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OLECULAR MECHANISMS OF BRAIN TUMOR EDEMA

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. C. PAPADOPOULOS,a,b,c* S. SAADOUN,a,b,c

. K. BINDER,c,d G. T. MANLEY,d S. KRISHNAb AND

. S. VERKMANc

Department of Neurosurgery, St. George’s Hospital Medical School,ondon, UK

Department of Cellular and Molecular Sciences, St. George’s Hospi-al Medical School, London SW17 0NE, UK

Department of Medicine and Physiology, University of California atan Francisco, San Francisco CA 94143, USA

Department of Neurological Surgery, University of California atan Francisco, San Francisco CA 94143, USA

bstract—Despite their diverse histological types, mostrain tumours cause brain oedema, which is a significantause of patient morbidity and mortality. Brain tumour oe-ema occurs when plasma-like fluid enters the brain extra-ellular space through impaired capillary endothelial tightunctions in tumours. Under-expression of the tight junctionroteins occludin, claudin-1 and claudin-5 are key molecularbnormalities responsible for the increased permeability ofumour endothelial tight junctions. Recent evidence sug-ests that the membrane water channel protein aquaporin-4AQP4) also plays a role in brain tumour oedema. AQP4-eficient mice show remarkably altered brain water balancefter various insults, including brain tumour implantation.QP4 expression is strongly upregulated around malignantuman brain tumours in association with reduced extracel-

ular volume, which may restrict the flow of extracellular fluidrom the tumour bed into the brain parenchyma. Eliminationf excess fluid leaking into brain parenchyma requires pas-age across three AQP4-rich barriers: a) the glia limitansxterna, b) the glia limitans interna/ependyma, and c) thelood–brain barrier. Modulation of the expression and/orunction of endothelial tight junction proteins and aquaporinsay provide novel therapeutic options for reducing brain

umour oedema. © 2004 IBRO. Published by Elsevier Ltd. Allights reserved.

ey words: aquaporin, blood–brain barrier, tight junction,asogenic edema, water channel.

rain oedema is defined as a net increase in waterontent of the brain parenchyma. Redistribution of wateretween the intracellular and extracellular compart-ents, or an increase in cerebral blood or cerebrospinal

uid volume does not therefore constitute brain oedema.

Correspondence to: M. C. Papadopoulos, Cellular and Molecularciences, St. George’s Hospital Medical School, London SW17 0NE,K. Tel: �1-415-476-8530 (USA) or �44-20-8725-0669 (UK); fax:1-415-665-3847 (USA) or �44-20-8725-3487 (UK).-mail address: mpapadop@sghms.ac.uk (M. C. Papadopoulos).bbreviations: AQP, aquaporin; BBB, blood–brain barrier; CSF, cere-rospinal fluid; ECS, extracellular space; GBM, glioblastomaultiforme; ICP, intracranial pressure; JAM, junctional adhesion

polecule; SF/HGF, scatter factor/hepatocyte growth factor; VEGF,

ascular endothelial growth factor; ZO, zonula occludens.

306-4522/04$30.00�0.00 © 2004 IBRO. Published by Elsevier Ltd. All rights reseroi:10.1016/j.neuroscience.2004.05.044

1011

ll aggressive brain tumours, such as malignant gliomasnd metastatic tumours, and many benign tumours,uch as meningiomas, produce brain oedema regard-

ess of their cell type of origin (Thapar and Rutka, 1995).ecause the skull is non-compliant, brain oedemaauses increased intracranial pressure (ICP), potentially

eading to brain ischaemia, herniation and death. Theethal effect of oedema is illustrated by the 10-fold re-uction in brain tumour operative mortality in the 1960s,hich resulted from treating brain oedema with cortico-teroids (Jelsma and Bucy, 1967).

