Blood-brain barrier basics: A selective passageway for bacterial pathogens and antimicrobial drugs

Meningitis is a serious complication of Streptococcus suisbacteremia in young piglets. This microorganism is also an emerging human pathogen that has been recently implicated in a case of meningitis in a swine handler (Wertheim et al., 2009; Fowler et al., 2013). In order to access and infect the brain meninges, blood-borne S. suis must penetrate the blood-brain barrier of affected animals or humans. The purpose of this article is to briefly review some key aspects of the locations and protective functions of this important physical barrier in swine and other animals and summarize our current understanding of how S. suis breaches this barrier. 

The blood-brain barrier plays several key roles in central nervous system (CNS) homeostasis in that it hinders the entry of microorganisms and potentially neuroactive or toxic substances into the brain, maintains concentrations of proteins and ions at levels optimal for neural activity, and orchestrates immune responses that incur a minimum of inflammation and cellular damage (that would disrupt barrier function, see below). There are in reality, three major barriers within the CNS, which include (1) the endothelial cells lining continuous capillaries supplying the brain parenchyma, which encompasses the largest surface area for barrier function; (2) a less extensive blood-cerebrospinal fluid (CSF) barrier consisting of CSF-secreting choroid plexus epithelial cells lying in close proximity to fenestrated capillaries; and (3) an arachnoid epithelium within the meninges (Saunders et al., 2012; Abbott, 2013). The microvascular endothelial cells of brain capillaries interact with closely apposed pericytes, and adjacent neurons, astrocytes and microglia (the “macrophages of the brain”) to form a so-called neurovascular unit (Iadecola, 2004). These cells act in a concerted manner to maintain barrier function. Compared with these three regions, the circumventricular organs possess a “leakier” barrier composed of ventricular ependymal cells. This more permeant cellular structure permits brain chemosensors to monitor and respond appropriately to changes in serum osmolality or the presence of blood-borne toxicants. 

Brain microvascular endothelial cells (BMECs) and epithelial cells of the choroid plexus and meninges are held together by a complex array of specialized membrane proteins to form tight junctions. Claudin-5, occludin, junctional adherens molecules (JAMs) and zona occludens adapter proteins are proteins in tight junctional complexes that are commonly expressed in BMECs and choroid plexus epithelial cells (CPECs) from pigs (Malina et al., 2009; Schroten et al., 2012). These tight junctions physically exclude proteins and other macromolecules as well as microbes present in the brain blood supply. Even blood-borne molecules that are essential for brain function are generally excluded by these intercellular junctions and normally cannot cross the barrier through a paracellular pathway. Instead, these substances move across the blood-brain interface by means of selective carrier proteins (e.g. ion and water channels, glucose and amino acid transporters, transferrin receptors, efflux pumps). Many inflammatory, immune and neuroactive substances can regulate blood-brain barrier permeability (Liu et al., 2012). For example tumor necrosis factor-α, a pro-inflammatory cytokine, has been reported to increase paracellular permeability to both small and large molecules in neonatal pigs (Megyeri et al., 1992; Abraham et al., 1996). On the other hand, transforming growth factor-β, which is secreted by astrocytes and pericytes present in the neurovascular unit, decreases blood-brain barrier permeability by increasing tight junction protein gene expression in BMECs (Takeshita and Ransohoff, 2012). 

Streptococcus suis appears to migrate across the bloodbrain and blood-CSF barriers of piglets after adhering to and invading BMECs and CPECs respectively. This process appears to involve several Streptococcal proteins, including adhesins, invasins and cell wall components as well as bacterial interactions with serum components such as fibronectin (Fittipaldi et al., 2012). The Streptococcal exotoxin suilysin may disrupt barrier function through cytotoxic effects on endothelial and epithelial cells, but it is not required for invasion. Capsular polysaccharide, which is otherwise considered a major virulence factor for this bacterium, may actually reduce S. suis invasion in BMECs and CPECs by covering up cell wall adhesins (Vanier et al., 2007, 2009a).

Interactions between Streptococcus suis serotype 2 and cultured porcine CPEC monolayers in a Transwell system have been the subject of recent investigations (Tenenbaum et al., 2009; Schroten et al., 2012). These have revealed that S. suis shuttles across the choroid plexus epithelium and enters the CSF in a polarized fashion. Bacteria cross the basolateral membranes of epithelial cells, are enclosed in endocytic vacuoles and trafficked to the apical membrane, and the cocci are subsequently released intact into the CSF via exocytosis. This process, which involves virulence factors and direct bacterial-epithelial contacts, disrupts blood-CSF barrier function and enhances the entry of additional bacteria and white blood cells into the CSF (Fittipaldi et al., 2012). Administration of an antiinflammatory glucocorticoid (dexamethasone) inhibits S. suis-induced alterations in tight junction re-organization and breakdown in CPECs (Tenenbaum et al., 2008). Although the mechanistic aspects of S. suis migration across BMECs are less well-defined, it is known that this organism can reside within porcine BMECs for up to 7 hours (Vanier et al., 2004). 

Cells comprising the neurovascular unit that participate in first line defense against pathogenic microbes must have the ability to recognize conserved, pathogen-associated molecular patterns, such as the presence of lipopolysaccharide or lipopeptides on bacterial surfaces. BMECs and neighboring cells express a number of pathogen recognition receptors, including Toll- and NOD-like receptors, and virus-detecting RIG helicases (Kristensson, 2011; van Sorge and Doran, 2012; Lampron et al., 2013). Streptococcal cell wall components can activate these receptors in BMECs and CPECs to produce pro-inflammatory responses, which can potentially degrade barrier function if unchecked (Schwerk et al., 2011; Fittipaldi et al. 2012). Inflammatory mediators, including cytokines, chemokines and arachidonic acid metabolites, facilitate the attachment of leukocytes to the basement membranes of BMECs and, by loosening tight junctions, allow the paracellular transmigration of these immunocytes into the CNS (Lampron et al., 2013). Neutrophils appear to move directly through porcine CPECs in response to inflammatory mediators formed during S. suis infection (Wewer et al., 2011). Streptococci c an d egrade t he c hemokine i nterleukin 8 generated by porcine BMECs and thereby slow the appearance of neutrophils at sites of infection (Vanier et al., 2009b). On the other hand, exposure of porcine CPECs to the inflammatory cytokine interferon-g increases the activity of epithelial indolamine dioxygenase, an enzyme that degrades tryptophan necessary for S. suis growth. CPECs can thus have a bacteriostatic effect on this organism (Adam et al., 2004). 

Although the blood-brain barrier function of BMECs and the overall neurovascular unit appears does not appear to mature with age, the blood-CSF barrier of neonates is generally more permeable than that of adults (Saunders et al., 2012). Relatively large proteins such as IgG from colostrum or bilirubin can enter the CSF in suckling piglets (Lee et al., 1995; Harada et al., 2002). This barrier may be particularly vulnerable to S. suis infection in neonatal swine. However, it may likewise be more permeable to antimicrobial drugs, such as b-lactam antibiotics that are routinely used to treat Streptococcal infections in swine and humans and to which microbial resistance is currently low (Nau et al., 2010; Varela et al., 2013). Meningitis is associated with increased permeability to hydrophilic drugs (e.g. penicillin) that normally penetrate the blood-brain barrier poorly (Lutsar et al., 1998). Drug accumulation in the CSF to concentrations effective in arresting Streptococcal meningeal infections may even be enhanced, based on studies in neonatal rodents reporting a reduced expression of drug efflux pumps such as P-glycoprotein (Tsai et al., 2002). 

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