Evaluation of Blood-brain Barrier Transporters by Co-culture of Brain Capillary Endothelial Cells with Astrocytes

Kido, Yasuto; Tamai, Ikumi; Nakanishi, Takeo; Kagami, Toru; Hirosawa, Iori; Sai, Yoshimichi; Tsuji, Akira

doi:10.2133/dmpk.17.34

Cited by 32 publications

(20 citation statements)

References 25 publications

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“…These cells also have efflux transporters for cationic compounds (e.g., Pgp) and organic anions (5,25,41). It is noteworthy that many of the endogenous anions reportedly effluxed by cultured brain capillary endothe- (17,19,20). The expression of Mrp4 in the lumen of brain capillaries is compatible with the premise that Mrp4 protects the brain by restricting the movement of cytotoxic organic anions out of the capillaries.…”

Section: Discussionmentioning

confidence: 99%

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Mrp4 Confers Resistance to Topotecan and Protects the Brain from Chemotherapy

Leggas

Adachi

Scheffer

et al. 2004

Molecular and Cellular Biology

388

363

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The role of the multidrug resistance protein MRP4/ABCC4 in vivo remains undefined. To explore this role, we generated Mrp4-deficient mice. Unexpectedly, these mice showed enhanced accumulation of the anticancer agent topotecan in brain tissue and cerebrospinal fluid (CSF). Further studies demonstrated that topotecan was an Mrp4 substrate and that cells overexpressing Mrp4 were resistant to its cytotoxic effects. We then used new antibodies to discover that Mrp4 is unique among the anionic ATP-dependent transporters in its dual localization at the basolateral membrane of the choroid plexus epithelium and in the apical membrane The endothelial cells of the brain's capillaries are tightly joined to form a hydrophobic permeability barrier (32) termed the blood-brain barrier. Pgp expression in these cells limits the movement of hydrophobic cationic drugs from the blood into the brain (36,42,43). However, in vitro, these capillary endothelial cells also transport organic anions unidirectionally toward the capillary lumen in an energy-dependent fashion (5, 25, 41). Therefore, the capillary endothelial cells appear to express an unidentified anionic ABC transporter. Currently, it is unknown whether an anionic ABC transporter is expressed at functional levels in vivo in the endothelium of brain capillaries.The ABC transporter Mrp4, originally described as a nucleotide transporter (37), is known to transport a diverse array of compounds (2,7,34) and is capable of transporting organic anions as well as antiviral and antiretroviral compounds that do not easily penetrate the central nervous system (CNS) (2, 3, 9, 27, 37). Mrp4 expression was previously demonstrated on the basolateral membrane of the prostate gland and the apical membrane of the kidney (21, 44). Studies in cultured epithelial cells have demonstrated basolateral localization of Mrp4 (22). Transporters typically route to one surface in polarized cells. For instance, the Mrp (ABCC) subfamily members localize to either the basolateral or apical membrane, but not to both. MRP1 is restricted to the basolateral membrane of the choroid plexus and intestine, whereas MRP2 is found on the apical membrane in the intestine and liver (26,29). Mrp4 might be unique among the Mrp transporters in having cell-or tissuedependent polarized expression, but the biological importance of this unique ability to localize either apically or basolaterally remains unknown.We have developed Mrp4 knockout mice, and here we report their first use to show that Mrp4 is expressed in the lumen of brain capillaries and in the basolateral membrane in the choroid plexus epithelium. In vivo, Mrp4 restricts topotecan movement from the blood into the CSF and from the capillaries into the brain tissue by virtue of its unique ability to traffic to either the apical or basolateral membrane. We further show that Mrp4 overexpression confers resistance to the camptothecin topotecan. These studies have specific therapeutic implications for targeting the CNS that might harbor tumors but have more general impl...

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Section: Discussionmentioning

confidence: 99%

“…The fact that naphthol-glucuronide inhibits Mrp4 transport (44) suggests that it is an Mrp4 substrate. Methotrexate is another organic anion and Mrp4 substrate (19). Importantly, the transport of intraventricularly injected methotrexate from the choroid plexus into the blood is inhibited by probenecid, a reported Mrp4 inhibitor (38,44).…”

Section: Discussionmentioning

confidence: 99%

Mrp4 Confers Resistance to Topotecan and Protects the Brain from Chemotherapy

Leggas

Adachi

Scheffer

et al. 2004

Molecular and Cellular Biology

388

363

View full text Add to dashboard Cite

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“…Brain endothelial cells control the transport of substances between brain and blood through transporters and tight junction (TJ) proteins [3]. TJ proteins regulate the transport of ions via the development of high transendothelial electrical resistance (TEER) as well as the transport of molecules [4, 5], which create a physical blockade to paracellular diffusion [6]. This specialized phenotype is fundamental for protection of the brain.…”