Current understanding of brain oedema is based onlatzo’s (1994) classification into cytotoxic and vasogenic

ypes. Cytotoxic oedema refers to cell swelling, as seen inarly hypoxia, which primarily affects the astrocytes. Va-ogenic oedema, of which brain tumour oedema is therchetypal example, is produced by fluid flow into thextracellular space (ECS) of the brain parenchyma throughn incompetent blood–brain barrier (BBB). The amount ofrain swelling is a dynamic balance between oedema fluidormation and absorption. Research into brain tumour oe-ema has concentrated on oedema fluid formation, and soelatively little is known about the mechanisms of oedemauid elimination.

In humans, peripheral tumour metastases to brain (Itot al., 1990) and primary brain tumours such as glioblas-oma multiforme (GBM) (Groger et al., 1994) produce up to0 ml of oedema fluid each day. The excess fluid is ac-ommodated within the skull, which contains the brain,erebrospinal fluid (CSF) and blood. In normal humandults the intracranial volume contains intracellular (1100–300 ml) and interstitial (100–150 ml) spaces, and theSF (75–100 ml) and blood (75–100 ml) spaces. Ex-hange of fluid amongst these compartments occurs inesponse to osmotic and hydrostatic forces at the inter-aces between intracellular and extracellular compart-ents (cell membranes), blood and brain (BBB), CSF andrain (ventricular ependyma, glia limitans), and blood andSF (choroid plexus and arachnoid granulations). In addi-

ion, up to 30 ml of water in brain is produced daily fromlucose metabolism. Because the skull is rigid, an increase

n brain parenchymal volume results in displacement ofuid from the low-pressure CSF (approximately 10 mm Hg

n man) and venous (approximately 10 mm Hg) compart-ents, and later the high-pressure arterial (approximately00 mm Hg mean) compartment (Monro-Kellie doctrine).he ability of the brain to compensate for a rise in ICP as

otal brain water increases is in large part due to theapacity of the CSF and venous compartments to be com-

ressed.

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M. C. Papadopoulos et al. / Neuroscience 129 (2004) 1011–10201012

rain tumour oedema: a defect in the blood–tumourarrier

he existence of the BBB was first demonstrated in 1885y the German pathologist Ehrlich (1885) who reportedhat i.v. injection of albumin-bound dyes into rats stained allody tissues except for the brain. Subsequent electronicroscopy studies showed that i.v. administered electronense particles, such as horseradish peroxidase, are pre-ented from entering the brain parenchyma by capillaryndothelial cells (Reese and Karnovsky, 1967). Cerebralapillary endothelial cells differ from those of peripheralapillaries in that they are not fenestrated and contain fewndocytotic vesicles, thus limiting transcellular flux (Saget al., 1998). Individual brain microvessel endothelial cellsake contact by tight junctions, further limiting intercellular

ux (Reese and Karnovsky, 1967; Rubin and Staddon,999). The permeability properties of the BBB reflect

argely the tightness of these junctions (Bar-Sella et al.,979; Rubin and Staddon, 1999). Astrocytes, whose pro-esses form end-feet that surround brain microvesselsJanzer and Raff, 1987; Hayashi et al., 1997), and peri-ytes, which contact microvessel endothelial cells, proba-ly secrete factors that confer BBB properties to adjacentndothelial cells. Transplanted astrocytes in vivo (Janzernd Raff, 1987; Hayashi et al., 1997) and astrocytes initro (Janzer and Raff, 1987; Hayashi et al., 1997) induceBB properties in adjacent non-neural endothelial cells

rom different species. Interestingly, some of the BBB-nducing properties of astrocytes are mimicked by cortico-teroids (Underwood et al., 1999; Antonetti et al., 2002;omero et al., 2003), which are used clinically to reducerain tumour oedema. The absence of brain pericytes inlatelet-derived growth factor receptor � deficient mice isssociated with endothelial cell proliferation and increasedicrovascular permeability (Hellstrom et al., 1999, 2001).

