191 results on '"Cecchelli, Roméo"'
Search Results
152. Viral neuraminidase and cellular ectosialytransferase in human lymphoblastoid cells infected with influenza virus
- Author
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Duc Dodon, Madeleine, primary, Cecchelli, Roméo, additional, Cacan, René, additional, Gazzolo, Louis, additional, and Verbert, André, additional
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- 1984
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153. Structural determination of the novel fragmentation routes of morphine opiate receptor antagonists using electrospray ionization quadrupole time-of-flight tandem mass spectrometry.
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Joly, Nicolas, Aneed, Anas El, Martin, Patrick, Cecchelli, Roméo, and Banoub, Joseph
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- 2006
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154. Protein kinase C restricts transport of carnitine by amino acid transporter ATB0,+ apically localized in the blood–brain barrier.
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Michalec, Katarzyna, Mysiorek, Caroline, Kuntz, Mélanie, Bérézowski, Vincent, Szczepankiewicz, Andrzej A., Wilczyński, Grzegorz M., Cecchelli, Roméo, and Nałęcz, Katarzyna A.
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PROTEIN kinase C , *CARNITINE , *AMINO acid transport , *BIOLOGICAL transport , *BLOOD-brain barrier , *ENDOTHELIAL cells - Abstract
Highlights: [•] Amino acid transporter B0,+ (ATB0,+) is apically localized in blood–brain barrier. [•] ATB0,+ in the blood–brain barrier is controlled by protein kinase C. [•] ATB0,+ is inactivated by exclusion from membrane rafts. [•] Brain capillary endothelial cells control carnitine gradient. [ABSTRACT FROM AUTHOR]
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- 2014
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155. A differential proteomic approach identifies structural and functional components that contribute to the differentiation of brain capillary endothelial cells
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Pottiez, Gwënaël, Duban-Deweer, Sophie, Deracinois, Barbara, Gosselet, Fabien, Camoin, Luc, Hachani, Johan, Couraud, Pierre-Olivier, Cecchelli, Roméo, Dehouck, Marie-Pierre, Fenart, Laurence, Karamanos, Yannis, and Flahaut, Christophe
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PROTEOMICS , *CELL differentiation , *ENDOTHELIUM , *ASTROCYTES , *BLOOD-brain barrier , *CYTOSKELETON , *CAPILLARY electrophoresis - Abstract
Abstract: When in the vicinity of astrocytes, brain capillary endothelial cells (BCECs) develop the characteristic structural and functional features of the blood-brain barrier (BBB). The latter has low cellular permeability and restricts various compounds from entering the brain. We recently reported that the cytoskeleton-related proteins actin, gelsolin and filamin-A undergo the largest quantitative changes in bovine BCECs after re-induction of BBB functions by co-culture with glial cells. In the present study, we used an in-depth, proteomic approach to quantitatively compare differences in Triton-X-100-solubilized proteins from bovine BCECs with limited or re-induced BBB functions (i.e. cultured in the absence or presence of glial cells, respectively). The 81 protein spots of differing abundance were linked to 55 distinct genes. According to the Protein ANalysis THrough Evolutionary Relationships classification system and an Ingenuity Pathway Analysis, these quantitative changes mainly affected proteins involved in (i) cell structure and motility and (ii) protein metabolism and modification. The fold-changes affecting HSPB1, moesin and ANXA5 protein levels were confirmed by western blot analysis but were not accompanied by changes in the corresponding mRNA expression levels. Our results reveal that the bovine BCECs'' phenotype adaptation to variations in their environment involves the reorganization of the actin cytoskeleton. [Copyright &y& Elsevier]
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- 2011
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156. New strategy for alerting central nervous system toxicity: Integration of blood–brain barrier toxicity and permeability in neurotoxicity assessment
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Hallier-Vanuxeem, Dorothée, Prieto, Pilar, Culot, Maxime, Diallo, Heidi, Landry, Christophe, Tähti, Hanna, and Cecchelli, Roméo
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IN vitro toxicity testing , *BLOOD-brain barrier , *NEUROTOXICOLOGY , *PERMEABILITY , *NERVOUS system blood-vessels , *CELL lines , *BIOLOGICAL mathematical modeling - Abstract
Abstract: The combination of an in vitro BBB model (4d/24w) with a neuronal cell line (SH-SY5Y) provides a convenient approach to explore the importance of BBB permeability in neurotoxicity assessment of compounds. The toxicity of 16 compounds on SH-SY5Y cells was evaluated after 24h incubation with each compound and compared to their toxicity on SH-SY5Y after passage through the BBB model. Nine out of 16 compounds were found toxic after direct exposure at 100μM while only three still induced toxicity on SH-SY5Y cells after BBB transport. The BBB permeability values of each compound revealed that in the case of compounds that did not induce toxicity, the amount that crossed the BBB was not enough to exert a toxic effect on the neuronal cells. Since disrupting the BBB may also cause unwanted effect on brain cells, the BBB toxicity of these compounds have been assessed. Our results prompted the importance of BBB permeability assessment in neurotoxicity evaluation, as it allows a better estimation of the actual concentration at the target site. [Copyright &y& Elsevier]
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- 2009
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157. A polarized localization of amino acid/carnitine transporter B0,+ (ATB0,+) in the blood–brain barrier
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Czeredys, Magdalena, Mysiorek, Caroline, Kulikova, Natalia, Samluk, Łukasz, Berezowski, Vincent, Cecchelli, Roméo, and Nałęcz, Katarzyna A.
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ENDOTHELIAL seeding , *VASCULAR grafts , *VITAMIN B complex , *AMINO acids - Abstract
Brain capillary endothelial cells control the uptake and efflux from the brain of many hydrophilic compounds due to highly specialized transporters often localized in a polarized way. Localization of Na+- and Cl−-dependent amino acid and carnitine transporter B0,+ (ATB0,+) was studied in a co-culture of bovine brain capillary endothelial cells (BBCEC) grown on filters above astrocytes (an in vitro blood–brain barrier model). Immunoblotting and three-dimensional immunocytochemistry analysis with anti-B0,+antibodies demonstrated the presence of this transporter and its prevalent co-localization with P-glycoprotein i.e. at the apical side. The sensitivity of leucine uptake through the apical membrane to 2-aminobicyclo-[2.2.1]-heptane-2-carboxylic acid (BCH), d-serine as well as sodium and chloride replacement confirm the functioning of ATB0,+ and suggests an important physiological role of ATB0,+ in controlling the delivery of amino acids and carnitine to the brain. [Copyright &y& Elsevier]
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- 2008
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158. An in vitro blood-brain barrier model for high throughput (HTS) toxicological screening
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Culot, Maxime, Lundquist, Stefan, Vanuxeem, Dorothée, Nion, Stéphane, Landry, Christophe, Delplace, Yannick, Dehouck, Marie-Pierre, Berezowski, Vincent, Fenart, Laurence, and Cecchelli, Roméo
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CELLS , *ENDOTHELIUM , *NEUROGLIA , *HIGH throughput screening (Drug development) , *PROTEINS , *GLYCOPROTEINS - Abstract
Abstract: There is a growing interest to use in vitro BBB cell assays in early safety assessment of compounds. By modifying a well-validated co-culture model of brain capillary endothelial and glial cells, developed by Dehouck et al. [Dehouck, M.P., Meresse, S., Delorme, P., Fruchart, J.C., Cecchelli, R., 1990. An easier, reproducible, and mass-production method to study the blood-brain barrier in vitro. Journal of Neurochemistry 54 (5), 1798–1801], it has been possible to develop a new in vitro BBB system suitable for high throughput screening (HTS). In addition, this new procedure substantially reduces the use of experimental animals and considerably facilitates the process of obtaining a functional in vitro BBB model. The model is ready to use after only 4 days of culture and then shows the typical expression and localization of tight junction proteins. The function of the P-glycoprotein and the transcriptional expression of other efflux transporters such as MRP 1, 4 and 5 have been demonstrated. In addition, the model produces a good in vitro/in vivo correlation for 10 compounds (R 2 =0.81). Furthermore, studies were undertaken within the European ACuteTox consortium with the objective to assess BBB toxicity and make risk assessments of potentially toxic compounds according to their predicted ability to reach the CNS compartment. These investigations demonstrated that the results produced in the HTS BBB model were similar to the standard co-culture model. [Copyright &y& Elsevier]
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- 2008
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159. Localization of organic cation/carnitine transporter (OCTN2) in cells forming the blood–brain barrier.
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Miecz, Dorota, Januszewicz, Elżbieta, Czeredys, Magdalena, Hinton, Barry T., Berezowski, Vincent, Cecchelli, Roméo, and Nałęcz, Katarzyna A.
