postnatal stages. A significant difference was noted only between LV CP (76%) and 4thV CP at P60 (100%). CONCLUSIONS All together these results indicate that the two barriers differ: The BBB develops after the birth and is surrounded by the lipid rich parenchyma environment, in agreement with the transporter preference for the lipid compounds and the strong post-natal developmental increase of the Pgp amount on MV. The CPs develops early in the foetal life, are involved in the guidance of the brain development and posses the highest phase II metabolising enzymes in the brain. The Mrp1 amount in CPs is similar to the adult level since the birth and may be involved in the transport of some compounds important in the brain development, in the detoxification or in the maintenance of the redox state (GS- sulfo- conjugates, LC4, etc.). In Gunn rats, as model for Kernicterus and Crigler-Najjar syndrome type I, the Pgp offered protection is not sufficiently modulate until P17, when the amount of the free bilirubin is elevated and could cross the brain barriers. The simultaneously down regulation of Mrp1 at the BCSFB may facilitate the entry of the bilirubin or strongly impair their clearance in the central nervous system, leading to the accumulation in brain and subsequent damage of tissue.
1974

Bibliography: leaves 318-367. Examines the blood-brain barrier in normal and pathological conditions induced by intravascular and extravascular insults. Intravascular insults were induced by administration of Clostridium perfringens prototoxin; extravascular insults were induced by an impact acceleration model for closed head injury to induce traumatic brain injury. Also examines the integrity of the blood-brain barrier ultrastructurally and by its ability to exclude endogenous and exogenous tracers. Also studies the expression of 2 blood-brain barrier specific proteins, endothelial barrier antigen (EBA) and glucose transporter 1 (GLUT1)

During development, netrin guidance cues control cell motility and cell adhesion. Cell-adhesion between endothelial cells at the blood-brain barrier makes the endothelium impermeable to blood-derivatives and immune cells. To establish and maintain this barrier during development, and adulthood, and as well as during disease, brain endothelial cells must develop and sustain these strong adhesive contacts, through expression of tight junction molecules. However, we do not know whether netrins support inter-endothelial cell adhesion at the blood-brain barrier. Given this, we hypothesize that netrin tightens the blood-brain barrier during development, adulthood, and protects it during disease. Methods: To test this, we used both human adult primary brain-derived endothelial cells and newborn netrin-1 knockout mice and evaluated netrin's effect on inter-endothelial cell adhesion and barrier permeability. We also assessed netrins' therapeutic potential to maintain the barrier and limit immune cell infiltration into the central nervous system (CNS) during experimental autoimmune encephalomyelitis (EAE). Results: Our results demonstrate that brain endothelial cells express netrins where they function in three ways. They help to form a tighter blood-brain barrier during development. They also maintain and protect the adult barrier by increasing the expression of endothelial junction molecules, thus promoting inter-endothelial adhesion and reducing protein leakage across the barrier. Netrins also reduce blood-brain barrier breakdown and diminish initial myeloid cell infiltration into the brain and spinal cord during EAE, which delays disease onset and ameliorates disease severity. However, during the chronic phase of EAE, netrin-1 treated mice have higher numbers and more activated T cells in their CNS and exhibit an ataxic gait and limb spasticity. Discussion: We conclude that netrins enhance BBB stability, but have dual functions on immune responses during neuroinflammatory disease. These findings favour the hypothesis that if netrin function was to be manipulated as a therapeutic, early short-term approaches would likely be the most effective.
Au cours du développement, les molécules de la famille des nétrines contribuent à la morphologénèse des organes en contrôlant la motilité et l'adhérence cellulaire. L'adhérence cellulaire entre les cellules endothéliales est une caractéristique importante de la barrière hémato-encéphalique (BHE), ce qui rend l'endothélium imperméable aux molécules sanguines et aux cellules immunitaires. Pour établir et maintenir cette barrière au cours du développement, à l'âge adulte et au cours de la maladie, les cellules endothéliales du cerveau doivent développer et maintenir ces contacts adhésifs en exprimant des molécules de jonction serrées. Cependant, nous ne savons pas si les molécules de la famille des nétrines influencent l'adhérence cellulaire inter-endothéliale de la BHE. Nous avons donc émis l'hypothèse que les nétrines resserrent la BHE au cours du développement, à l'âge adulte, et la protège au cours de la maladie.Méthodes: Pour valider notre hypothèse, nous avons utilisé des cellules endothéliales primaires dérivées des cerveaux humains adultes ou des cerveaux de souris nouveau-nés déficientes en nétrine-1 et évalué l'effet de la nétrine sur l'adhésion cellulaire endothéliale et inter-perméabilité de la barrière. Nous avons également évalué le potentiel thérapeutique des nétrines a restaurer la barrière et l'infiltration de cellules immunitaires limite dans le système nerveux central (SNC) pendant encéphalomyélite allergique expérimentale, un modèle animal de sclérose en plaques. Résultats: Nos résultats démontrent que les nétrines sont exprimées par les cellules endothéliales du cerveau, exprimes nétrines. Au cours du développement les nétrines aident à assurer l'étanchéité de la BHE. Chez les adultes, ils maintiennent et protègent la barrière adulte en augmentant l'expression des molécules de jonctions serrées, favorisant ainsi l'adhérence inter-endothéliale et diminuant les fuites de protéines à travers la BHE. Dans la pathologie de l'EAE, le rôle des nétrins diffère en fonction de la phase de la maladie. Au cours de la phase aigue, les nétrines atténuent la perte de l'intégrité de la BHE et diminuent l'infiltration des cellules myéloïdes dans le SNC. Ceci retarde l'apparition de la maladie et réduit sa sévérité. Au cours de la phase chronique de l'EAE, les souris traitées avec netrin-1 ont un plus grand nombre des cellules T activées dans leurs SNC et présentent une démarche ataxique ainsi qu'une spasticité des membres. Discussion: Nous concluons que les nétrins améliorent la stabilité de la BHE. Ces résultats suggèrent que les nétrines peuvent être envisagée comme agent thérapeutique dans les maladies neuroinflammatoire. Dans ce cas une approche précoce et à court terme serait probablement plus efficace.