Increased vascular permeability in human brain tu-ours has been demonstrated by computed tomographiceasurements of blood–tumour barrier permeability

Groothuis et al., 1991) and by immunohistochemical de-ection of plasma proteins in tumour parenchyma (Seitznd Wechsler, 1987). The leakiness of the blood–tumourarrier occurs primarily because of defects in inter-ndothelial tight junctions, although there may be otherontributing abnormalities in endothelial pinocytosis andndothelial fenestrations (Long, 1979; Nir et al., 1989;hibata, 1989). Abnormalities in the appearance of tight

unctions in human gliomas correlate with increasing ma-ignancy (Shibata, 1989). Electron microscopy has alsohown that i.v.-injected horseradish peroxidase in rodentsith GBM is excluded locally from the brain parenchymay morphologically normal tight junctions, but not by de-ective tight junctions (Nir et al., 1989). In human meta-tatic tumours the blood vessels fail to form tight junctionsLong, 1979) and exhibit the characteristics of their parentissue such as expression of peripheral endothelial mark-rs like aquaporin (AQP) 1 (Saadoun et al., 2002a). This is

nteresting because metastatic tumours recruit endothelial

ells from the surrounding brain to form blood vessels. o

erebral endothelial cells that vascularise brain tumoursrobably dedifferentiate in response to signals producedy the surrounding tumour cells. Cultured endothelial cellsrom brain (Dropulic and Masters, 1987; Stolz and Jacob-on, 1991) and other organs (Stolz and Jacobson, 1991)an be induced to lose features characteristic of theirissue of origin and adopt an immature phenotype byhanging culture media and exposing to growth factors.

There is no general agreement about whether the mi-rovessels of peritumoral brain are leaky. Long (1979) didot find morphological evidence of increased permeability

n microvessels around metastatic adenocarcinomas,hereas Stewart et al. (1985) reported structural defects inndothelial tight junctions around human GBM. The latterndings may be related to the ill-defined margins in GBMnd cell infiltration in surrounding brain.

The importance of the vascular bed in the initiation androgression of vasogenic oedema has been shown in an-

mal models. Excision of the lesioned cortex immediatelyfter focal freeze injury completely abolishes the expectedasogenic oedema and removal of the lesion a few hoursfter freeze injury prevents advancement of the oedemaront (Aarabi and Long, 1979). There is good evidence thatasogenic oedema fluid arises from plasma. In cat modelsf tumour implantation, brain abscess and focal freeze

njury, the sodium content of the oedema fluid was theame as plasma (129–135 mM; Bothe et al., 1984) and

.v.-injected 99mTc-labelled albumin was found in the oe-ema fluid (Gazendam et al., 1979).

Recently the molecular constituents of brain endothe-ial tight junctions have been elucidated (Huber et al.,001; Fig. 1). Tight junctions consist of transmembranend intracellular protein components (Rubin and Staddon,999; Wolburg and Lippoldt, 2002; Vorbrodt and Dobro-owska, 2003). The transmembrane proteins, which in-lude the claudin family, occludin and junctional adhesionolecule (JAM), bind to their counterparts on adjacent

ells, in effect ‘gluing’ two cells together. Although occludins expressed ubiquitously in tight junctions, members of thelaudin family are expressed in different tissues. Brainndothelial cells express occludin, JAM, claudin-1 andlaudin-5 (Rubin and Staddon, 1999; Wolburg and Lip-oldt, 2002; Vorbrodt and Dobrogowska, 2003). At theytoplasmic surface of cells, the claudins and occludin bindonula occludens 1 (ZO1), ZO2, ZO3 and other less well-haracterised proteins, each of which anchor to the actinytoskeleton. ZO1, ZO2 and ZO3 are members of theembrane-activated guanylate cyclase protein family

Stevenson et al., 1986; Haskins et al., 1998), suggestingheir involvement in intracellular signalling. This complexrganisation confers plasticity to the tight junction, allowing

t to function as a gate that can be transiently opened byhe cell in response to second messengers to permit pas-age of leucocytes or solutes (Vestweber, 2002). Phos-horylation of occludin and claudins may regulate tight

unction permeability (Rubin and Staddon, 1999). Proteininase C (Andreeva et al., 2001) and casein kinase 2Smales et al., 2003) have been shown to phosphorylate

ccludin on tyrosine residues.

