Analysis of bioactive oxysterols in newborn mouse brain by LC/MS

Meljon, Anna; Theofilopoulos, Spyridon; Shackleton, Cedric; Watson, Gordon; Javitt, Norman B.; Knölker, Hans‐Joachim; Saini, Ratni; Arenas, Ernest; Wang, Yuqin; Griffiths, William J.

doi:10.1194/jlr.d028233

Cited by 49 publications

(66 citation statements)

References 65 publications

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“…Until now, squalene 2,3;22,23-diepoxide was the only known biosynthetic precursor to 24 S ,25-epoxycholesterol ( 43 ), but our study reveals a novel pathway for the formation of this biologically important oxysterol. 24 S ,25-epoxycholesterol was found to be the most potent and abundant oxysterol ligand of LXRs in developing mouse brain (signifi cantly higher than the level of 24 Shydroxycholesterol), and it promotes neurogenesis and neuronal survival in an LXR-dependent fashion ( 44,45 ). Notably, the levels of desmosterol are also high in developing brains (up to 30% of total brain sterols) (46)(47)(48) and defi ciency in cholesterol.…”

Section: Discussionmentioning

confidence: 99%

“…The oxysterols derived from these enzymatic pathways could have signifi cant biological consequences in related physiological or pathological conditions where the levels of these two cholesterol precursors are high, such as in SLOS (7-dehydrocholesterol) ( 7,8 ), desmosterolosis (desmosterol) ( 7,54 ), and developing brain (desmosterol) (46)(47)(48). The direct conversion of desmosterol to 24 S ,25-epoxycholesterol by P450 46A1 represents a distinctly new pathway for the formation of this potent oxysterol ligand of LXRs ( 4,44,45 ). low in the adult brain ( ‫ف‬ 0.1% of cholesterol) ( 49 ).…”

Section: Discussionmentioning

confidence: 99%

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Oxidation of 7-dehydrocholesterol and desmosterol by human cytochrome P450 46A1

Goyal

Xiao

Porter

et al. 2014

Journal of Lipid Research

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Oxidation of 7-dehydrocholesterol and desmosterol by human cytochrome P450 46A1

Goyal

Xiao

Porter

et al. 2014

Journal of Lipid Research

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“…Charge-tagging of sterols. The sterols were charge-tagged with the GP-hydrazine as described previously (7,55,56). This greatly enhances their response when analyzed by LC-ESI-MS and tandem mass spectrometry (MS n ).…”

Section: Methodsmentioning

confidence: 99%

“…Sterols were extracted from CSF, plasma, or mouse brain into ethanol and fractionated by reversed phase solid phase extraction (SPE) to give a cholestenoic acid-and oxysterol-rich fraction devoid of cholesterol (7,55,56). Charge-tagging of sterols.…”

Section: Methodsmentioning

confidence: 99%

Cholestenoic acids regulate motor neuron survival via liver X receptors

Theofilopoulos¹,

Griffiths²,

Crick³

et al. 2014

J. Clin. Invest.

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160

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tion-mass spectrometry (LC-ESI-MS) analysis. We hereby report the exact identity of 16 oxysterols and downstream metabolites, including cholestenoic acids, found in human CSF (Supplemental Table 1; supplemental material available online with this article; doi:10.1172/JCI68506DS1). The most abundant of these metabolites (19.48-0.40 ng/ml; Supplemental Figure 1) were 7α-hydroxy-3-oxocholest-4-en-26-oic acid (7αH,3O-CA), 3β-hydroxycholest-5-en-26-oic acid (3β-HCA), and 2 newly identified metabolites in CSF, 3β,7α-diHCA and 3β,7β-dihydroxycholest-5-en-26-oic acid (3β,7β-diHCA). Precursors of these acids, including 26-HC and newly identified 7α,26-dihydroxycholesterol (7α,26-diHC; cholest-5-ene-3β,7α,26-triol) and 7α,26-dihydroxycholest-4-en-3-one (7α,26-diHCO), were also found, but at lower levels (0.15-0.03 ng/ml). Our results thus identified 4 novel oxysterol metabolites in human CSF that were downstream of 26-HC ( Figure 1A). 26-HC is metabolized via 7α,26-diHC and 7α,26-diHCO, or via 3β-HCA and 3β,7α-diHCA, to 7αH,3O-CA. While 26-HC can cross the blood-brain barrier (BBB) and enter the brain from the circulation (25), 7αH,3O-CA traverses the BBB and is exported from the brain (26). Very low levels of 24S-hydroxycholesterol (24S-HC; cholest-5-ene-3β,24S-diol), 25-hydroxycholesterol (25-HC; cholest-5-ene-3β,25-diol), and newly identified 7α,25-dihydroxycholesterol (7α,25-diHC; cholest-5-ene-3β,7α,25-triol) and 7α,25-dihydroxycholest-4-en-3-one (7α,25-diHCO) were also found in CSF (0.08-0.03 ng/ml).Reduced levels of 7α-hydroxylated cholestenoic acids in CSF and plasma/serum of human patients with SPG5. SPG5 presents with upper motor neuron signs and results from mutations in CYP7B1, encoding the oxysterol 7α-hydroxylase responsible for 7α-hydroxylation of side-chain oxidized sterols that is required for extrahepatic synthesis of 7αH,3O-CA and its precursor, 3β,7α-diHCA ( Figure 1A and ref. 18). In order to examine the pathogenic role of such mutations, we sought to identify alterations in oxysterol and cholestenoic acid profiles in CSF and plasma from these patients and then examine the biological activities of the altered metabolites. We first studied the CSF from 3 patients with SPG5

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“…Girard reagents ( 23 ), a family of quaternary ammonium hydrazides ( 24 ) typically comprising pyridine or trimethylamine (TMA) moieties, were employed for the derivatization of 3-ketone groups in mammalian oxysterols produced by their enzymatic oxidation (25)(26)(27)(28): converting charge-neutral oxysterols to strongly positively charged hydrazones enhanced their solubility and ionization capacity (an approach recently termed "charge-tagging") ( 29 ). If subjected to MS/MS, Girard hydrazones undergo LC-MS/MS ( 8,20 ); however, they appeared to be far less sensitive compared with RIA.…”

Section: Supplementary Abstract Ecdysone • Drosophila Melanogaster •mentioning

confidence: 99%