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CARNITINE , *BLOOD-brain barrier , *ASTROCYTES , *FATTY acids , *OXIDATION , *MESSENGER RNA , *BRAIN - Abstract
Carnitine β-hydroxy-γ-(trimethylammonio)butyrate – a compound necessary in the peripheral tissues for a transfer of fatty acids for their oxidation within the cell, accumulates in the brain despite low β-oxidation in this organ. In order to enter the brain, carnitine has to cross the blood–brain barrier formed by capillary endothelial cells which are in close interaction with astrocytes. Previous studies, demonstrating expression of mRNA coding two carnitine transporters – organic cation/carnitine transporter 2 (OCTN2) and B0,+ in endothelial cells, did not give any information on carnitine transporters polarity in endothelium. Therefore more detailed experiments were performed on expression and localization of a high affinity carnitine transporter OCTN2 in an in vitro model of the blood–brain barrier by real-time PCR, western blot analysis, and immunocytochemistry. The amount of mRNA was comparable in endothelial cells and kidney, when referred to house-keeping genes, it was, however, significantly lower in astrocytes. Polarity of OCTN2 localization was further studied in an in vitro model of the blood–brain barrier with use of anti-OCTN2 antibodies. Z-axis analysis of the confocal microscope pictures of endothelial cells, with anti-P-glycoprotein antibodies as the marker of apical membrane, showed OCTN2 localization at the basolateral membrane and in the cytoplasmic region in the vicinity of nuclei. Localization of OCTN2 suggest that carnitine can be also transported from the brain, playing an important role in removal of certain acyl esters. [ABSTRACT FROM AUTHOR]
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- 2008
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160. Evidence of lowest brain penetration of an antiemetic drug, metopimazine, compared to domperidone, metoclopramide and chlorpromazine, using an in vitro model of the blood–brain barrier
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Jolliet, Pascale, Nion, Stéphane, Allain-Veyrac, Gwenaëlle, Tilloy-Fenart, L., Vanuxeem, Dorothée, Berezowski, Vincent, and Cecchelli, Roméo
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CHROMATOGRAPHIC analysis , *HIGH performance liquid chromatography , *NERVOUS system , *LIQUID chromatography - Abstract
Abstract: Purpose: The objective of the current study was to determine the ability of some antiemetic compounds to cross the blood–brain barrier (BBB) and thereby to determine possible side effects of compounds for the central nervous system (CNS). Methods: We compared the brain penetration of some antiemetic compounds using an in vitro BBB model consisting in brain capillary endothelial cells co-cultured with primary rat glial cells. Results: This study clearly demonstrated that the metopimazine metabolite, metopimazine acid, has a very low brain penetration, lower than metopimazine and even less than the other antiemetic compounds tested in this study. Conclusions: The poor brain penetration of metopimazine acid, metopimazine biodisponible form, seems very likely related to the clinically observed difference in therapeutic and safety profile. [Copyright &y& Elsevier]
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- 2007
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161. The MAP kinase pathway mediates transcytosis induced by TNF-α in an in vitro blood–brain barrier model.
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Miller, Florence, Fenart, Laurence, Landry, Valérie, Coisne, Caroline, Cecchelli, Roméo, Dehouck, Marie‐Pierre, and Buée‐Scherrer, Valérie
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PROTEIN kinases , *PHOSPHOTRANSFERASES , *TUMOR necrosis factors , *BRAIN , *CYTOKINES , *GLYCOPROTEINS - Abstract
Cerebral capillary endothelial cells constitute the blood–brain barrier (BBB). In these highly specialized cells, transcellular transports rarely occur, and the presence of tight junctions between them leads to a low paracellular permeability. In order to understand the functions of this barrier, an in vitro model of the BBB has been developed and consists in a co-culture of primary cerebral capillary endothelial cells and glial cells. When these endothelial cells are subjected to an inflammatory agent, such as tumor necrosis factor-alpha (TNF-α), in vitro BBB permeability is increased, as indicated by the increase in holotransferrin transcytosis. However, no significant change in the paracellular permeability is observed. In order to understand the molecular mechanisms that underlie these transcytosis processes, we investigated the implication of the mitogen-activated protein kinase (MAPK) signalling pathway, as TNF-α is known to activate this kinase family. In the present study, an increase in the activation of p42–44 MAPK is observed after TNF-α treatment. Holotransferrin transcytosis as well as p42–44 MAPK activation are inhibited after addition of a p42–44 MAPK pathway inhibitor (UO126) during TNF-α challenge. These data suggest that the MAPK pathway is involved in the transcytosis regulation in endothelial cells from an in vitro BBB model. [ABSTRACT FROM AUTHOR]
- Published
- 2005
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162. Adamantoylated monosaccharides: new compounds for modification of the properties of cyclodextrin-containing materials
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Binkowski, Cécile, Lequart, Vincent, Hapiot, Frédéric, Tilloy, Sébastien, Cecchelli, Roméo, Monflier, Eric, and Martin, Patrick
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GLYCOSIDES , *MONOSACCHARIDES , *SACCHARIDES , *GLUCOSIDES - Abstract
Abstract: Adamantoyl glycosides were obtained in good yields by coupling adamantanecarboxylic acid with monosaccharides. They form very stable inclusion complexes with β-cyclodextrin, as shown by 1H NMR measurements. [Copyright &y& Elsevier]
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- 2005
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163. Carnitine: transport and physiological functions in the brain
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Nalęcz, Katarzyna A., Miecz, Dorota, Berezowski, Vincent, and Cecchelli, Roméo
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CARNITINE , *VITAMIN B complex , *BIOMOLECULES , *FATTY acids - Abstract
Carnitine (4-N-trimethylammonium-3-hydroxybutyric acid), a compound necessary for a transfer of fatty acids for their oxidation within the cell, accumulates in brain although β-oxidation of fatty acids is very low in neurons. Carnitine accumulates to lower extent in the brain than in peripheral tissues and the mechanism of its transport through the blood–brain barrier is discussed, with the involvement of two transporters, OCTN2 and B0,+ being presented. A limitation by the blood–brain barrier of carnitine supply for the brain and the mechanism of its transport to neural cells by a protein belonging to neurotransmitters'' transporters superfamily is further discussed.Due to the beneficial effects of administration of acetylcarnitine in case of patients with dementia, the role of this acylcarnitine is presented in the context of neuronal cell metabolism and the role of acetylcarnitine in the synthesis of acetylcholine. The roles of long-chain acyl derivatives of carnitine, in particular palmitoylcarnitine, responsible for interaction with the membranes, lipids acylation and specific interactions with proteins have been summarized. Stimulation of protein palmitoylation and a possibility of changing the acylation status of G proteins is described, as well as interaction of palmitoylcarnitine with protein kinase C. Diminished interaction of the isoform δ of this kinase with GAP-43 (B-50, neuromodulin), whose expression increases upon accumulation of either carnitine or palmitoylcarnitine points to a possible regulation of differentiation by these compounds and their role in neuroregeneration. [Copyright &y& Elsevier]
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- 2004
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164. Synthesis of chacotriose analogues
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Lequart, Vincent, Goethals, Gérard, Usubillaga, Alfredo, Villa, Pierre, Cecchelli, Roméo, and Martin, Patrick
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CARBOHYDRATES , *SUGAR synthesis , *ORGANIC synthesis , *GLUCOSE - Abstract
We report here the synthesis of three chacotriose analogues, namely β-l-fucopyranosyl-(1 → 2)-[β-l-fucopyranosyl-(1 → 4)]-d-glucopyranose, β-l-fucopyranosyl-(1 → 2)-[β-l-fucopyranosyl-(1 → 4)]-d-galactopyranose, and α-l-rhamnopyranosyl-(1 → 2)-[α-l-rhamnopyranosyl-(1 → 4)]-α-d-galactopyranose. [Copyright &y& Elsevier]
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- 2004
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165. Astrocyte mediated modulation of blood-brain barrier permeability does not correlate with a loss of tight junction proteins from the cellular contacts.
