Ryan W. Mitchell

Fatty acid transport protein expression in human brain and potential role in fatty acid transport across human brain microvessel endothelial cells

J. Neurochem. (2011) 117, 735–746.

Fatty acid transport into the brain: Of fatty acid fables and lipid tails

The blood-brain barrier formed by the brain capillary endothelial cells provides a protective barrier between the systemic blood and the extracellular environment of the central nervous system. Brain capillaries are a continuous layer of endothelial cells with highly developed tight junctional complexes and a lack of fenestrations. The presence of these tight junctions in the cerebral microvessel endothelial cells aids in the restriction of movement of molecules and solutes into the brain. Fatty acids are important components of biological membranes, are precursors for the biosynthesis of phospholipids and sphingolipids and are utilized for mitochondrial β-oxidation. The brain is capable of synthesizing only a few fatty acids. Hence, most fatty acids must enter into the brain from the blood. Here we review current mechanisms of transport of free fatty acids into cells and describe how free fatty acids move from the blood into the brain. We discuss both diffusional as well as protein-mediated movement of fatty acids across biological membranes.

On the mechanism of oleate transport across human brain microvessel endothelial cells

The blood–brain barrier formed by the brain capillary endothelial cells provides a protective barrier between the systemic blood and the extracellular environment of the CNS. As most fatty acids in the brain enter from the blood, we examined the mechanism of oleate (C18:1) transport across primary human brain microvessel endothelial cells (HBMEC). The permeability of [1‐14C]oleate was determined using confluent cells grown on Transwell® inserts in both the absence or presence of bovine serum albumin in the basolateral media, and following inhibition of various fatty acid transporters. The passage of [1‐14C]oleate across confluent HBMEC monolayers was significantly enhanced when fatty acid free albumin was present in the basolateral media. The presence of the non‐specific fatty acid uptake inhibitor phloretin significantly decreased [1‐14C]oleate uptake by HBMEC and the subsequent release of [1‐14C]oleate into the basolateral medium. Knockdown of fatty acid transport protein‐1 or fatty acid translocase/CD36 significantly decreased [1‐14C]oleate transport across the HBMEC monolayer from either apical as well as basolateral sides. The findings indicate that a fatty acid acceptor is a requirement for oleate transport across HBMEC monolayers. In addition, transport of oleate across HBMEC is, in part, a transcellular process mediated by fatty acid transport proteins.

Human Trifunctional Protein Alpha Links Cardiolipin Remodeling to Beta-Oxidation

Cardiolipin (CL) is a mitochondrial membrane phospholipid which plays a key role in apoptosis and supports mitochondrial respiratory chain complexes involved in the generation of ATP. In order to facilitate its role CL must be remodeled with appropriate fatty acids. We previously identified a human monolysocardiolipin acyltransferase activity which remodels CL via acylation of monolysocardiolipin (MLCL) to CL and was identical to the alpha subunit of trifunctional protein (αTFP) lacking the first 227 amino acids. Full length αTFP is an enzyme that plays a prominent role in mitochondrial β-oxidation, and in this study we assessed the role, if any, which this metabolic enzyme plays in the remodeling of CL. Purified human recombinant αTFP exhibited acyl-CoA acyltransferase activity in the acylation of MLCL to CL with linoleoyl-CoA, oleoyl-CoA and palmitoyl-CoA as substrates. Expression of αTFP increased radioactive linoleate or oleate or palmitate incorporation into CL in HeLa cells. Expression of αTFP in Barth Syndrome lymphoblasts, which exhibit reduced tetralinoleoyl-CL, elevated linoleoyl-CoA acylation of MLCL to CL in vitro, increased mitochondrial respiratory Complex proteins and increased linoleate-containing species of CL. Knock down of αTFP in Barth Syndrome lymphoblasts resulted in greater accumulation of MLCL than those with normal αTFP levels. The results clearly indicate that the human αTFP exhibits MLCL acyltransferase activity for the resynthesis of CL from MLCL and directly links an enzyme of mitochondrial β-oxidation to CL remodeling.

Delineating the role of alterations in lipid metabolism to the pathogenesis of inherited skeletal and cardiac muscle disorders Thematic Review Series: Genetics of Human Lipid Diseases

As the specific composition of lipids is essential for the maintenance of membrane integrity, enzyme function, ion channels, and membrane receptors, an alteration in lipid composition or metabolism may be one of the crucial changes occurring during skeletal and cardiac myopathies. Although the inheritance (autosomal dominant, autosomal recessive, and X-linked traits) and underlying/defining mutations causing these myopathies are known, the contribution of lipid homeostasis in the progression of these diseases needs to be established. The purpose of this review is to present the current knowledge relating to lipid changes in inherited skeletal muscle disorders, such as Duchenne/Becker muscular dystrophy, myotonic muscular dystrophy, limb-girdle myopathic dystrophies, desminopathies, rostrocaudal muscular dystrophy, and Dunnigan-type familial lipodystrophy. The lipid modifications in familial hypertrophic and dilated cardiomyopathies, as well as Barth syndrome and several other cardiac disorders associated with abnormal lipid storage, are discussed. Information on lipid alterations occurring in these myopathies will aid in the design of improved methods of screening and therapy in children and young adults with or without a family history of genetic diseases.

Exogenous arachidonic acid mediates permeability of human brain microvessel endothelial cells through prostaglandin E2 activation of EP3 and EP4 receptors.

The blood–brain barrier, formed by microvessel endothelial cells, is the restrictive barrier between the brain parenchyma and the circulating blood. Arachidonic acid (ARA; 5,8,11,14‐cis‐eicosatetraenoic acid) is a conditionally essential polyunsaturated fatty acid [20:4(n − 6)] and is a major constituent of brain lipids. The current study examined the transport processes for ARA in confluent monolayers of human brain microvascular endothelial cells (HBMEC). Addition of radioactive ARA to the apical compartment of HBMEC cultured on Transwell® inserts resulted in rapid incorporation of radioactivity into the basolateral medium. Knock down of fatty acid transport proteins did not alter ARA passage into the basolateral medium as a result of the rapid generation of prostaglandin E2 (PGE2), an eicosanoid known to facilitate opening of the blood–brain barrier. Permeability following ARA or PGE2 exposure was confirmed by an increased movement of fluorescein‐labeled dextran from apical to basolateral medium. ARA‐mediated permeability was attenuated by specific cyclooxygenase‐2 inhibitors. EP3 and EP4 receptor antagonists attenuated the ARA‐mediated permeability of HBMEC. The results indicate that ARA increases permeability of HBMEC monolayers likely via increased production of PGE2 which acts upon EP3 and EP4 receptors to mediate permeability. These observations may explain the rapid influx of ARA into the brain previously observed upon plasma infusion with ARA. The blood–brain barrier, formed by microvessel endothelial cells, is a restrictive barrier between the brain parenchyma and the circulating blood. Radiolabeled arachidonic acid (ARA) movement across, and monolayer permeability in the presence of ARA, was examined in confluent monolayers of primary human brain microvessel endothelial cells (HBMECs) cultured on Transwell® plates. Incubation of HBMECs with ARA resulted in a rapid increase in HBMEC monolayer permeability. The mechanism was mediated, in part, through increased prostaglandin E2 production from ARA which acted upon EP3 and EP4 receptors to increase HBMEC monolayer permeability.