Fatty Acid-Binding Protein 5 Facilitates the Blood-Brain Barrier Transport of Docosahexaenoic Acid

The brain has a limited ability to synthesize the essential polyunsaturated fatty acid (PUFA) docosahexaenoic acid (DHA) from its omega-3 fatty acid precursors. Therefore, to maintain brain concentrations of this PUPA at physiological levels, plasma-derived DHA must be transported across the blood brain barrier (BBB). While DHA is able to partition into the luminal membrane of brain endothelial cells, its low aqueous solubility likely limits its cytosolic transfer to the abluminal membrane, necessitating the requirement of an intracellular carrier protein to facilitate trafficking of this PUFA across the BBB. As the intracellular carrier protein fatty acid-binding protein 5 (FABPS) is expressed at the human BBB, the current study assessed the putative role of FABPS in the brain endothelial cell uptake and BBB transport of DHA in vitro and in vivo, respectively. hFAPB5 was recombinantly expressed and purified from Escherichia coli C41(DE3) cells and the binding affinity of DHA to hFABPS assessed using isothermal titration calorimetry. The impact of FABPS siRNA on uptake of C-14-DHA into immortalized human brain microvascular endothelial (hCMEC/D3) cells was assessed. An in situ transcardiac perfusion method was optimized in C57BL/6 mice and subsequently used to compare the BBB influx rate MO of C-14-DHA between FABPS-deficient (FABP5(-/-)) and wild-type (FABP5(+/+)) C57BL/6 mice. DHA bound to hFABPS with an equilibrium dissociation constant of 155 +/- 8 nM (mean +/- SEM). FABPS siRNA transfection decreased hCMEC/D3 mRNA and protein expression of FABPS by 53.2 +/- 5.5% and 44.8 +/- 13.7%, respectively, which was associated with a 14.1 +/- 2.7% reduction in C-14-DHA cellular uptake. By using optimized conditions for the in situ transcardiac perfusion (a 1 min preperfusion (10 mL/min) followed by perfusion of C-14-DHA (1 min)), the Kin of C-14-DHA was 0.04 +/- 0.01 mL/g/s. Relative to FABP5(+/+) mice, the K-in of C-14-DHA decreased 36.7 +/- 12.4% in FABP5(-/-)mice. This study demonstrates that FABPS binds to DHA and is involved in the brain endothelial cell uptake and subsequent BBB transport of DHA, confirming the importance of this cytoplasmic carrier protein in the CNS exposure of this PUFA essential for neuronal function.