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Hamm, Stefan, Dehouck, Bénédicte, Kraus, Jörg, Wolburg-Buchholz, Karen, Wolburg, Hartwig, Risau, Werner, Cecchelli, Roméo, Engelhardt, Britta, and Dehouck, Marie-Pierre
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BLOOD-brain barrier , *TIGHT junctions , *CENTRAL nervous system , *ASTROCYTES , *PROTEINS , *BRAIN blood-vessels , *ELECTRON microscopy - Abstract
In the central nervous system (CNS) complex endothelial tight junctions (TJs) form a restrictive paracellular diffusion barrier, the blood-brain barrier (BBB). Pathogenic changes within the CNS are frequently accompanied by the loss of BBB properties, resulting in brain edema. In order to investigate whether BBB leakiness can be monitored by a loss of TJ proteins from cellular borders, we used an in vitro BBB model where brain endothelial cells in co-culture with astrocytes form a tight permeability barrier for 3H-inulin and 14C-sucrose. Removal of astrocytes from the co-culture resulted in an increased permeability to small tracers across the brain endothelial cell monolayer and an opening of the TJs to horseradish peroxidase as detected by electron microscopy. Strikingly, opening of the endothelial TJs was not accompanied by any visible change in the molecular composition of endothelial TJs as junctional localization of the TJ-associated proteins claudin-3, claudin-5, occludin, ZO-1 or ZO-2 or the adherens junction-associated proteins β-catenin or p120cas did not change. Thus, opening of BBB TJs is not readily accompanied by the complete loss of the junctional localization of TJ proteins. [ABSTRACT FROM AUTHOR]
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- 2004
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166. P-glycoprotein in blood–brain barrier endothelial cells: interaction and oligomerization with caveolins.
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Jodoin, Julie, Demeule, Michel, Fenart, Laurence, Cecchelli, Roméo, Farmer, Sarah, Linton, Kenneth J., Higgins, Christopher F., and Béliveau, Richard
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GLYCOPROTEINS , *BLOOD-brain barrier , *ASTROCYTES - Abstract
P-glycoprotein (P-gp), an adenosine triphosphate (ATP)-binding cassette transporter which acts as a drug efflux pump, is highly expressed at the blood–brain barrier (BBB) where it plays an important role in brain protection. Recently, P-gp has been reported to be located in the caveolae of multidrug-resistant cells. In this study, we investigated the localization and the activity of P-gp in the caveolae of endothelial cells of the BBB. We used an in vitro model of the BBB which is formed by co-culture of bovine brain capillary endothelial cells (BBCEC) with astrocytes. Caveolar microdomains isolated from BBCEC are enriched in P-gp, cholesterol, caveolin-1, and caveolin-2. Moreover, P-gp interacts with caveolin-1 and caveolin-2; together, they form a high molecular mass complex. P-gp in isolated caveolae is able to bind its substrates, and the caveolae-disrupting agents filipin III and nystatin decrease P-gp transport activity. In addition, mutations in the caveolin-binding motif present in P-gp reduced the interaction of P-gp with caveolin-1 and increased the transport activity of P-gp. Thus, P-gp expressed at the BBB is mainly localized in caveolae and its activity may be modulated by interaction with caveolin-1. [ABSTRACT FROM AUTHOR]
- Published
- 2003
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167. Tumor Necrosis Factor-α Increases Lactoferrin Transcytosis Through the Blood-Brain Barrier.
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Fillebeen, Carine, Dehouck, Bénédicte, Benaïssa, Monique, Dhennin-Duthille, Isabelle, Cecchelli, Roméo, and Pierce, Annick
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LACTOFERRIN , *INFLAMMATION , *TRANSFORMING growth factors , *PHYSIOLOGY - Abstract
Abstract: Lactoferrin (Lf) is an iron-binding protein involved in host defense against infection and severe inflammation, which accumulates in the brain during neurodegenerative disorders. Prior to determining Lf function in pathological brain tissues, we investigated its transport through the blood-brain barrier (BBB) in inflammatory conditions. For this purpose, we used a reconstituted BBB model consisting of the coculture of bovine brain capillary endothelial cells (BBCECs) and astrocytes in the presence of tumor necrosis factor-α (TNF-α). As TNF-α can be either synthesized by brain glial cells or present in circulating blood, BBCECs were exposed to this cytokine at their luminal or abluminal side. We have been able to demonstrate that in the presence of TNF-α, whatever the type of exposure, BBCECs were activated and Lf transport through the activated BBCECs was markedly increased. Lf was recovered intact at the abluminal side of the cells, suggesting that increased Lf accumulation may occur in immune-mediated pathophysiology. This process was transient as 20 h later, cells were in a resting state and Lf transendothelial traffic was back to normal. The enhancement of Lf transcytosis seems not to involve the up-regulation of the Lf receptor but rather an increase in the rate of transendothelial transport. [ABSTRACT FROM AUTHOR]
- Published
- 1999
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168. Transport of Cationized Anti-Tetanus Fab'2 Fragments Across an In Vitro Blood-Brain Barrier Model: Involvement of the Transcytosis Pathway.
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Girod, Julie, Fenart, Laurence, Régina, Anthony, Dehouck, Marie-Pierre, Hong, Guying, Scherrmann, Jean-Michel, Cecchelli, Roméo, and Roux, Françoise
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TETANUS toxin , *CENTRAL nervous system , *AXONAL transport , *BLOOD-brain barrier - Abstract
Abstract: Tetanus neurotoxin reaches the CNS by axonal retrograde transport and thus becomes inaccessible to current treatments. A possible strategy to improve current therapy for tetanus disease would be the vectorization of Fab'2 fragments, allowing their delivery into the CNS. The purpose of this study was to investigate whether after cationization anti-tetanus Fab'2 fragments are able to cross the blood-brain barrier, the first obstacle to CNS delivery. We used primary cocultures of bovine brain capillary endothelial cells and newborn rat astrocytes as an in vitro model to study the binding and transport of cationized Fab'2 (cFab'2) fragments across the brain endothelium. We first show that cationization does not alter Fab'2 affinity for tetanus toxin. Then we demonstrate that after cationization Fab'2 fragments are able to bind to the negative charges on the surface of endothelial cells and subsequently to be transported across the endothelial cell monolayer without any modification of affinity. Finally, using fluorescence microscopy, we show that cFab'2 fragments are transported through endocytotic vesicles. The present study demonstrates that cationization allows Fab'2 directed against tetanus toxin to be transported through brain endothelium by adsorptive-mediated transcytosis. We suggest that this vectorization way could be a promising delivery strategy for carrying anti-tetanic immunoglobulin fragments across the blood-brain barrier to improve... [ABSTRACT FROM AUTHOR]
- Published
- 1999
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169. Receptor-Mediated Transcytosis of Cyclophilin B Through the Blood—Brain Barrier.
- Author
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Carpentier, Mathieu, Descamps, Laurence, Allain, Fabrice, Denys, Agnès, Durieux, Sandrine, Fenart, Laurence, Kieda, Claudine, Cecchelli, Roméo, and Spik, Geneviève
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CYCLOPHILINS , *BLOOD-brain barrier - Abstract
Cyclophilin B (CyPB) is a cyclosporin A (CsA)-binding protein mainly located in intracellular vesicles and secreted in biological fluids. In previous works, we demonstrated that CyPB interacts with T lymphocytes and enhances in vitro cellular incorporation and activity of CsA. In addition to its immunosuppressive activity, CsA is able to promote regeneration of damaged peripheral nerves. However, the crossing of the drug from plasma to neural tissue is restricted by the relative impermeability of the blood—brain barrier. To know whether CyPB might also participate in the delivery of CsA into the brain, we have analyzed the interactions of CyPB with brain capillary endothelial cells. First, we demonstrated that CyPB binds to two types of binding sites present at the surface of capillary endothelial cells from various species of tissues. The first type of binding sites (KD = 300 nM; number of sites = 3 × 106) is related to interactions with negatively charged compounds such as proteoglycans. The second type of binding sites, ∼50,000 per cell, exhibits a higher affinity for CyPB (KD = 15 nM) and is involved in an endocytosis process, indicating it might correspond to a functional receptor. Finally, the use of an in vitro model of blood—brain barrier allowed us to demonstrate that CyPB is transcytosed by a receptor-mediated pathway (flux = 16.5 fmol/cm2/h). In these conditions, CyPB did not significantly modify the passage of CsA, indicating that it is unlikely to provide a pathway for CsA brain delivery. [ABSTRACT FROM AUTHOR]
- Published
- 1999
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170. Pharmacokinetics and In Vitro Blood-Brain Barrier Screening of the Plant-Derived Alkaloid Tryptanthrin.
