title: Marked change in the balance between CYP27A1 and CYP46A1 mediated elimination of cholesterol during differentiation of human neuronal cells.
abstract: Cholesterol metabolism in the brain is distinct from that in other tissues due to the fact that cholesterol itself is unable to pass across the blood-brain barrier. Elimination of brain cholesterol is mainly dependent on a neuronal-specific cytochrome P450, CYP46A1, catalyzing the conversion of cholesterol into 24(S)-hydroxycholesterol (24OHC), which is able to pass the blood-brain barrier. A suitable model for studying this elimination from human neuronal cells has not been described previously. It is shown here that differentiated Ntera2/clone D1 (NT2) cells express the key genes involved in brain cholesterol homeostasis including CYP46A1, and that the expression profiles of the genes observed during neuronal differentiation are those expected to occur in vivo. Thus there was a decrease in the mRNA levels corresponding to cholesterol synthesis enzymes and a marked increase in the mRNA level of CYP46A1. The latter increase was associated with increased levels of CYP46A1 protein and increased production of 24OHC. The magnitude of the secretion of 24OHC from the differentiated NT2 cells into the medium was similar to that expected to occur under in vivo conditions. An alternative to elimination of cholesterol by the CYP46A1 mechanism is elimination by CYP27A1, and the product of this enzyme, 27-hydroxycholesterol (27OHC), is also known to pass the blood-brain barrier. The CYP27A1 protein level decreased during the differentiation of the NT2 cells in parallel with decreased production of 27OHC. The ratio between 24OHC and 27OHC in the medium from the cultured cells increased, by a factor of 13, during the differentiation process. The results suggest that progenitor cells eliminate cholesterol in the form of 27OHC while neurogenesis induces a change to the CYP46A1 dependent pathway. Furthermore this study demonstrates that differentiated NT2 cells are suitable for studies of cholesterol homeostasis in human neurons.
authors.affiliation: Research Institute for Medicines and Pharmaceutical Sciences (iMedUL), Faculty of Pharmacy, University of Lisbon, Lisbon, Portugal.
authors.forename: Inês
authors.initials: I
authors.lastname: Milagre
authors.forename: Maria
authors.initials: M
authors.lastname: Olin
authors.forename: Maria João
authors.initials: MJ
authors.lastname: Nunes
authors.forename: Miguel
authors.initials: M
authors.lastname: Moutinho
authors.forename: Anita
authors.initials: A
authors.lastname: Lövgren-Sandblom
authors.forename: Maria João
authors.initials: MJ
authors.lastname: Gama
authors.forename: Ingemar
authors.initials: I
authors.lastname: Björkhem
authors.forename: Elsa
authors.initials: E
authors.lastname: Rodrigues
mesh_terms: D002454:Cell Differentiation / Q000235:genetics / Q000502:physiology*; D045744:Cell Line, Tumor; D053493:Cholestanetriol 26-Monooxygenase / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D000072556:Cholesterol 24-Hydroxylase; D018507:Gene Expression Regulation, Developmental / Q000235:genetics / Q000502:physiology; D006706:Homeostasis / Q000235:genetics; D006801:Humans; D055495:Neurogenesis / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000096:biosynthesis; D013250:Steroid Hydroxylases / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*
chemical_list: D012333:RNA, Messenger; D013250:Steroid Hydroxylases; D000072556:Cholesterol 24-Hydroxylase; C506831:CYP27A1 protein, human; D053493:Cholestanetriol 26-Monooxygenase