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M. C. Papadopoulos et al. / Neuroscience 129 (2004) 1011–1020 1013

Some defects in brain tumour endothelial vessel tightunctions have been characterised. Occludin, claudin-1nd claudin-5 are downregulated in human gliomas andre absent in metastatic tumour vessel tight junctionsLiebner et al., 2000; Papadopoulos et al., 2001b; Raschert al., 2002). The occludin protein content of human astro-ytomas is inversely related to the Daumas-Duport histo-ogical grade (Papadopoulos et al., 2001b). It has beenuggested that in human high-grade gliomas, occludin isownregulated and phosphorylated (Papadopoulos et al.,001a,b). Phosphorylation of occludin inhibits its interac-ions with ZO1, ZO2 and ZO3 (Kale et al., 2003) andncreases tight junction permeability (Rubin and Staddon,999).

Two principal reasons why tumours have defectivendothelial tight junctions include reduced numbers of nor-al astrocytes in tumour tissue, and excessive secretion ofngiogenic factors. Because aggressive tumours are defi-ient in normal astrocytes, they lack the astrocyte derivedactors required for the formation of a normal BBB (Janzernd Raff, 1987). Brain tumour cells secrete a mixture ofngiogenic factors, such as vascular endothelial growthactor (VEGF, originally called vascular permeability factor;ates et al., 1999) and scatter factor/hepatocyte growth

actor (SF/HGF; Lamszus et al., 1999; Arrieta et al., 2002).

ig. 1. Molecular constituents of endothelial tight junctions. Transmendothelial cells, thus ‘gluing’ the cells together. Intracellularly, occlud

o the actin cytoskeleton.

igh VEGF immunoreactivity has been reported in human t

naplastic astrocytoma and GBM (Lafuente et al., 1999;udwig et al., 2000b), oedematous meningioma (Goldmant al., 1997) and metastatic tumours to the brain (Ludwig etl., 2000a). In cultured brain or retinal endothelial cells,EGF (Antonetti et al., 1999; Wang et al., 2001; Behzadiant al., 2003) and SF/HGF (Jiang et al., 1999) cause down-egulation and phosphorylation of the tight junction com-onent proteins occludin and ZO1, with a corresponding

ncrease in monolayer permeability. In contrast, corticoste-oids cause de-phosphorylation of occludin and ZO1, andecreased endothelial monolayer permeability (Under-ood et al., 1999; Antonetti et al., 2002; Romero et al.,003). Many malignant tumours outgrow their blood supplynd become hypoxic, which is probably a potent stimulusor secretion of angiogenic factors by brain tumour cells.he VEGF promoter has hypoxia responsive elementsShibata et al., 2000), and hypoxia has been shown totimulate human GBM cells to secrete VEGF (Plate et al.,994).

About 60% of meningiomas cause brain oedema, evenhough they do not originate from brain cells, but arise fromhe coverings of the brain (extra-axial tumours). Meningi-mas grow in extra-cerebral spaces, have extra-axiallood supply, and are separated from the brain by the piaater. Electron microscopy of human peri-meningioma

proteins (occludin and claudins) bind their counterparts on adjacentudins are attached to ZO1, ZO2 and ZO3, which are in turn attached

mbrane

issue showed increased ECS volume, suggesting that the

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ssociated oedema is of the vasogenic type (Gilbert et al.,983). The presence of brain oedema may be related tohe secretion of VEGF by meningioma cells (Yoshioka etl., 1999; Paek et al., 2002; Pistolesi et al., 2002). It ishought that VEGF enters the peritumoral space andauses the formation of a cerebral/pial blood supply to theumour (Yoshioka et al., 1999; Paek et al., 2002; Pistolesit al., 2002). Human peri-meningioma endothelial cellshow morphological changes, such as nuclear enlarge-ent and many pinocytotic vesiles, consistent with de-ifferentiation due to exposure to VEGF (Vaz et al., 1998).he cerebral/pial-derived microvessels lack tight junctionsllowing brain oedema to develop in the same way as itoes in aggressive gliomas and metastases.