Thesis (S.M.)--Massachusetts Institute of Technology, Dept. of Nuclear Science and Engineering, 2007.
Includes bibliographical references (p. 53-56).
Selective vascular irradiation enables the critical examination of the vasculature and its role in the onset of late radiation effects. It is a novel approach to expose the endothelial cells to much higher levels of ionizing radiation relative to normal cells by utilizing the boron neutron capture reaction. When boron-containing compounds are restricted to the lumen of the blood vessel, the resulting high-LET alpha and lithium particles cannot deposit their energy in the normal cells beyond the vasculature after the target is exposed to thermal neutrons. This allows for a 2- to 3-fold increase in the calculated dose to the endothelial cells. However, this technique has been criticized because there is no direct evidence that the endothelial cells receive an absorbed dose from the selective vascular irradiation. The objective of this work is to provide corroborating experimental evidence that selective vascular irradiation physically damages the endothelial cells. An established assay utilizing blood-brain barrier disruption was adopted to quantify the radiation damage to the endothelial cells in female BALB/C mice, 8-12 weeks of age. A dye that attaches to the plasma proteins in the blood and that is ordinarily kept out of the brain by the blood-brain barrier is injected into the blood supply before the irradiation, and following irradiation, damage to the vasculature will result in disruption of the blood-brain barrier that allows blood stained with the dye to enter the brain. After sacrificing, the blood in the vessel lumen is cleared by performing a trans-cardiac perfusion, and the brain is homogenized and prepared for analysis. The absorbance of the resulting supernatant of each brain sample is measured with a spectrophotometer at the optimal wavelength of the dye.
(cont.) The absorbance is related to the quantity of blood that leaked through the blood-brain barrier, which is also related to the damage caused to the vasculature from exposure to ionizing radiation. Increased leakage through the blood-brain barrier was observed for those mice exposed to selective vascular irradiation, indicating a direct relationship between the leakage through the blood-brain barrier and the 10B concentration in the blood. The most significant increase in the leakage through the blood-brain barrier (p<0.002) was observed at the highest lOB concentration in the blood (161 ppm). The compound biological effectiveness (CBE) for sulfhydryl borane (BSH) was calculated to be 0.28, which is consistent with the published value of the CBE for BSH in the rat spinal cord. This suggests that the assumptions used for calculating the absorbed doses for selective vascular irradiation are reasonable and approximate to what the endothelial cells receive.
by Rebecca L. Raabe.
S.M.

