Tight junctions at the blood brain barrier: physiological architecture and disease-associated dysregulation

Fluids and Barriers of the CNS Tight junctions at the blood brain barrier: physiological architecture and disease-associated dysregulation Anny-Claude Luissint Cdric Artus Fabienne Glacial Kayathiri Ganeshamoorthy Pierre-Olivier Couraud The Blood-brain barrier (BBB), present at the level of the endothelium of cerebral blood vessels, selectively restricts the blood-to-brain paracellular diffusion of compounds; it is mandatory for cerebral homeostasis and proper neuronal function. The barrier properties of these specialized endothelial cells notably depend on tight junctions (TJs) between adjacent cells: TJs are dynamic structures consisting of a number of transmembrane and membrane-associated cytoplasmic proteins, which are assembled in a multimolecular complex and acting as a platform for intracellular signaling. Although the structural composition of these complexes has been well described in the recent years, our knowledge about their functional regulation still remains fragmentary. Importantly, pericytes, embedded in the vascular basement membrane, and perivascular microglial cells, astrocytes and neurons contribute to the regulation of endothelial TJs and BBB function, altogether constituting the so-called neurovascular unit. The present review summarizes our current understanding of the structure and functional regulation of endothelial TJs at the BBB. Accumulating evidence points to a correlation between BBB dysfunction, alteration of TJ complexes and progression of a variety of CNS diseases, such as stroke, multiple sclerosis and brain tumors, as well as neurodegenerative diseases like Parkinson's and Alzheimer's diseases. Understanding how TJ integrity is controlled may thus help improve drug delivery across the BBB and the design of therapeutic strategies for neurological disorders. Blood-brain barrier; Tight junction; Neurovascular unit; Kinases; Signaling pathways - Review Background The BBB maintains the homeostasis of the central nervous system (CNS) by (i) strictly limiting the passive diffusion of polar substances from the blood to the brain, (ii) mediating the transport of nutrients to the brain parenchyma as well as the efflux from the brain of toxic metabolites and xenobiotics, (iii) regulating the migration of circulating immune cells [1-3]. Formed by specialized vascular endothelial cells, the BBB is tightly controlled by pericytes, embedded in the vascular basement membrane, perivascular microglial cells, astrocytes and neurons which * Correspondence: 1INSERM U1016, Institut Cochin, Paris, France 2CNRS, UMR 8104, Paris, France Full list of author information is available at the end of the article altogether constitute the neurovascular unit (NVU), a concept highlighting the functional cell-cell interactions supporting BBB function. BBB endothelial cells display a unique phenotype characterized by the presence of TJs and the expression of specific polarized transport systems. TJs constitute the most apical intercellular junctional complex in polarized epithelium and endothelium, with three key biological functions: a barrier to paracellular diffusion of bloodborne polar substances [4], a fence preventing the lateral diffusion of lipids and integral membrane proteins, thus maintaining cell polarization [5-7] and an intracellular signaling platform which will be described below. Brain endothelial TJ strands, like epithelial TJs, are composed of integral membrane proteins (occludin, claudins and junctional adhesion molecules (JAMs)) involved in intercellular contacts and interactions with cytoplasmic scaffolding proteins such as zonula occludens (ZO) proteins, actin cytoskeleton and associated proteins, such as protein kinases, small GTPases [8] and heterotrimeric G-proteins [9]. Excellent reviews have recently been published on the architecture of TJ complexes in epithelial and brain endothelial cells [10,11]. Here we will briefly recall the main features of the structural organization of TJs at the BBB and will focus on transcriptional regulation, posttranslational modifications and subcellular localization of TJ proteins and their consequences for BBB integrity with exposure to various environmental stimuli and during CNS disorders. Components of TJs in brain endothelial cells As in polarized epithelial cells where TJs have been mostly studied, the TJ backbone in brain endothelial cells consists of transmembrane proteins (occludin, claudins and JAMs) which recruit a number of membrane-associated cytoplasmic proteins. Transmembrane proteins as the BBB TJ backbone Occludin (60kDa), a tetraspan integral membrane protein, was the first TJ-specific protein identified [12,13] in epithelial cells and shown to be functionally important for barrier function [14]. It is a member of the family of TJ-associated marvel proteins (TAMP) with tricellulin (marvelD2) [15] and marvelD3 [16,17]. Both the MARVEL transmembrane domain of occludin, encompassing the four transmembrane helices, and its coiled coil cytosolic C-terminus were recently described to mediate its lateral (i.e. cis-) oligomerization in epithelial MDCK cells [18-20]. More precisely, cystein residues in these domains are directly involved in oligomerization through disulfide bridge formation. This process being redox-sensitive, oligomerization of occludin likely contributes to the redox-dependency of the TJ assembly [20,21]: whereas normoxia conditions support occludin oligomerization and contribute to TJ assembly, oxidative stress associated with hypoxia-reoxygenation [22] or inflammation [23,24] results in TJ disruption. This novel concept that occludin plays a key role in the redox regulation of TJs has been very recently reviewed [25]. In addition, the second extracellular domain of occludin is required for its stable assembly in TJs [26]. Indeed, synthetic peptides corresponding to this domain were shown to perturb TJ permeability barrier in epithelial cells [27-29]. The important contribution of occludin to TJ function is illustrated by the observations that ectopic expression of chicken occludin induced the formation of TJ-like structures in Sf9 insect cells [30], while increasing electrical resistance in MDCK cells [31]. Conversely, occludin degradation induced by viruses or bacteria (like HIV-1 Tat protein or Neisseria meningitidis), is associated with increased permeability in primary or immortalized human brain microvascular endothelial cells, respectively [32,33]. However, well-developed TJ strands were reported in cells lacking occludin (human or guinea pig testis) [34] and between adjacent occludin-deficient epithelial cells [34,35]; together with the report that occludin deficientmice are viable, exhibiting normal TJs morphology as well as intestinal epithelium barrier function, these observations indicate that occludin is dispensable for TJ formation [36,37]. Claudins constitute a large family of 20-27kDa membrane proteins (with four transmembrane domains) expressed in TJ (...truncated)