The effect of astrocyte-derived fatty acid-binding protein 7 on blood-brain barrier integrity in LPS-induced inflammation

Sepsis is a life-threatening systemic inflammatory disease caused by infection, which causes disruption of blood-brain barrier (BBB) integrity. Fatty acid-binding protein 7 (FABP7), a transporter of fatty acids across the plasma membrane in astrocytes of the brain, modulates the caveolae formation in response to damage. However, the role of FABP7 in BBB disruption in the setting of septic inflammatory conditions caused by lipopolysaccharide (LPS) has not been clear yet. This study investigates the in vitro and in vivo effects of exogenous FABP7 administration on LPS-induced BBB disruption. bEnd.3 cells were treated with 0.5 µg/mL LPS for 24 hours followed by FABP7 (0.5 µg/mL) for 24 hours. Transendothelial electrical resistance (TEER) was measured to evaluate BBB integrity, whereas BBB permeability was assessed using sodium fluorescein (NaFl) tracer. Septic mice model was induced by intraperitoneally injecting LPS (3 mg/kg) and one hour later, the animals were treated with FABP7 (40 and 80 μg/kg). Body temperature was measured, and blood samples were collected from all experimental groups to determine the sepsis parameters. The anxiety levels of animals were measured by elevated plus maze test. The BBB permeability was evaluated by assessing the extravasation of albumin Alexa fluor 594 tracer injected after 23 hours of LPS and/or FABP7 treatments by IVIS imaging and confocal scanning fluorescence microscopy. The expression levels of tight junction (TJ) proteins claudin-5 and zonula occludens (ZO)-1, transcytosis-related proteins caveolin-1 and major facilitator superfamily domain containing 2a (mfsd2a), and inflammatory mediators NF-κB and interleukin-1β were determined by immunofluorescence staining and western blotting. In vitro FABP7 administration to bEnd.3 cells following LPS challenge significantly increased the lowered TEER values (p< 0.05) and decreased the enhanced NaFl leakage (p< 0.05). In LPS-injected animals, 80 μg/kg FABP7 treatment decreased the elevated body temperature from 40⁰C to 37-38⁰C. Serum levels of procalcitonin (PCT), C-reactive protein (CRP), and reactive oxygen species (ROS) were significantly increased by LPS injection, whereas high dose FABP7 markedly decreased the levels of sepsis parameters (p< 0.05). In LPS-treated animals, a significant increase was observed in the severity of anxiety-like behavior which was significantly decreased by 80 μg/kg FABP7 (p< 0.05). LPS treatment increased the extravasation of albumin Alexa fluor 594 in the brain while high dose FABP7 decreased the BBB leakage of the tracer (p< 0.05). When the expressions of TJ proteins were evaluated following the LPS challenge, a substantial increase in the decreased claudin-5 expression was noted by FABP7 treatment in vitro and in vivo (p< 0.0001), while no statistically significant change was observed in the expression level of ZO-1 in bEnd.3 cells. In vitro and in vivo FABP7 administration following LPS significantly decreased the increased expression level of caveolin-1 (p< 0.0001), while a significant increase was found in the decreased expression level of mfsd2a (p< 0.0001). In terms of alterations in the inflammatory response, the increased levels of NF-κB in vitro and in vivo (p< 0.01) and interleukin-1β in vitro (p< 0.001) were markedly reduced by FABP7 treatment following LPS-induced BBB disruption. Our results suggest, for the first time, that exogenous FABP7 may account for a new pharmacological tool in the treatment of inflammation- and sepsis-related BBB disruption.