The blood-brain barrier (BBB) is located at the level of the cerebral microvasculature and is critical to maintain central nervous system (CNS) homeostasis. The tight junction (TJ) protein complexes between endothelial cells at the BBB are primarily responsible for limiting paracellular diffusion of substances from the blood to the CNS. The BBB’s functional integrity is compromised in a number of disease states which affect the CNS, suggesting BBB dysfunction causes or contributes to many diseases of the CNS. A common component of most of these diseases is oxidative stres. Oxidative stress is associated with hypoxia-reoxygenation (HR) and peripheral inflammatory pain (PIP). Both HR and PIP have been shown to compromise BBB functional integrity. Using in vivo rat models of HR and PIP, we examined the role of ROS on BBB permeability as well as the TJ protein occludin using the free radical scavenger tempol. First, we subjected rats to HR with or without pre-treatment with tempol (200 mg/kg). We showed that tempol prevents up-regulation of the cellular stress marker heat shock protein 70 at the BBB during HR. Next we showed tempol reverses HR-mediated BBB permeability increase to ¹⁴C-sucrose, a marker of BBB paracellular permeability. Tempol also attenuated changes in the structure and localization of occludin, suggesting ROS produced during HR alter occludin and lead to disruption of BBB. We then investigated whether ROS production have similar effects on occludin and BBB permeability during PIP by administering 3% λ-carrageenan into the hind paw of rats. We found tempol attenuated carrageenan-induced increase in paw edema and thermal hyperalgesia. Tempol also attenuated up-regulation of the cellular stress marker NF-κB in cerebral microvessels. Tempol significantly decreased BBB permeability to ¹⁴C sucrose during PIP. We found PIP reduces disulfide bonds in occludin oligomeric assemblies thought to be important in maintaining the structural integrity of the BBB. Tempol significantly inhibited disulfide bond reduction, suggesting ROS mediate BBB disruption during inflammatory pain by reducing occludin disulfide bonding. Taken together, these findings show the involvement of ROS during HR and PIP contributes to BBB dysfunction by altering the structure of high molecular weight occludin oligomeric assemblies.

Dans les systèmes biologiques, les barrières tissulaires permettent le transport sélectif de molécules du sang au tissu approprié. Un exemple de barrière tissulaire est la barrière hémato-encéphalique (BHE). La BHE protège le cerveau du sang et maintient l'homéostasie du microenvironnement du cerveau, ce qui est essentiel à l'activité et à la fonction neuronale. La caractérisation de cette BHE est importante, car un dysfonctionnement de cette barrière est souvent révélateur de toxicité ou de maladie. Bien que le nombre d'articles publiés dans le domaine du développement et de la caractérisation de la BHE ait été multiplié ces dernières années, la validité des modèles utilisés est encore un sujet de débat. L'avènement de l'électronique organique a créé une occasion unique pour coupler les mondes de l'électronique et de la biologie, à l'aide de dispositifs tels que le transistor électrochimique organique (OECT). OECT constitue un outil très sensible et économique pour diagnostiquer l’intégrité d’une barrière tissulaire. Dans cette étude, nous avons tout d’abord développé trois différents modèles de BHE. Nous avons optimisé l’adhésion des cellules endothéliales cérébrales sur la matière active du transistor. Nous avons ainsi pu établir l'intégration des OECTs avec des cellules immortalisées humaines micro vasculaires cérébrales endothéliales (h CMEC/D3) en tant que modèle in vitro de BHE. Nous avons démontré que la fonction de tissu de la BHE peut être détectée en utilisant cette nouvelle technique. En outre, par cette technique, une perturbation de la barrière (par exemple, provoquée par un composé toxique) pourra être détectée électriquement au moyen d'une mesure de courant
In biological systems many tissue types have evolved a barrier function to selectively allow the transport of matter from the lumen to the tissue beneath; one example is the Blood Brain Barrier (BBB). The BBB protects the brain from the blood and maintains homeostasis of the brain microenvironment, which is crucial for neuronal activity and function. Characterization of the BBB is very important as its disruption or malfunction is often indicative of toxicity/disease. Though the number of published papers in the field of in vitro BBB has multiplied in recent years, the validity of the models used is still a subject of debate.The advent of organic electronics has created a unique opportunity to interface the worlds of electronics and biology, using devices such as the Organic ElectroChemical Transistor (OECT), which provide a very sensitive way to detect minute ionic currents in an electrolyte as the transistor amplifies the gate current.In this study, we test three different type of BBB in order to develop a stable BBB model. We optimize the adhesion of brain endothelial cell on OECT conducting polymer. We show the integration of OECTs with immortalized human cerebral microvascular endothelial cells as a model of human blood brain barrier, and demonstrate that the barrier tissue function can be detected. Moreover, by this technique, a disruption in the barrier (e.g. caused by a toxic compound) is assessed electrically through a measurement of the drain current. Results show the successful development and validation of an in vitro BBB model. Dynamic monitoring of the barrier properties of the BBB barrier tissue was possible using the OECT