low of oedema fluid in brain ECS

he hydrostatic pressure gradient (systemic arterial pres-ure minus ICP) is the primary force driving vasogenicedema fluid entry into the ECS. After focal cortical freeze

njury in cats, elevation of the mean arterial pressure dra-atically accelerated the spread of oedema, and oedema

pread was slow with reduced blood pressure (Klatzo,994). Having crossed the defective blood–tumour barrier,edema fluid enters the ECS of the tumour bed. Comparedith normal brain tissue, aggressive human brain tumoursuch as GBM and medulloblastomas have an apparentlyxpanded ECS volume (Bruehlmeier et al., 2003; Vargovat al., 2003), probably resulting from necrosis, abnormal

ig. 2. AQP4 expression (brown) in human brain and brain tumoursrocesses, and (B) the glia limitans. Note massive upregulation of AQD) the gliotic region around metastatic carcinoma. Haematoxylin cou

umour cell shape and defective ECS matrix. Increased t

CS volume may provide a low resistance pathway facil-tating the flow of oedema fluid throughout the tumour bed.

Prior to entering the ECS of the brain parenchyma, theedema fluid must cross a rim of reactive glia surroundinghe tumour (Zhang and Olsson, 1997). The role of reactiveliosis in brain tumour biology is poorly understood, butay limit the excess fluid entering the brain parenchyma.he gliotic tissue presents a significant barrier to the flow ofxtracellular fluid (Roitbak and Sykova, 1999), probablyue to the high density of hypertrophied astrocytic pro-esses and enhanced formation of extracellular matrix.fter forebrain stab injury in mice, gliosis has been shown

o reduce BBB permeability, leucocyte infiltration, and neu-onal death (Bush et al., 1999).

Once inside the ECS of the brain parenchyma, furtherovement of oedema fluid is resisted by the cells and theirrocesses. Dilation of the ECS by the hydrostatic pressuref the oedema fluid creates a pathway for its movement.asogenic oedema spreads by bulk flow (Reulen et al.,977) preferentially through the white matter, which pre-ents a more orderly arrangement of extracellular chan-els and thus offers less resistance to oedema fluid flowhan the densely tangled cellular structures of the greyatter (Thapar and Rutka, 1995).

limination of oedema fluid: role of AQP waterhannels?

everal groups have proposed the involvement of AQPs in

al brain, there is immunostaining of (A) pericapillary astrocyte footnoreactivity and loss of polarized expression in (C) glioblastoma, andScale bars�10 �m (A), 35 �m (B), 30 �m (C), 10 �m (D).

. In norm

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M. C. Papadopoulos et al. / Neuroscience 129 (2004) 1011–1020 1015

apadopoulos et al., 2002; Agre and Kozono, 2003; Amiry-oghaddam and Ottersen, 2003). The AQPs are a familyf water channel proteins, which comprises at least 12embers in mammals (Verkman, 2002; Agre and Kozono,003). The AQPs provide a major pathway for osmotically-riven water movement across plasma membranes inome cell types. AQP4 is the predominant water channel inormal brain; it is strongly expressed in astrocyte plasmaembranes, and has been shown to increase the waterermeability of cell membranes isolated from mouse brainMa et al., 1997) and cultured cortical astrocytes (Solenovt al., 2004). As shown in Fig. 2, the sites of AQP4 expres-ion include the pericapillary astrocyte foot processes, thexternal glial limiting membrane, the ependyma, and theubependymal glia limitans interna (Nielsen et al., 1997;ash et al., 1998). The localisation of AQP4 at the CSF–rain and blood–brain interfaces suggests its involvement