- Author
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Jähne EA, Eigenmann DE, Sampath C, Butterweck V, Culot M, Cecchelli R, Gosselet F, Walter FR, Deli MA, Smieško M, Hamburger M, and Oufir M
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- Animals, Cell Line, Chromatography, High Pressure Liquid methods, Humans, Isatis chemistry, Male, Molecular Structure, Plant Extracts pharmacokinetics, Quinazolines chemical synthesis, Quinazolines chemistry, Rats, Rats, Sprague-Dawley, Blood-Brain Barrier metabolism, Quinazolines pharmacokinetics
- Abstract
The indolo[2,1-b]quinazoline alkaloid tryptanthrin was previously identified as a potent anti-inflammatory compound with a unique pharmacological profile. It is a potent inhibitor of cyclooxygenase-2, 5-lipooxygenase-catalyzed leukotriene synthesis, and nitric oxide production catalyzed by the inducible nitric oxide synthase. To characterize the pharmacokinetic properties of tryptanthrin, we performed a pilot in vivo study in male Sprague-Dawley rats (2 mg/kg bw i. v.). Moreover, the ability of tryptanthrin to cross the blood-brain barrier was evaluated in three in vitro human and animal blood-brain barrier models. Bioanalytical UPLC-MS/MS methods used were validated according to current international guidelines. A half-life of 40.63 ± 6.66 min and a clearance of 1.00 ± 0.36 L/h/kg were found in the in vivo pharmacokinetic study. In vitro data obtained with the two primary animal blood-brain barrier models showed a good correlation with an immortalized human monoculture blood-brain barrier model (hBMEC cell line), and were indicative of a high blood-brain barrier permeation potential of tryptanthrin. These findings were corroborated by the in silico prediction of blood-brain barrier penetration. P-glycoprotein interaction of tryptanthrin was assessed by calculation of the efflux ratio in bidirectional permeability assays. An efflux ratio below 2 indicated that tryptanthrin is not subjected to active efflux., (Georg Thieme Verlag KG Stuttgart · New York.)
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- 2016
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171. Selection of a Relevant In Vitro Blood-Brain Barrier Model to Investigate Pro-Metastatic Features of Human Breast Cancer Cell Lines.
- Author
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Drolez A, Vandenhaute E, Julien S, Gosselet F, Burchell J, Cecchelli R, Delannoy P, Dehouck MP, and Mysiorek C
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- Animals, Antigens, CD34 metabolism, Cattle, Cell Differentiation physiology, Cell Line, Tumor, Cells, Cultured, Female, Fetal Blood cytology, Humans, Mice, Neuroglia cytology, Neuroglia metabolism, Pericytes cytology, Pericytes metabolism, Rats, Blood-Brain Barrier cytology, Breast Neoplasms pathology
- Abstract
Around 7-17% of metastatic breast cancer patients will develop brain metastases, associated with a poor prognosis. To reach the brain parenchyma, cancer cells need to cross the highly restrictive endothelium of the Blood-Brain Barrier (BBB). As treatments for brain metastases are mostly inefficient, preventing cancer cells to reach the brain could provide a relevant and important strategy. For that purpose an in vitro approach is required to identify cellular and molecular interaction mechanisms between breast cancer cells and BBB endothelium, notably at the early steps of the interaction. However, while numerous studies are performed with in vitro models, the heterogeneity and the quality of BBB models used is a limitation to the extrapolation of the obtained results to in vivo context, showing that the choice of a model that fulfills the biological BBB characteristics is essential. Therefore, we compared pre-established and currently used in vitro models from different origins (bovine, mice, human) in order to define the most appropriate tool to study interactions between breast cancer cells and the BBB. On each model, the BBB properties and the adhesion capacities of breast cancer cell lines were evaluated. As endothelial cells represent the physical restriction site of the BBB, all the models consisted of endothelial cells from animal or human origins. Among these models, only the in vitro BBB model derived from human stem cells both displayed BBB properties and allowed measurement of meaningful different interaction capacities of the cancer cell lines. Importantly, the measured adhesion and transmigration were found to be in accordance with the cancer cell lines molecular subtypes. In addition, at a molecular level, the inhibition of ganglioside biosynthesis highlights the potential role of glycosylation in breast cancer cells adhesion capacities.
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- 2016
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172. Development and validation of a LC-MS/MS method for assessment of an anti-inflammatory indolinone derivative by in vitro blood-brain barrier models.
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Jähne EA, Eigenmann DE, Culot M, Cecchelli R, Walter FR, Deli MA, Tremmel R, Fricker G, Smiesko M, Hamburger M, and Oufir M
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- ATP Binding Cassette Transporter, Subfamily B, Member 1 metabolism, Animals, Biological Transport, Cattle, Cells, Cultured, Endothelial Cells metabolism, Endothelium, Vascular metabolism, Fluoresceins chemistry, Fluoresceins metabolism, Humans, Male, Rats, Rats, Sprague-Dawley, Rats, Wistar, Swine, Anti-Inflammatory Agents chemistry, Anti-Inflammatory Agents metabolism, Blood-Brain Barrier metabolism, Chromatography, Liquid methods, Indoles chemistry, Indoles metabolism, Tandem Mass Spectrometry methods
- Abstract
The compound (E,Z)-3-(4-hydroxy-3,5-dimethoxybenzylidene)indolin-2-one (indolinone) was identified from lipophilic woad extracts (Isatis tinctoria L., Brassicaceae) as a compound possessing potent histamine release inhibitory and anti-inflammatory properties [1]. To further evaluate the potential of indolinone in terms of crossing the blood-brain barrier (BBB), we screened the compound in several in vitro cell-based human and animal BBB models. Therefore, we developed a quantitative LC-MS/MS method for the compound in modified Ringer HEPES buffer (RHB) and validated it according to FDA and EMA guidelines [2,3]. The calibration curve of indolinone in the range between 30.0 and 3000ng/ml was quadratic, and the limit of quantification was 30.0ng/ml. Dilution of samples up to 100-fold did not affect precision and accuracy. The carry-over was within acceptance criteria. Indolinone proved to be stable in RHB for 3h at room temperature (RT), and for three successive freeze/thaw cycles. The processed samples could be stored in the autosampler at 10°C for at least 28h. Moreover, indolinone was stable for at least 16 days in RHB when stored below -65°C. This validation study demonstrates that our method is specific, selective, precise, accurate, and capable to produce reliable results. In the immortalized human BBB mono-culture model, the apparent permeability coefficient from apical to basolateral (PappA→B), and the Papp from basolateral to apical (PappB→A) were 19.2±0.485×10(-6)cm/s and 21.7±0.326×10(-6)cm/s, respectively. For the primary rat/bovine BBB co-culture model a PappA→B of 27.1±1.67×10(-6)cm/s was determined. In the primary rat BBB triple co-culture model, the PappA→B and the PappB→A were 56.2±3.63×10(-6)cm/s and 34.6±1.41×10(-6)cm/s, respectively. The data obtained with the different models showed good correlation and were indicative of a high BBB permeation potential of indolinone confirmed by in silico prediction calculations. P-glycoprotein (P-gp) interaction for indolinone was studied with the aid of a calcein-AM uptake assay, and by calculation of the efflux ratio (ER) from the bidirectional permeability assays. For both bidirectional BBB models an ER below 2 was calculated, indicating that no active mediated transport mechanism is involved for indolinone. In porcine brain capillary endothelial cells (PBCECs), the calcein-AM uptake assay demonstrated that indolinone is neither a P-gp substrate nor a P-gp inhibitor and is accumulated into cells at high extent., (Copyright © 2014 Elsevier B.V. All rights reserved.)
- Published
- 2014
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173. Stroke-induced brain parenchymal injury drives blood-brain barrier early leakage kinetics: a combined in vivo/in vitro study.
- Author
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Kuntz M, Mysiorek C, Pétrault O, Pétrault M, Uzbekov R, Bordet R, Fenart L, Cecchelli R, and Bérézowski V
- Subjects
- Animals, Blood-Brain Barrier metabolism, Blood-Brain Barrier pathology, Cell Death, Cells, Cultured, Coculture Techniques, Disease Models, Animal, Endothelial Cells metabolism, Endothelial Cells ultrastructure, Glucose metabolism, Kinetics, Magnetic Resonance Imaging, Male, Mice, Mice, Inbred C57BL, Neuroglia metabolism, Neuroglia ultrastructure, Oxygen metabolism, Stroke metabolism, Stroke pathology, Blood-Brain Barrier physiopathology, Capillary Permeability physiology, Stroke physiopathology
- Abstract
The disappointing clinical outcomes of neuroprotectants challenge the relevance of preclinical stroke models and data in defining early cerebrovascular events as potential therapeutic targets. The kinetics of blood-brain barrier (BBB) leakage after reperfusion and the link with parenchymal lesion remain debated. By using in vivo and in vitro approaches, we conducted a kinetic analysis of BBB dysfunction during early reperfusion. After 60 minutes of middle cerebral artery occlusion followed by reperfusion times up to 24 hours in mice, a non-invasive magnetic resonance imaging method, through an original sequence of diffusion-weighted imaging, determined brain water mobility in microvascular compartments (D*) apart from parenchymal compartments (apparent diffusion coefficient). An increase in D* found at 4 hours post reperfusion concurred with the onset of both Evans blue/Dextran extravasations and in vitro BBB opening under oxygen-glucose deprivation and reoxygenation (R). The BBB leakage coincided with an emerging cell death in brain tissue as well as in activated glial cells in vitro. The co-culture of BBB endothelial and glial cells evidenced a recovery of endothelium tightness when glial cells were absent or non-injured during R. Preserving the ischemic brain parenchymal cells within 4 hours of reperfusion may improve therapeutic strategies for cerebrovascular protection against stroke.