"Intranasal drug administration is an effective method that has shown promise for delivering drugs directly to the brain. This approach is associated with many challenges, and efficacy in bypassing blood-brain barrier (BBB) is debated. This review describes the pathways of nose-to-brain drug delivery, physicochemical drug properties that influence drug uptake through the nasal epithelium, physiological barriers, methods to enhance nose-to-brain absorption, drug bioavailability and biodistribution, and intranasal devices for nose-to-brain drug delivery. The mechanism of each device is described and supporting evidence from clinical trials is presented. This paper summarizes strategies involved in nose-to-brain drug delivery and provides evidence of potential efficacy of nose-braindelivery from clinical trials."

Background: Antibiotics exposure is related to gastrointestinal tract dysbiosis and appearance of systemic repercussions. Due to the correlation between Enteric Nervous System (ENS) and Central Nervous System (CNS), abnormalities in the gut microbiota have been associated with neurological disorders including Parkinson’s Disease (PD). Objectives: Search evidence in the scientific literature relating antibiotic therapy and Parkinson’s disease. Methods: A systematic review has been done using the descriptors “Parkinson’s disease”, “antibiotics” and “gut microbiota” in PubMed’s database. The research was conducted in april 2021, without temporal limitations, in english and portuguese. Results: Studies suggest that PD begins with intestinal inflammation and abnormal alpha-synuclein deposition in the ENS that follows, through nerves, to the CNS. Results show that leaky gut and dysbiosis preceded 5-10 years PD’s initial symptoms, while the intense exposure to antibiotics preceded 10-15 years the diagnostic. On average, PD patients received larger amounts of antibiotics than controls (p=0.021). Dysbiosis post-antibiotics presented reduced diversity of Bacteroidetes, Firmicutes and Prevotellaceae and growthing of Enterobacteriaceae, resulting in higher risk of gastrointestinal infections, higher rates of pro-inflammatory cytokines, increased permeability of gastrointestinal and brain-blood barriers and hyperexpression of the alpha-synuclein protein in the colon. Conclusion: Poorly controlled antibiotic therapy and its subsequent damage to gut microbiota anticipates PD’s early symptoms.

Cerebral blood vessels are critical in maintaining the health and function of the brain, but their function can be disrupted by traumatic brain injury (TBI), which commonly includes damage to these vessels [1]. However, even in cases where there is not apparent mechanical damage to the cerebral vasculature, TBI can induce physiological disruptions that can lead to breakdown of the blood brain barrier or loss of cerebral autoregulation.

A number of studies found that the virus can activate the endothelial cells and affect the structure and function of the blood?brain barrier, promoting immune cell migration to benefit the virus nervous system target cells infected by flaviviruses.