Effects of Immunomodulatory and Organism-Associated Molecules on the Permeability of an In Vitro Blood-Brain Barrier Model to Amphotericin B and Fluconazole

Amphotericin B (AMB) is used to treat fungal infections of the central nervous system (CNS). However, AMB shows poor penetration into the CNS and little is known about the factors affecting its permeation through the blood-brain barrier (BBB). Therefore, we studied immunomodulatory and organism-associated molecules affecting the permeability of an in vitro BBB model to AMB. We examined the effects of interleukin-1 beta (IL-1β), tumor necrosis factor alpha (TNF-α), lipopolysaccharide (LPS), lipoteichoic acid (LTA), zymosan (ZYM), dexamethasone (DEX), cyclosporine, and tacrolimus on transendothelial electrical resistance (TEER); endothelial tight junctions; filamentous actin; and permeability to deoxycholate AMB (DAMB), liposomal AMB (LAMB), and fluconazole. Proinflammatory cytokines and organism-associated molecules significantly decreased the mean TEER by 40.7 to 100% (P ≤ 0.004). DEX increased the mean TEER by 18.2 to 26.4% (P ≤ 0.04). TNF-α and LPS increased the permeability to AMB by 8.2 to 14.5% compared to that for the controls (1.1 to 2.4%) (P ≤ 0.04). None of the other molecules affected the model''s permeability to AMB. By comparison, the BBB model''s permeability to fluconazole was >78% under all conditions studied, without significant differences between the controls and the experimental groups. LPS and TNF-α decreased tight-junction protein zona occludens 1 (ZO-1) between endothelial cells. In conclusion, IL-1β, ZYM, and LTA increased the permeability of the BBB to small ions but not to AMB, whereas TNF-α and LPS, which disrupted the endothelial layer integrity, increased the permeability to AMB.Amphotericin B (AMB) and fluconazole are important antifungal agents used to treat invasive fungal infections of the central nervous system (CNS). AMB is commonly used in the treatment of cryptococcal meningitis, rhinocerebral zygomycosis, hematogenous candida meningoencephalitis, and other invasive fungal infections of the CNS (7, 16, 18). AMB was effective for the treatment of experimental hematogenous candida meningoencephalitis in rabbits, but it demonstrated poor penetration into the cerebrospinal fluid (CSF) and brain tissue (10). While the penetration of AMB into CSF and brain parenchyma is poor, it remains an effective agent against CNS mycoses (20). By comparison, fluconazole displays a relatively high level of penetration into the CNS (2). Variables which might influence the permeability of the blood-brain barrier (BBB) to AMB, such as cytokines, organism-associated molecules, and immunosuppressants, have not been fully evaluated. We therefore studied the effects of several such factors on an in vitro mammalian BBB model. We examined the effects of those variables on interendothelial tight junctions; the structure of filamentous actin; and the permeability of the in vitro BBB model to AMB, fluconazole, and small ions.