title: Profiling and Imaging Ion Mobility-Mass Spectrometry Analysis of Cholesterol and 7-Dehydrocholesterol in Cells Via Sputtered Silver MALDI.
abstract: Profiling and imaging of cholesterol and its precursors by mass spectrometry (MS) are important in a number of cholesterol biosynthesis disorders, such as in Smith-Lemli-Opitz syndrome (SLOS), where 7-dehydrocholesterol (7-DHC) is accumulated in affected individuals. SLOS is caused by defects in the enzyme that reduces 7-DHC to cholesterol. However, analysis of sterols is challenging because these hydrophobic olefins are difficult to ionize for MS detection. We report here sputtered silver matrix-assisted laser desorption/ionization (MALDI)-ion mobility-MS (IM-MS) analysis of cholesterol and 7-DHC. In comparison with liquid-based AgNO3 and colloidal Ag nanoparticle (AgNP), sputtered silver NP (10-25 nm) provided the lowest limits-of-detection based on the silver coordinated [cholesterol + Ag](+) and [7-DHC + Ag](+) signals while minimizing dehydrogenation products ([M + Ag-2H](+)). When analyzing human fibroblasts that were directly grown on poly-L-lysine-coated ITO glass plates with this technique, in situ, the 7-DHC/cholesterol ratios for both control and SLOS human fibroblasts are readily obtained. The m/z of 491 (specific for [7-DHC + (107)Ag](+)) and 495 (specific for [cholesterol + (109)Ag](+)) were subsequently imaged using MALDI-IM-MS. MS images were co-registered with optical images of the cells for metabolic ratio determination. From these comparisons, ratios of 7-DHC/cholesterol for SLOS human fibroblasts are distinctly higher than in control human fibroblasts. Thus, this strategy demonstrates the utility for diagnosing/assaying the severity of cholesterol biosynthesis disorders in vitro.
authors.affiliation: Department of Chemistry, Vanderbilt University, Nashville, TN, 37235, USA.
authors.forename: Libin
authors.initials: L
authors.lastname: Xu
authors.forename: Michal
authors.initials: M
authors.lastname: Kliman
authors.forename: Jay G
authors.initials: JG
authors.lastname: Forsythe
authors.forename: Zeljka
authors.initials: Z
authors.lastname: Korade
authors.forename: Anthony B
authors.initials: AB
authors.lastname: Hmelo
authors.forename: Ned A
authors.initials: NA
authors.lastname: Porter
authors.forename: John A
authors.initials: JA
authors.lastname: McLean
mesh_terms: D002460:Cell Line; D002784:Cholesterol / Q000032:analysis*; D003684:Dehydrocholesterols / Q000032:analysis*; D005347:Fibroblasts / Q000473:pathology*; D006801:Humans; D061848:Optical Imaging / Q000379:methods; D012834:Silver / Q000737:chemistry; D019082:Smith-Lemli-Opitz Syndrome / Q000473:pathology*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
chemical_list: D003684:Dehydrocholesterols; D012834:Silver; D002784:Cholesterol; C016705:7-dehydrocholesterol
references.citation: J Lipid Res. 2011 Jan;52(1):6-34
references.pmid: 20929975
references.citation: Rapid Commun Mass Spectrom. 2010 Apr 30;24(8):1147-54
references.pmid: 20301106
references.citation: Prague Med Rep. 2007;108(3):263-9
references.pmid: 18399064
references.citation: Anal Chem. 2003 Jul 15;75(14):3429-34
references.pmid: 14570193
references.citation: Neuropharmacology. 2008 Dec;55(8):1265-73
references.pmid: 18402986
references.citation: Anal Chem. 2006 Feb 1;78(3):734-42
references.pmid: 16448046
references.citation: Microsc Res Tech. 2004 Dec;65(6):282-6
references.pmid: 15662659
references.citation: Anal Chem. 2013 Feb 5;85(3):1926-31
references.pmid: 23320778
references.citation: FEBS Lett. 2004 May 21;566(1-3):291-3
references.pmid: 15147911
references.citation: Anal Chem. 2013 Mar 19;85(6):3318-24
references.pmid: 23425078
references.citation: Science. 1996 Oct 11;274(5285):255-9
references.pmid: 8824192
references.citation: Science. 1997 Jul 11;277(5323):228-31
references.pmid: 9211849
references.citation: Anal Chem. 1998 Oct 15;70(20):4394-400
references.pmid: 9796422
references.citation: J Am Chem Soc. 2010 Feb 24;132(7):2222-32
references.pmid: 20121089
references.citation: J Microsc. 2004 Aug;215(Pt 2):156-61
references.pmid: 15315502
references.citation: J Am Chem Soc. 2009 Sep 16;131(36):13037-44
references.pmid: 19705847
references.citation: J Lipid Res. 1991 Apr;32(4):603-12
references.pmid: 1856606
references.citation: J Inherit Metab Dis. 2008 Dec;31 Suppl 2:S387-93
references.pmid: 18949577
references.citation: Mol Cancer Ther. 2008 Dec;7(12):3707-18
references.pmid: 19074846
references.citation: Anal Chem. 2008 Sep 1;80(17):6796-9
references.pmid: 18671412
references.citation: J Am Chem Soc. 2005 Apr 20;127(15):5304-5
references.pmid: 15826152
references.citation: J Am Soc Mass Spectrom. 2005 Oct;16(10):1654-9
references.pmid: 16095912
references.citation: J Lipid Res. 2013 Apr;54(4):1135-43
references.pmid: 23381570
references.citation: Neuroscience. 2010 Oct 27;170(3):858-64
references.pmid: 20670678
references.citation: Curr Opin Lipidol. 2001 Apr;12(2):105-12
references.pmid: 11264981
references.citation: Annu Rev Cell Dev Biol. 1998;14:111-36
references.pmid: 9891780
references.citation: Angew Chem Int Ed Engl. 1999 Apr 1;38(7):992-995
references.pmid: 29711860
references.citation: J Lipid Res. 2010 Nov;51(11):3259-69
references.pmid: 20702862
references.citation: Annu Rev Biochem. 1983;52:223-61
references.pmid: 6311077