- Published
- 2014
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174. Maternal calorie restriction modulates placental mitochondrial biogenesis and bioenergetic efficiency: putative involvement in fetoplacental growth defects in rats.
- Author
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Mayeur S, Lancel S, Theys N, Lukaszewski MA, Duban-Deweer S, Bastide B, Hachani J, Cecchelli R, Breton C, Gabory A, Storme L, Reusens B, Junien C, Vieau D, and Lesage J
- Subjects
- Animals, Efficiency physiology, Female, Fetal Growth Retardation metabolism, Male, Maternal-Fetal Exchange physiology, Mitochondria metabolism, Placenta physiology, Placenta ultrastructure, Placental Circulation physiology, Pregnancy, Rats, Rats, Wistar, Caloric Restriction adverse effects, Energy Metabolism physiology, Fetal Growth Retardation etiology, Maternal Nutritional Physiological Phenomena, Mitochondria physiology, Placenta metabolism
- Abstract
Low birth weight is associated with an increased risk for developing type 2 diabetes and metabolic diseases. The placental capacity to supply nutrients and oxygen to the fetus represents the main determiner of fetal growth. However, few studies have investigated the effects of maternal diet on the placenta. We explored placental adaptive proteomic processes implicated in response to maternal undernutrition. Rat term placentas from 70% food-restricted (FR30) mothers were used for a proteomic screen. Placental mitochondrial functions were evaluated using molecular and functional approaches, and ATP production was measured. FR30 drastically reduced placental and fetal weights. FR30 placentas displayed 14 proteins that were differentially expressed, including several mitochondrial proteins. FR30 induced a marked increase in placental mtDNA content and changes in mitochondrial functions, including modulation of the expression of genes implicated in biogenesis and bioenergetic pathways. FR30 mitochondria showed higher oxygen consumption but failed to maintain their ATP production. Maternal undernutrition induces placental mitochondrial abnormalities. Although an increase in biogenesis and bioenergetic efficiency was noted, placental ATP level was reduced. Our data suggest that placental mitochondrial defects may be implicated in fetoplacental pathologies.
- Published
- 2013
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175. Brain pericytes from stress-susceptible pigs increase blood-brain barrier permeability in vitro.
- Author
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Vandenhaute E, Culot M, Gosselet F, Dehouck L, Godfraind C, Franck M, Plouët J, Cecchelli R, Dehouck MP, and Ruchoux MM
- Abstract
Background: The function of pericytes remains questionable but with improved cultured technique and the use of genetically modified animals, it has become increasingly clear that pericytes are an integral part of blood-brain barrier (BBB) function, and the involvement of pericyte dysfunction in certain cerebrovascular diseases is now emerging. The porcine stress syndrome (PSS) is the only confirmed, homologous model of malignant hyperthermia (MH) in veterinary medicine. Affected animals can experience upon slaughter a range of symptoms, including skeletal muscle rigidity, metabolic acidosis, tachycardia and fever, similar to the human syndrome. Symptoms are due to an enhanced calcium release from intracellular stores. These conditions are associated with a point mutation in ryr1/hal gene, encoding the ryanodine receptor, a calcium channel. Important blood vessel wall muscle modifications have been described in PSS, but potential brain vessel changes have never been documented in this syndrome., Methods: In the present work, histological and ultrastructural analyses of brain capillaries from wild type and ryr1 mutated pigs were conducted to investigate the potential impairment of pericytes, in this pathology. In addition, brain pericytes were isolated from the three porcine genotypes (wild-type NN pigs; Nn and nn pigs, bearing one or two (n) mutant ryr1/hal alleles, respectively), and tested in vitro for their influence on the permeability of BBB endothelial monolayers., Results: Enlarged perivascular spaces were observed in ryr1-mutant samples, corresponding to a partial or total detachment of the astrocytic endfeet. These spaces were electron lucent and sometimes filled with lipid deposits and swollen astrocytic feet. At the ultrastructural level, brain pericytes did not seem to be affected because they showed regular morphology and characteristics, so we aimed to check their ability to maintain BBB properties in vitro. Our results indicated that pericytes from the three genotypes of pigs had differing influences on the BBB. Unlike pericytes from NN pigs, pericytes from Nn and nn pigs were not able to maintain low BBB permeability., Conclusions: Electron microscopy observations demonstrated brain capillary modifications in PSS condition, but no change in pericyte morphology. Results from in vitro experiments suggest that brain pericytes from ryr1 mutated pigs, even if they are not affected by this condition at the ultrastructural level, are not able to maintain BBB integrity in comparison with pericytes from wild-type animals.
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- 2012
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176. Case study: adapting in vitro blood-brain barrier models for use in early-stage drug discovery.
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Vandenhaute E, Sevin E, Hallier-Vanuxeem D, Dehouck MP, and Cecchelli R
- Subjects
- Animals, Biological Transport, Case-Control Studies, Humans, Pharmacokinetics, Blood-Brain Barrier metabolism, Drug Discovery, Models, Biological, Pharmaceutical Preparations chemistry, Pharmaceutical Preparations metabolism
- Abstract
Several parameters influencing the brain distribution of compounds must be considered when designing potential neuropharmaceuticals in early-stage drug discovery. The blood-brain barrier (BBB) represents an obstacle for drug penetration into the brain. Many in vitro BBB models have proven useful for predicting the BBB permeation rate, but do not meet all criteria for use in early-stage drug discovery: feasibility, rapidity, reliability and a low requirement for human resources. To meet this demand, we have developed a robust, higher-throughput, cell-based model exhibiting BBB features (low paracellular permeability, functional efflux pumps and the correct endothelial phenotype). This system comes in a ready-to-use, frozen format, appropriate for in-house use by large pharmaceutical firms and small biotech companies during early-stage drug discovery., (Copyright © 2011 Elsevier Ltd. All rights reserved.)
- Published
- 2012
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177. [Dysfunction of the blood-brain barrier during ischaemia: a therapeutic concern].
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Bérézowski V, Mysiorek C, Kuntz M, Pétrault O, and Cecchelli R
- Subjects
- Endothelial Cells physiology, Fibrinolytic Agents adverse effects, Humans, Neuroprotective Agents, Reperfusion Injury physiopathology, Stroke drug therapy, Thrombolytic Therapy, Blood-Brain Barrier physiopathology, Brain Ischemia drug therapy
- Abstract
Since it was discovered and its brain-protective role characterized, the blood-brain barrier (BBB), through the permeability-restricting action of the brain capillary endothelial cells, has been representing a hurdle for 95% of new medical compounds targeting the central nervous system. Recently, a BBB dysfunction is being found in an increasing number of pathologies such as brain ischaemic stroke, whose only therapy consists in a pharmacological thrombolysis limited to a small percentage of the admitted patients, because of the toxical effects of thrombolytics. And since the clinical failure of promising neuroprotectants, numerous studies of brain ischaemia were carried out, with physiopathological or pharmacological approaches refocused on the BBB, whose structural complexity is now expanded to perivascular cells, all forming a functional unit named the neurovascular unit (NVU). Nevertheless, in spite of the numerous molecular mechanisms identified, the process of BBB dysfunction in the ischaemia/reperfusion cascade remains insufficiently established to explain the pleiotropic action exerted by new pharmacological compounds, possibly protecting the entire NVU and representing potential treatments., (© Société de Biologie, 2012.)
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- 2012
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178. [Role of the blood-brain barrier in Alzheimer's disease].
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Gosselet F, Candela P, Cecchelli R, and Fenart L
- Subjects
- Alzheimer Disease etiology, Alzheimer Disease metabolism, Alzheimer Disease therapy, Amyloid beta-Peptides metabolism, Amyloid beta-Peptides physiology, Animals, Blood-Brain Barrier metabolism, Blood-Brain Barrier pathology, Brain blood supply, Brain metabolism, Brain physiopathology, Cell Membrane Permeability physiology, Drug Delivery Systems methods, Humans, Models, Biological, Pericytes metabolism, Pericytes physiology, Protein Transport physiology, Alzheimer Disease physiopathology, Blood-Brain Barrier physiology
- Abstract
The blood-brain barrier (BBB), which isolates the brain from the whole body, restricts exchanges between the brain and the peripheral compartments. Several studies highlight the importance of this barrier in neurodegenerative diseases such as Alzheimer's disease (AD). This pathology is characterized by an abnormal accumulation and aggregation of Aβ peptides which contribute to the neurodegenerative processes and the BBB seems to play a key role for the brain Aβ peptides metabolism. This review focuses on recent data demonstrating the important role of the BBB in AD, suggesting that this barrier is a possible new therapeutic target in this disease., (© 2011 médecine/sciences – Inserm / SRMS.)