n fluid transport into and out of the brain. Although AQP1nd AQP9 have also been found in the brain, their expres-ion is limited. In normal brain, AQP1 is found on theentricular-facing surface of the choroid plexus, where itacilitates the secretion of CSF (Oshio et al., 2003b). AQP9as been detected in the glia limitans and tanycytes, but itsrecise expression pattern remains unclear (Elkjaer et al.,000; Badaut et al., 2001). The possible role of AQP9 in

he brain is discussed further in the review by Badaut and

ig. 3. Brain swelling in mice with implanted melanoma brain tumours.�/�, E) mice. However, the AQP4-null mice have (B) worse (lower)P

egli (2004) in this issue of Neuroscience. t

Immunohistochemical studies of human brain tumoursSaadoun et al., 2002b, 2003; Badaut et al., 2003) reporttrong upregulation of AQP4 in astrocytes in the vicinity ofedematous brain tumours. Fig. 2A shows AQP4 expres-ion around a normal human brain microvessel and Fig. 2Bhows expression in the glia limitans. There is marked

ncrease in AQP4 expression in astrocytes in GBM (Fig.C) and reactive astrocytes around carcinoma metastasiso brain (Fig. 2D). Unlike normal brain, brain tumour-ssociated AQP4 expression is not polarized to astrocyteoot processes, but is seen throughout the entire astrocyteell membrane. Correlation between increased AQP4 ex-ression and brain tumour oedema provides indirect evi-ence for the importance of AQP4 in the pathophysiologyf brain tumour oedema. Due to the lack of AQP4 inhibi-ors, it is not yet possible to verify a role of AQP4 directly inasogenic brain oedema. However, AQP4-null mice haveeen valuable in defining the role of AQP4 in fluid transportnd balance in brain [see review by Manley et al. (2004) onQP4 knockout studies].

Despite the strong expression of AQP4 in the brain,QP4-null mice are surprisingly normal in terms of grossehaviour, brain morphology, blood vessel anatomy, ICP,ulk intracranial compliance, and expression of otherQPs (Ma et al., 1997; Manley et al., 2000; Papadopoulost al., 2004). Experiments using AQP4-null mice suggest

umours grow at comparable rates in wild-type (�/�, F) and AQP4-nullical score, and (C) higher ICP, measured 1 week post-implantation.

(A) The t

he involvement of AQP4 in both cytotoxic and vasogenic

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rain oedema. AQP4 deletion protected mice from brainwelling in two models of primarily cytotoxic oedema: wa-er intoxication and permanent focal cerebral ischaemiaManley et al., 2000). Similar findings were reported inice lacking �-syntrophin (Amiry-Moghaddam et al., 2003)r dystrophin (Vajda et al., 2002), which secondarily man-

fest altered distribution of brain AQP4 expression. In con-rast, AQP4 deletion caused increased brain swelling inhree models of vasogenic brain oedema: intraparenchy-al fluid infusion, focal freeze injury and brain tumour

Papadopoulos et al., 2004). Intracerebral fluid infusionimics the resolution phase of brain swelling by producingcontrolled amount of brain oedema without the poten-

ially confounding variations in BBB permeability. Cold in-ury is a highly reproducible model of ‘vasogenic’ brainedema resulting from increased BBB permeability anduid extravasation. In the tumour model, melanoma cellsere stereotactically implanted into the brain. As illustrated

n Fig. 3A, 7 days post-implantation, the tumours werequal in size in AQP4-null and wild-type mice (Papado-oulos et al., 2004). However, the AQP4-null mice devel-ped worse neurological score (Fig. 3B) and higher ICPFig. 3C) than wild-type controls. These experiments pro-ided evidence that AQP4 deletion limits brain swelling inytotoxic oedema, but aggravates brain swelling in vaso-enic oedema.

