| Myelin is formed by oligodendrocytes in the central nervous system and Schwann cells in the peripheral nervous system. This unique membrane is comprised of approximately 70% lipid and 30% protein. The high lipid content is thought to be vital for its insulatory function. Galactosylceramide (Ga1Cer) and sulfatide make up approximately 30% of total myelin lipids, with more than half of these galactolipids containing fatty acids hydroxylated at the C2 position (2-hydroxy fatty acids). Despite their high abundance, very little is know about the biosynthesis of these 2-hydroxy galactolipids, and specific functions of the 2-hydroxyl group in myelin galactolipids remain speculative. To fill this gap, we recently cloned and characterized a human fatty acid 2-hydroxylase gene, FA2H, that is highly expressed in brain. To study the roles of fatty acid 2-hydoxylase and 2-hydroxy sphingolipids in the brain and other tissues, we have developed highly sensitive methodologies to measure in vitro fatty acid 2-hydroxylase and free 2-hydroxy fatty acids by gas chromatography/mass spectrometry (GC/MS). Utilizing these novel methods, here we demonstrate that FA2H is required for the formation of 2-hydroxy fatty acids (precursors of 2-hydroxy galactolipids) in the central and peripheral nervous systems, and 2-hydroxylation of free fatty acids is the first step for the biosynthesis of 2-hydroxy galactolipids. During the course of the study we developed FA2H knockdown using siRNA and shRNA. RNAi against FA2H revealed unexpected cellular phenotypes indicative of altered differentiation. Cells were 2.8-fold more migratory, exhibited increased proliferation, and were strongly resistant to cAMP induced differentiation.{09}These findings open an exciting area of research involving 2-hydroxy lipids in cell growth, migration, and differentiation. |