- Published
- 2011
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179. PPACK-Desmodus rotundus salivary plasminogen activator (cDSPAalpha1) prevents the passage of tissue-type plasminogen activator (rt-PA) across the blood-brain barrier and neurotoxicity.
- Author
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Roussel BD, Hommet Y, Macrez R, Schulz T, Petersen KU, Berezowski V, Cecchelli R, Ali C, and Vivien D
- Subjects
- Animals, Biotinylation, Chiroptera, Fibrinolytic Agents therapeutic use, Humans, Ischemia therapy, Mice, Recombinant Proteins therapeutic use, Stroke therapy, Blood-Brain Barrier metabolism, Neurons pathology, Plasminogen Activators metabolism, Tissue Plasminogen Activator metabolism
- Published
- 2009
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180. Peroxisome-proliferator-activated receptor-alpha activation protects brain capillary endothelial cells from oxygen-glucose deprivation-induced hyperpermeability in the blood-brain barrier.
- Author
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Mysiorek C, Culot M, Dehouck L, Derudas B, Staels B, Bordet R, Cecchelli R, Fenart L, and Berezowski V
- Subjects
- Animals, Animals, Newborn, Blood-Brain Barrier metabolism, Capillary Permeability drug effects, Capillary Permeability genetics, Cell Proliferation drug effects, Cells, Cultured, Claudin-5, Coculture Techniques methods, Dose-Response Relationship, Drug, Endothelial Cells drug effects, Fenofibrate analogs & derivatives, Fenofibrate pharmacology, Gene Expression Regulation drug effects, Gene Expression Regulation genetics, Hypolipidemic Agents pharmacology, Hypoxia pathology, Hypoxia prevention & control, Membrane Proteins metabolism, Mice, Mice, Inbred C57BL, Mice, Knockout, Neuroglia drug effects, Neuroglia physiology, PPAR alpha deficiency, Time Factors, von Willebrand Factor immunology, von Willebrand Factor metabolism, Blood-Brain Barrier physiology, Brain cytology, Capillary Permeability physiology, Endothelial Cells metabolism, Glucose deficiency, PPAR alpha metabolism
- Abstract
That promising neuroprotectants failed to demonstrate benefit against stroke highlights the great difficulties to translate preclinical pharmacological effects in clinical outcomes. Part of this hurdle implies the complex response to injury of the neurovascular unit increasing the cerebrovascular permeability at the level of the blood-brain barrier (BBB). Previous studies reported neuroprotection in animal models upon activation of the nuclear receptor PPARalpha(peroxisome proliferator-activated receptor)alpha, but the cellular targets at the BBB level remain largely unexplored. Here, to study whether PPAR-alpha activation acts on BBB permeability, we adapted a mouse BBB cell model to ischaemic conditions at the stage of occlusion defined in vitro as oxygen-glucose deprivation (OGD). This model consists of a co-culture of brain capillary endothelial cells (ECs) on a filter insert placed upon a rat glial cell culture. The EC monolayer permeability increase induced by 4 h of OGD was significantly restricted after treatment with the PPAR-alpha agonist fenofibric acid (FA) 24 h before or at the onset of OGD. Treatments of separated ECs or glial cells showed that this protective effect was conferred by BBB ECs but not glial cells. Furthermore, co-cultures with ECs from PPAR-alpha-deficient mice revealed that FA had no effect on OGD-induced hyperpermeability. No transcriptional modulation of classical PPAR-alpha target genes such as SOD, ICAM-1, VCAM-1, ACO, CPT-1, PDK-4 or ET-1 was observed in wild type mouse ECs. In conclusion, these results suggest that part of the preventive PPAR-alpha-mediated protection may occur via BBB ECs by limiting hyperpermeability.
- Published
- 2009
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181. A polarized localization of amino acid/carnitine transporter B(0,+) (ATB(0,+)) in the blood-brain barrier.
- Author
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Czeredys M, Mysiorek C, Kulikova N, Samluk Ł, Berezowski V, Cecchelli R, and Nałecz KA
- Subjects
- Amino Acids, Cyclic pharmacology, Animals, Blood-Brain Barrier drug effects, Carnitine metabolism, Cattle, Leucine metabolism, Rats, Amino Acid Transport System ASC metabolism, Amino Acid Transport Systems, Neutral metabolism, Blood-Brain Barrier metabolism
- Abstract
Brain capillary endothelial cells control the uptake and efflux from the brain of many hydrophilic compounds due to highly specialized transporters often localized in a polarized way. Localization of Na(+)- and Cl(-)-dependent amino acid and carnitine transporter B(0,+) (ATB(0,+)) was studied in a co-culture of bovine brain capillary endothelial cells (BBCEC) grown on filters above astrocytes (an in vitro blood-brain barrier model). Immunoblotting and three-dimensional immunocytochemistry analysis with anti-B(0,+)antibodies demonstrated the presence of this transporter and its prevalent co-localization with P-glycoprotein i.e. at the apical side. The sensitivity of leucine uptake through the apical membrane to 2-aminobicyclo-[2.2.1]-heptane-2-carboxylic acid (BCH), D-serine as well as sodium and chloride replacement confirm the functioning of ATB(0,+) and suggests an important physiological role of ATB(0,+) in controlling the delivery of amino acids and carnitine to the brain.
- Published
- 2008
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182. Recombinant Desmodus rotundus salivary plasminogen activator crosses the blood-brain barrier through a low-density lipoprotein receptor-related protein-dependent mechanism without exerting neurotoxic effects.
- Author
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López-Atalaya JP, Roussel BD, Ali C, Maubert E, Petersen KU, Berezowski V, Cecchelli R, Orset C, and Vivien D
- Subjects
- Animals, Blood-Brain Barrier drug effects, Cattle, Cells, Cultured, Chiroptera, Coculture Techniques, Fibrinolytic Agents toxicity, Humans, Male, Plasminogen Activators toxicity, Rats, Rats, Sprague-Dawley, Recombinant Proteins toxicity, Blood-Brain Barrier metabolism, Fibrinolytic Agents pharmacokinetics, Low Density Lipoprotein Receptor-Related Protein-1 physiology, Plasminogen Activators pharmacokinetics, Recombinant Proteins pharmacokinetics
- Abstract
Background and Purpose: Desmoteplase, a recombinant form of the plasminogen activator DSPAalpha1 from Desmodus rotundus, may offer improved clinical benefits for acute ischemic stroke treatment over the current therapy, recombinant tissue plasminogen activator (rtPA). Accumulating evidence suggests that clinical use of rtPA could be limited by unfavorable properties, including its ability to cross the blood-brain barrier (BBB), thus potentially adding to the pro-excitotoxic effect of endogenous tPA in cerebral parenchyma. Here, to investigate whether desmoteplase may display a safer profile than the structurally-related tPA, both agents were compared for their ability to cross the BBB and promote neurotoxicity., Methods: First, the passage of vascular DSPA and rtPA was investigated in vitro in a model of BBB, subjected or not to oxygen and glucose deprivation. Second, we studied DSPA- and rtPA-mediated effects in an in vivo paradigm of excitotoxic necrosis., Results: The rtPA and desmoteplase cross the intact BBB by LRP-mediated transcytosis. Under conditions of oxygen and glucose deprivation, translocation rates of both compounds increased; however, unlike rtPA, desmoteplase transport remained LRP-dependent. Additionally, neither intracerebral nor intravenous desmoteplase administration enhanced NMDA-induced excitotoxic striatal damage in vivo. Interestingly, intravenous but not intrastriatal coadministration of desmoteplase and rtPA reduced the pro-excitotoxic effect of rtPA., Conclusions: We show that desmoteplase crosses the BBB but does not promote neuronal death. Moreover, intravenous administration of desmoteplase antagonizes the neurotoxicity induced by vascular rtPA. This action may be caused by competition of desmoteplase with rtPA for LRP binding at the BBB, thus effectively blocking rtPA access to the brain parenchyma.
- Published
- 2007
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183. Oxygen glucose deprivation switches the transport of tPA across the blood-brain barrier from an LRP-dependent to an increased LRP-independent process.