These opposing actions of AQP4 in cytotoxic and va-ogenic oedema are probably related to the bidirectionalater transport through the AQP4 channel (Meinild et al.,998; Amiry-Moghaddam et al., 2003). We hypothesisehat influx of oedema fluid into the brain parenchyma in

ig. 4. Proposed mechanism of AQP4 involvement in oedema fluidarenchyma is AQP4 dependent, because oedema fluid flows fromrocesses, and accumulates primarily in astrocytes. (B) In vasogenicermitting the entry of plasma fluid directly into the brain ECS circum

ytotoxic oedema involves water movement through the v

ntact BBB and astrocyte foot processes. Fluid entry intohe brain in vasogenic oedema involves movement acrosshe leaky tumour microvascular endothelium, which is noturrounded by astrocyte foot processes. Therefore, theormation of cytotoxic oedema is AQP4-dependent, buthe formation of tumour-associated oedema is AQP4-ndependent (Fig. 4). However, elimination of excess brainater in both cytotoxic and vasogenic brain oedema is

ikely to be AQP4-dependent because excess fluid flowshrough AQP4-containing barriers. Fluid flows through thelia limitans into the subarachnoid space, through thependyma into the ventricles, and through the BBB andstrocyte foot processes into the blood (Reulen et al.,977; Marmarou et al., 1994; Fig. 5).

In brain tumours, AQP4 may also be involved in limitinghe entry of oedema fluid from the tumour bed into therain parenchyma. Indirect evidence suggests a correla-ion between the size of the ECS and the level of AQP4xpression, with AQP4-expressing gliotic tissue (Saadount al., 2002b) presenting a significant barrier to extracellu-

ar fluid flow (Roitbak and Sykova, 1999). In the first feweeks after birth, rat brain ECS volume decreases (Leh-enkuhler et al., 1993) in parallel to increased AQP4xpression (Wen et al., 1999). There is strong AQP4 ex-ression in the glia limitans (Nielsen et al., 1997; Saadount al., 2003), which consists of tightly packed astrocyte footrocesses (Brightman, 2002). AQP4-null mice have higherCS volume compared with wild-type controls (D. K.inder, unpublished observations). The reported in-reased AQP4 expression in peritumoral reactive astro-ytes (Saadoun et al., 2002b) may therefore reduce ECS

n. (A) In cytotoxic oedema, the entry of excess fluid into the brainular compartment, through intact BBB and AQP4-rich astrocyte footwater accumulation is AQP4-independent because the BBB is leaky,strocyte foot processes.

formatiothe vasc

olume in the peritumoral gliotic rim. This would slow the

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ow of oedema fluid from the tumour bed into the brainCS.

It has been reported recently that AQP1 is expressedn tumour cells and peritumoral astrocytes in human high-rade gliomas (Saadoun et al., 2002a; Oshio et al.,003a). This observation is intriguing because AQP1 is notormally found in astrocytes; its expression is restricted to

he choroid plexus, where it provides the major waterathway for secretion of CSF (Oshio et al., 2003b). It is notnown whether tumour-associated AQP1 is functional orhether it plays a role in the pathophysiology of brain

umour oedema. Experiments comparing brain tumour oe-ema in AQP1 knockout mice versus wild-type controls are

ikely to clarify the role of AQP1 in brain tumour oedema.

CONCLUSIONS

rain tumour oedema results from imbalance betweenater moving into and out of the brain. Two recent discov-ries have advanced our understanding of both processes.irst, molecular abnormalities of tumour endothelial tight

unctions increase blood–tumour barrier permeability, thusccelerating the rate of water entry into the brain. Second,he elimination of excess water from the brain is controlledy the water channel AQP4. We therefore suggest thatndothelial tight junction proteins and AQP4 representargets for the design of novel drugs to treat brain tumouredema. Such compounds might slow oedema fluid for-ation (by upregulating occludin, claudin-1 and claudin-5

n endothelial cells) or increase oedema fluid elimination

ig. 5. Proposed role of AQP4 in elimination of cytotoxic and vasogenuid is eliminated through (A) the glia limitans externa into the subaracnd ependyma into the ventricles.

by upregulating AQP4 in astrocytes).

cknowledgements—Supported by a Wellcome Trust Clinician-cientist Fellowship (to M.C.P.).

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(Accepted 25 May 2004)