Section: Introductionmentioning

confidence: 99%

Hydralazine is a Suitable Mimetic Agent of Hypoxia to Study the Impact of Hypoxic Stress on In Vitro Blood-Brain Barrier Model

Chatard

Puech

Perek

et al. 2017

Cell Physiol Biochem

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Background/Aims: Understanding cellular mechanisms induced by hypoxia is fundamental to reduce blood-brain barrier (BBB) disruption. Nevertheless, the investigation of hypoxia on cellular pathways is complex with true hypoxia because HIF-1α has a short lifetime and rapidly reverts back to a normoxic state. To overcome this difficulty, mimetic agents of the hypoxia pathway have been developed, including the gold standard CoCl₂. In this study, we proposed to compare CoCl₂ and hydralazine in order to determine a suitable mimetic agent of hypoxia for the study on the BBB. Methods: We studied the cytotoxicity and the impact of these molecules on the integrity of an in vitro BBB model by comparing them to hypoxia controls. Results: We showed that the impact of hypoxic stress in our in vitro BBB model is rather similar between hydralazine and CoCl₂. Chemical hypoxic stress led to an increase of BBB permeability either with CoCl₂ or hydralazine. Tight junction protein expressions showed that this chemical hypoxic stress decreased ZO-1 but not occluding expressions, and cells had set up a defence mechanism by increasing expression and activity of their efflux transporters. Conclusion: Our results demonstrated that hydralazine is a better mimetic agent and more suitable than CoCl₂ because it had the same effect but without the cytotoxic effect on in vitro BBB cells.

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“…Although most research has focused on the development and characterisation of BBEC cultures, porcine brain endothelial cells (PBECs) are another convenient source with a sufficient number of BCECs to allow extensive testing. The use of primary cultures of PBECs as an in vitro permeability model of the BBB has been pioneered by Franke et al (1999), and more recent studies have also used co-cultured PBECs with astrocytes in order to improve the restrictiveness of the culture system (Kido et al, 2002;Zhang et al, 2006). Rat brain endothelial cells (RBECs) have also been described and may be relevant to facilitate in vitro/in vivo correlations if the rat is primarily used to generate in vivo data (Demeuse et al, 2002;Perriere et al, 2007).…”

Section: Primary or Low Passage Brain Capillary Endothelial Cell Cultmentioning

confidence: 99%

In vitro blood brain barrier models as a screening tool for colloidal drug delivery systems and other nanosystems

Garcel

Martel

Carrupt

et al. 2010

IJBNN

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The brain is one of the least accessible organs of the body due to the presence of the blood-brain barrier (BBB), thus making drug delivery to the brain quite a challenge. Various strategies have been explored to circumvent this physiological barrier, including the use of colloidal carriers. These carriers hold great promise as they may increase the delivery of drugs into the brain by protecting them from degradation and prolonging their circulation in the blood, as well as promoting their transport through the BBB. Moreover, functionalisation of these carriers with various ligands allows specific targeting of the central nervous system compartment. Additionally, various in vitro BBB models have been developed and are increasingly useful for screening of drug delivery systems, especially cell-based models that provide mechanistic information. In fact, this paper specifically reviews selected in vitro BBB models as a screening tool for drug delivery colloidal systems.Keywords: drug delivery; colloidal systems; nanocarriers; nanoparticles; liposomes; micelles; blood-brain barrier; BBB; in vitro model.Reference to this paper should be made as follows: Garcel, A., Martel, S., Carrupt, P.A., Doelker, E. and Delie, F. (2010) 'In vitro blood brain barrier models as a screening tool for colloidal drug delivery systems and other nanosystems', Int.

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Evaluation of Blood-brain Barrier Transporters by Co-culture of Brain Capillary Endothelial Cells with Astrocytes

Cited by 32 publications

References 25 publications

Mrp4 Confers Resistance to Topotecan and Protects the Brain from Chemotherapy

Mrp4 Confers Resistance to Topotecan and Protects the Brain from Chemotherapy

Hydralazine is a Suitable Mimetic Agent of Hypoxia to Study the Impact of Hypoxic Stress on In Vitro Blood-Brain Barrier Model

In vitro blood brain barrier models as a screening tool for colloidal drug delivery systems and other nanosystems

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