- Author
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Benchenane K, Berezowski V, Fernández-Monreal M, Brillault J, Valable S, Dehouck MP, Cecchelli R, Vivien D, Touzani O, and Ali C
- Subjects
- Animals, Brain Ischemia etiology, Cell Hypoxia, Cytoplasmic Vesicles chemistry, Endothelium, Vascular cytology, Endothelium, Vascular ultrastructure, Glucose physiology, Infarction, Middle Cerebral Artery complications, LDL-Receptor Related Proteins antagonists & inhibitors, Male, Mice, Protein Transport, Tissue Plasminogen Activator analysis, Tissue Plasminogen Activator pharmacology, Blood-Brain Barrier metabolism, Brain Ischemia metabolism, LDL-Receptor Related Proteins physiology, Tissue Plasminogen Activator metabolism
- Abstract
Background and Purpose: Despite uncontroversial benefit from its thrombolytic activity, the documented neurotoxic effect of tissue plasminogen activator (tPA) raises an important issue: the current emergency stroke treatment might not be optimum if exogenous tPA can enter the brain and thus add to the deleterious effects of endogenous tPA within the cerebral parenchyma. Here, we aimed at determining whether vascular tPA crosses the blood-brain barrier (BBB) during cerebral ischemia, and if so, by which mechanism., Methods: First, BBB permeability was assessed in vivo by measuring Evans Blue extravasation following intravenous injection at 0 or 3 hours after middle cerebral artery electrocoagulation in mice. Second, the passage of vascular tPA was investigated in an in vitro model of BBB, subjected or not to oxygen and glucose deprivation (OGD)., Results: We first demonstrated that after focal permanent ischemia in mice, the BBB remains impermeable to Evans Blue in the early phase (relative to the therapeutic window of tPA), whereas at later time points massive Evans Blue extravasation occurs. Then, the passage of tPA during these 2 phases, was investigated in vitro and we show that in control conditions, tPA crosses the intact BBB by a low-density lipoprotein (LDL) receptor-related protein (LRP)-dependent transcytosis, whereas OGD leads to an exacerbation of tPA passage, which switches to a LRP-independent process., Conclusions: We evidence 2 different mechanisms through which vascular tPA can reach the brain parenchyma, depending on the state of the BBB. As discussed, these data show the importance of taking the side effects of blood-derived tPA into account and offer a basis to improve the current thrombolytic strategy.
- Published
- 2005
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184. Specific AHNAK expression in brain endothelial cells with barrier properties.
- Author
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Gentil BJ, Benaud C, Delphin C, Remy C, Berezowski V, Cecchelli R, Feraud O, Vittet D, and Baudier J
- Subjects
- Angiopoietin-1 metabolism, Angiopoietin-1 pharmacology, Animals, Animals, Newborn, Blood-Brain Barrier ultrastructure, Brain Neoplasms blood supply, Brain Neoplasms ultrastructure, Cattle, Cell Communication physiology, Cell Differentiation physiology, Cell Line, Cell Membrane ultrastructure, Choroid Plexus metabolism, Choroid Plexus ultrastructure, Coculture Techniques, Cytosol metabolism, Endothelial Cells ultrastructure, Male, Mice, Neuroglia metabolism, Phosphoproteins metabolism, Protein Transport drug effects, Protein Transport physiology, Rats, Rats, Wistar, Tight Junctions metabolism, Tight Junctions ultrastructure, Up-Regulation drug effects, Up-Regulation physiology, Zonula Occludens-1 Protein, Blood-Brain Barrier metabolism, Brain blood supply, Brain Neoplasms metabolism, Cell Membrane metabolism, Endothelial Cells metabolism, Membrane Proteins metabolism, Neoplasm Proteins metabolism
- Abstract
The blood-brain barrier (BBB) is essential for maintaining brain homeostasis and low permeability. Because disruption of the BBB may contribute to many brain disorders, they are of considerable interests in the identification of the molecular mechanisms of BBB development and integrity. We here report that the giant protein AHNAK is expressed at the plasma membrane of endothelial cells (ECs) forming specific blood-tissue barriers, but is absent from the endothelium of capillaries characterized by extensive molecular exchanges between blood and extracellular fluid. In the brain, AHNAK is widely distributed in ECs with BBB properties, where it co-localizes with the tight junction protein ZO-1. AHNAK is absent from the permeable brain ECs of the choroid plexus and is down-regulated in permeable angiogenic ECs of brain tumors. In the choroid plexus, AHNAK accumulates at the tight junctions of the choroid epithelial cells that form the blood-cerebrospinal fluid (CSF) barrier. In EC cultures, the regulation of AHNAK expression and its localization corresponds to general criteria of a protein involved in barrier organization. AHNAK is up-regulated by angiopoietin-1 (Ang-1), a morphogenic factor that regulates brain EC permeability. In bovine cerebral ECs co-cultured with glial cells, AHNAK relocates from the cytosol to the plasma membrane when endothelial cells acquire BBB properties. Our results identify AHNAK as a protein marker of endothelial cells with barrier properties., (Copyright 2004 Wiley-Liss, Inc.)
- Published
- 2005
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185. [Cerebral transfer and neuroprotection].
- Author
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Fenart L, Berezowski V, Dehouck MP, and Cecchelli R
- Subjects
- Animals, Brain anatomy & histology, Brain physiology, Brain Chemistry, Endothelium, Vascular physiology, Humans, Neuroglia physiology, Neuroprotective Agents metabolism, Blood-Brain Barrier physiology, Neuroprotective Agents pharmacology
- Abstract
In contrast to other vascular beds, the endothelial cells in brain capillaries, which constitute the blood-brain barrier, are sealed together by continuous tight junctions and have little transcellular vesicular transport. In addition to these morphological properties, the presence of specific enzymes and proteins highly restricts the passage of molecules from the blood to the brain. To provide an in vitro system for studying brain capillary functions, we have developed a process of coculture that closely mimics the in vivo situation by culturing brain capillary endothelial cells on one side of a filter and glial cells on the other. In these culture conditions, endothelial cells retain all the endothelial cell markers and the characteristics of the blood-brain barrier, including gamma-glutamyl transpeptidase and P-glycoprotein activities. Moreover, the close correlation between the results obtained in vitro with our model and in vivo allows us to conclude that our in vitro blood-brain barrier model is a relevant model for the screening of new molecules to the brain.
- Published
- 2004
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186. [3H]BHDP as a novel and selective ligand for sigma1 receptors in liver mitochondria and brain synaptosomes of the rat.
- Author
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Klouz A, Tillement JP, Boussard MF, Wierzbicki M, Berezowski V, Cecchelli R, Labidalle S, Onténiente B, and Morin D
- Subjects
- Adenosine Triphosphate metabolism, Animals, Astrocytes cytology, Astrocytes drug effects, Astrocytes metabolism, Brain cytology, Cells, Cultured, Hydrogen-Ion Concentration, Hypoxia metabolism, Ligands, Piperazines pharmacology, Protein Binding, Rats, Substrate Specificity, Time Factors, Tritium metabolism, Brain metabolism, Mitochondria, Liver metabolism, Piperazines metabolism, Receptors, sigma metabolism, Synaptosomes metabolism
- Abstract
The binding profile of [(3)H]BHDP ([(3)H]N-benzyl-N'-(2-hydroxy-3,4-dimethoxybenzyl)-piperazine) was evaluated. [(3)H]BHDP labelled a single class of binding sites with high affinity (K(d)=2-3 nM) in rat liver mitochondria and synaptic membranes. The pharmacological characterization of these sites using sigma reference compounds revealed that these sites are sigma receptors and, more particularly, sigma1 receptors. Indeed, BHDP inhibited [(3)H]pentazocine binding, a marker for sigma1 receptors, with high affinity in a competitive manner. BHDP is selective for sigma1 receptors since it did not show any relevant affinity for most of the other receptors, ion channels or transporters tested. Moreover, in an in vitro model of cellular hypoxia, BHDP prevented the fall in adenosine triphosphate (ATP) levels caused by 24 h hypoxia in cultured astrocytes. Taken together, these results demonstrate that [(3)H]BHDP is a potent and selective ligand for sigma1 receptors showing cytoprotective effects in astrocytes.
- Published
- 2003
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187. Peroxisome proliferator-activated receptor-alpha activation as a mechanism of preventive neuroprotection induced by chronic fenofibrate treatment.
