Effective tight junctions are an important element of a fully differentiated BBB. This is largely because the physical barrier to hydrophilic molecules thus provided is one of the defining roles of the BBB. As mentioned above, polarization of transporters to apical and basolateral membrane domains also requires effective tight junctions. Hence the two features for pharmaceutical applications, tightness and polarized expression of transporters, depend on the tight junctions. This readily explains why the two models most commonly used in pharmaceutical assays are the bovine brain endothelial model (with or without astrocytes)37,46,61 and the porcine model (with or without hydrocortisone).59 Furthermore, scientific study of the mechanisms underlying polarized transendothelial transport, whether mediated by membrane carrier proteins or by vesicles (RMTor AMT), requires tight junctions restricting enough to allow resolution of the transport without excessive shunting (short-circuiting flux) via the junctions.94
The tightness of an endothelial or epithelial layer can be assessed by two measures of paracellular permeability, the transendothelial (or epithelial) electrical resistance (TEER, ohm cm2) and the transmonolayer permeability to a hydrophilic tracer molecule with negligible cell uptake or binding. Suitable permeability markers of a size similar to small drug molecules (typically 100-400 Da mol.wt.) are sucrose and mannitol (used as radiolabeled probes) or fluorescent indicators such as fluorescein and Lucifer yellow.76 As electrical resistance is the reciprocal of conductance, related to the permeability of small ions, TEER is inversely proportional to the permeability or conductance of Na+ and Cl_ through the tight junctions, these being the major charge carriers in extracellular fluids.92
A plot of TEER versus small solute permeability through an epithelial or endothelial monolayer will give an asymptotic curve reflecting this relationship.95 Net solute flux increases as the sum of fluxes across individual junctional clefts, while TEER is most affected by areas with the lowest resistance, which shunt current flow. Hence a small flaw in a monolayer can cause a large drop in TEER with little drop in sucrose or fluorescein permeability. Monolayers with TEER greater than ~ 100-200 ohm cm2 give the expected relation between TEER and solute permeability,37'76'95 so TEER assessment is a useful way of checking the uniformity and tightness of a monolayer before starting a drug permeability assay. The ability of a monolayer to rank drug permeabilities (its dynamic range) will depend partly on its tightness, as this will determine the degree of paracellular leak of the drug, and hence the dynamic range of the model.
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