- Author
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Deplanque D, Gelé P, Pétrault O, Six I, Furman C, Bouly M, Nion S, Dupuis B, Leys D, Fruchart JC, Cecchelli R, Staels B, Duriez P, and Bordet R
- Subjects
- Animals, Apolipoproteins E deficiency, Apolipoproteins E genetics, Cerebral Infarction etiology, Cerebrovascular Circulation drug effects, Disease Models, Animal, Infarction, Middle Cerebral Artery complications, Infarction, Middle Cerebral Artery drug therapy, Intercellular Adhesion Molecule-1 metabolism, Mice, Mice, Inbred C57BL, Mice, Knockout, Mice, Transgenic, Middle Cerebral Artery drug effects, Middle Cerebral Artery physiopathology, Nitric Oxide Synthase metabolism, Nitric Oxide Synthase Type II, Nitric Oxide Synthase Type III, Oxidative Stress drug effects, Rats, Rats, Inbred WKY, Receptors, Cytoplasmic and Nuclear drug effects, Stroke etiology, Stroke physiopathology, Time, Transcription Factors drug effects, Vascular Cell Adhesion Molecule-1 metabolism, Vasodilation drug effects, Cerebral Infarction prevention & control, Fenofibrate pharmacology, Neuroprotective Agents pharmacology, Receptors, Cytoplasmic and Nuclear metabolism, Stroke drug therapy, Transcription Factors metabolism
- Abstract
The treatment of ischemic strokes is limited to the prevention of cerebrovascular risk factors and to the modulation of the coagulation cascade during the acute phase. A new therapeutic strategy could be to preventively protect the brain against noxious biological reactions induced by cerebral ischemia such as oxidative stress and inflammation to minimize their neurological consequences. Here, we show that a peroxisome proliferator-activated receptor (PPAR-alpha) activator, fenofibrate, protects against cerebral injury by anti-oxidant and anti-inflammatory mechanisms. A 14 d preventive treatment with fenofibrate reduces susceptibility to stroke in apolipoprotein E-deficient mice as well as decreases cerebral infarct volume in C57BL/6 wild-type mice. The neuroprotective effect of fenofibrate is completely absent in PPAR-alpha-deficient mice, suggesting that PPAR-alpha activation is involved as a mechanism of the protection against cerebral injury. Furthermore, this neuroprotective effect appears independently of any improvement in plasma lipids or glycemia and is associated with (1) an improvement in middle cerebral artery sensitivity to endothelium-dependent relaxation unrelated to an increase in nitric oxide synthase (NOS) type III expression, (2) a decrease in cerebral oxidative stress depending on the increase in numerous antioxidant enzyme activities, and (3) the prevention of ischemia-induced expression of vascular cell adhesion molecule-1 and intercellular adhesion molecule-1 in cerebral vessels without any change in NOS II expression. These data demonstrate that PPAR-alpha could be a new pharmacological target to preventively reduce the deleterious neurological consequences of stroke in mice and suggest that PPAR-alpha activators could preventively decrease the severity of stroke in humans.
- Published
- 2003
188. Protein transport in cerebral endothelium. In vitro transcytosis of transferrin.
- Author
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Fenart L and Cecchelli R
- Subjects
- Animals, Biological Transport, Active, Capillaries cytology, Capillaries metabolism, Cattle, Endocytosis, Endothelium, Vascular cytology, Fluorescein-5-isothiocyanate, In Vitro Techniques, Iodine Radioisotopes, Iron Radioisotopes, Kinetics, Microscopy, Fluorescence, Models, Biological, Temperature, Blood-Brain Barrier physiology, Brain blood supply, Brain metabolism, Endothelium, Vascular metabolism, Transferrin pharmacokinetics
- Published
- 2003
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189. Intercommunications between brain capillary endothelial cells and glial cells increase the transcellular permeability of the blood-brain barrier during ischaemia.
- Author
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Brillault J, Berezowski V, Cecchelli R, and Dehouck MP
- Subjects
- Animals, Apoproteins pharmacokinetics, Blood-Brain Barrier physiology, Brain blood supply, Capillaries cytology, Cattle, Cell Hypoxia, Cells, Cultured, Coculture Techniques, Endothelium, Vascular cytology, Glucose deficiency, Glucose metabolism, Inulin pharmacokinetics, Neuroglia cytology, Rats, Sucrose pharmacokinetics, Transferrin pharmacokinetics, Brain Ischemia metabolism, Capillary Permeability physiology, Cell Communication physiology, Endothelium, Vascular metabolism, Neuroglia metabolism
- Abstract
Increased cerebrovascular permeability is an important factor in the development of cerebral oedema after stroke, implicating the blood-brain barrier (BBB). To investigate the effect of hypoxia on the permeability changes, we used a cell culture model of the BBB consisting of a co-culture of brain capillary endothelial cells and glial cells. When endothelial cells from this co-culture model were submitted alone to hypoxic conditions, long exposures (48 h) were necessary to result in an increase in endothelial cell monolayer permeability to [3H]inulin. When endothelial cells were incubated in presence of glial cells, a huge increase in permeability occurred after 9 h of hypoxic conditions. Oxygen glucose deprivation (OGD) resulted in a much shorter time (i.e. 2 h) required for an increase in permeability. We have demonstrated that this OGD-induced permeability increase involves a transcellular rather than a paracellular pathway. Conditioned medium experiments showed that glial cells secrete soluble permeability factors during OGD. However, endothelial cells have to be made sensitive by OGD in order to respond to these glial soluble factors. This work shows that an early cross-talk between glial and endothelial cells occurs during ischaemic stroke and alters BBB transcellular transport by means of glial factor secretions.
- Published
- 2002
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190. High transcytosis of melanotransferrin (P97) across the blood-brain barrier.
- Author
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Demeule M, Poirier J, Jodoin J, Bertrand Y, Desrosiers RR, Dagenais C, Nguyen T, Lanthier J, Gabathuler R, Kennard M, Jefferies WA, Karkan D, Tsai S, Fenart L, Cecchelli R, and Béliveau R
- Subjects
- Animals, Antigens, Neoplasm, Astrocytes cytology, Astrocytes metabolism, Brain blood supply, Brain metabolism, Capillaries cytology, Cells, Cultured, Coculture Techniques, Endothelium, Vascular cytology, Endothelium, Vascular metabolism, Female, Humans, Iodine Radioisotopes, Low Density Lipoprotein Receptor-Related Protein-1 metabolism, Male, Melanoma-Specific Antigens, Mice, Models, Biological, Neoplasm Proteins pharmacokinetics, Protein Binding physiology, Protein Transport drug effects, Protein Transport physiology, Rats, Sucrose pharmacokinetics, Transferrin pharmacology, Blood-Brain Barrier physiology, Neoplasm Proteins metabolism
- Abstract
The blood-brain barrier (BBB) performs a neuroprotective function by tightly controlling access to the brain; consequently it also impedes access of proteins as well as pharmacological agents to cerebral tissues. We demonstrate here that recombinant human melanotransferrin (P97) is highly accumulated into the mouse brain following intravenous injection and in situ brain perfusion. Moreover, P97 transcytosis across bovine brain capillary endothelial cell (BBCEC) monolayers is at least 14-fold higher than that of holo-transferrin, with no apparent intra-endothelial degradation. This high transcytosis of P97 was not related to changes in the BBCEC monolayer integrity. In addition, the transendothelial transport of P97 was sensitive to temperature and was both concentration- and conformation-dependent, suggesting that the transport of P97 is due to receptor-mediated endocytosis. In spite of the high degree of sequence identity between P97 and transferrin, a different receptor than the one for transferrin is involved in P97 transendothelial transport. A member of the low-density lipoprotein receptor protein family, likely LRP, seems to be involved in P97 transendothelial transport. The brain accumulation, high rate of P97 transcytosis and its very low level in the blood suggest that P97 could be advantageously employed as a new delivery system to target drugs directly to the brain.
- Published
- 2002
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191. Prediction of drug transport through the blood-brain barrier in vivo: a comparison between two in vitro cell models.
- Author
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Lundquist S, Renftel M, Brillault J, Fenart L, Cecchelli R, and Dehouck MP
- Subjects
- Animals, Astrocytes cytology, Biological Transport drug effects, Biological Transport physiology, Caco-2 Cells cytology, Cattle, Cells, Cultured, Endothelium, Vascular cytology, Forecasting, Humans, Rats, Astrocytes metabolism, Blood-Brain Barrier physiology, Caco-2 Cells metabolism, Endothelium, Vascular metabolism, Pharmaceutical Preparations metabolism
- Abstract
Purpose: Studies were conducted to evaluate whether the use of an in vitro model of the blood-brain barrier (BBB) resulted in more accurate predictions of the in vivo transport of compounds compared to the use of a human intestinal cell line (Caco-2)., Methods: The in vitro BBB model employs bovine brain capillary endothelial cells co-cultured with primary rat astrocytes. The Caco-2 cells originate from a human colorectal carcinoma. The rat was used as experimental animal for the in vivo studies., Results: Strong correlations (r = 0.93-0.95) were found between the results generated by the in vitro model of the BBB and two different methodologies to measure the permeability across the BBB in vivo. In contrast, a poor correlation (r = 0.68) was obtained between Caco-2 cell data and in vivo BBB transport. A relatively poor correlation (r = 0.74) was also found between the two in vitro models., Conclusion: The present study illustrates the limitations of the Caco-2 model to predict BBB permeability of compounds in vivo. The results emphasize the fact that the BBB and the intestinal mucosa are two fundamentally different biologic barriers, and to be able to make accurate predictions about the in vivo CNS penetration of potential drug candidates, it is important that the in vitro model possesses the main characteristics of the in vivo BBB.
- Published
- 2002
- Full Text
- View/download PDF
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