Summary of study ST000917

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR000633. The data can be accessed directly via it's Project DOI: 10.21228/M8V961 This work is supported by NIH grant, U2C- DK119886.

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Study IDST000917
Study TitleBiomarkers of NAFLD progression: a lipidomics approach to an epidemic. Part 3:Urine
Study TypeLipidomics Study
Study SummaryThe spectrum of nonalcoholic fatty liver disease (NAFLD) includes steatosis, nonalcoholic steatohepatitis (NASH), and cirrhosis. Recognition and timely diagnosis of these different stages, particularly NASH, is important for both potential reversibility and limitation of complications. Liver biopsy remains the clinical standard for definitive diagnosis. Diagnostic tools minimizing the need for invasive procedures or that add information to histologic data are important in novel management strategies for the growing epidemic of NAFLD. We describe an 'omics' approach to detecting a reproducible signature of lipid metabolites, aqueous intracellular metabolites, SNPs, and mRNA transcripts in a double-blinded study of patients with different stages of NAFLD that involves profiling liver biopsies, plasma, and urine samples. Using linear discriminant analysis, a panel of 20 plasma metabolites that includes glycerophospholipids, sphingolipids, sterols, and various aqueous small molecular weight components involved in cellular metabolic pathways, can be used to differentiate between NASH and steatosis. This identification of differential biomolecular signatures has the potential to improve clinical diagnosis and facilitate therapeutic intervention of NAFLD.
Institute
LIPID MAPS
DepartmentBioengineering
Last NameFahy
First NameEoin
Address9500 Gilman, La Jolla, CA, 92093, USA
Emailefahy@ucsd.edu
Phone858-534-4076
Submit Date2018-01-14
Publicationshttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC4340319/
Analysis Type DetailGC/LC-MS
Release Date2018-04-05
Release Version1
Eoin Fahy Eoin Fahy
https://dx.doi.org/10.21228/M8V961
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:


Project:

Project ID:PR000633
Project DOI:doi: 10.21228/M8V961
Project Title:Biomarkers of NAFLD progression: a lipidomics approach to an epidemic
Project Type:Lipidomics Study
Project Summary:The spectrum of nonalcoholic fatty liver disease (NAFLD) includes steatosis, nonalcoholic steatohepatitis (NASH), and cirrhosis. Recognition and timely diagnosis of these different stages, particularly NASH, is important for both potential reversibility and limitation of complications. Liver biopsy remains the clinical standard for definitive diagnosis. Diagnostic tools minimizing the need for invasive procedures or that add information to histologic data are important in novel management strategies for the growing epidemic of NAFLD. We describe an 'omics' approach to detecting a reproducible signature of lipid metabolites, aqueous intracellular metabolites, SNPs, and mRNA transcripts in a double-blinded study of patients with different stages of NAFLD that involves profiling liver biopsies, plasma, and urine samples. Using linear discriminant analysis, a panel of 20 plasma metabolites that includes glycerophospholipids, sphingolipids, sterols, and various aqueous small molecular weight components involved in cellular metabolic pathways, can be used to differentiate between NASH and steatosis. This identification of differential biomolecular signatures has the potential to improve clinical diagnosis and facilitate therapeutic intervention of NAFLD.
Institute:University of California, San Diego
Department:Bioengineering
Last Name:Fahy
First Name:Eoin
Address:9500 Gilman, La Jolla, CA, 92093, USA
Email:efahy@ucsd.edu
Phone:858-534-4076
Funding Source:NIGMS Grant GM U54069338
Publications:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4340319/
Contributors:LIPID MAPS Consortium

Subject:

Subject ID:SU000955
Subject Type:Human clinical study
Subject Species:Homo sapiens
Taxonomy ID:9606
Age Or Age Range:23-83
Gender:Male and Female
Human Ethnicity:Mixed

Factors:

Subject type: Human clinical study; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id local_sample_id Diagnosis
SA053978NASH055Cirrhosis
SA053979NASH049Cirrhosis
SA053980NASH048Cirrhosis
SA053981NASH064Cirrhosis
SA053982NASH068Cirrhosis
SA053983NASH005Cirrhosis
SA053984NASH069Cirrhosis
SA053985NASH047Cirrhosis
SA053986NASH065Cirrhosis
SA053987NASH052Cirrhosis
SA053988NASH016Cirrhosis
SA053989NASH013Cirrhosis
SA053990NASH040Cirrhosis
SA053991NASH007Cirrhosis
SA053992NASH022Cirrhosis
SA053993NASH009Cirrhosis
SA053994NASH026Cirrhosis
SA053995NASH029Cirrhosis
SA053996NASH028Cirrhosis
SA053997NASH027Cirrhosis
SA053998NASH072NASH
SA053999NASH057NASH
SA054000NASH044NASH
SA054001NASH074NASH
SA054002NASH088NASH
SA054003NASH039NASH
SA054004NASH090NASH
SA054005NASH087NASH
SA054006NASH084NASH
SA054007NASH031NASH
SA054008NASH018NASH
SA054009NASH012NASH
SA054010NASH038NASH
SA054011NASH010NASH
SA054012NASH019NASH
SA054013NASH015NASH
SA054014NASH037NASH
SA054015NASH035NASH
SA054016NASH030NASH
SA054017NASH021NASH
SA054018NASH067Normal
SA054019NASH070Normal
SA054020NASH066Normal
SA054021NASH060Normal
SA054022NASH077Normal
SA054023NASH053Normal
SA054024NASH054Normal
SA054025NASH086Normal
SA054026NASH091Normal
SA054027NASH051Normal
SA054028NASH089Normal
SA054029NASH085Normal
SA054030NASH080Normal
SA054031NASH082Normal
SA054032NASH078Normal
SA054033NASH014Normal
SA054034NASH017Normal
SA054035NASH020Normal
SA054036NASH006Normal
SA054037NASH004Normal
SA054038NASH050Normal
SA054039NASH003Normal
SA054040NASH024Normal
SA054041NASH011Normal
SA054042NASH043Normal
SA054043NASH045Normal
SA054044NASH034Normal
SA054045NASH042Normal
SA054046NASH046Normal
SA054047NASH041Normal
SA054048NASH036Normal
SA054049NASH075Steatosis
SA054050NASH073Steatosis
SA054051NASH071Steatosis
SA054052NASH076Steatosis
SA054053NASH081Steatosis
SA054054NASH083Steatosis
SA054055NASH062Steatosis
SA054056NASH079Steatosis
SA054057NASH002Steatosis
SA054058NASH032Steatosis
SA054059NASH023Steatosis
SA054060NASH001Steatosis
SA054061NASH033Steatosis
SA054062NASH056Steatosis
SA054063NASH059Steatosis
SA054064NASH058Steatosis
SA054065NASH061Steatosis
Showing results 1 to 88 of 88

Collection:

Collection ID:CO000949
Collection Summary:Human samples were collected according to a protocol approved by Vanderbilt University Medical Center's Internal Review Board (#120829) and under informed written patients' consent prior to inclusion in this study. Sample sizes were selected to minimize the invasive procedures. Plasma samples were obtained from patients' blood collected during standard of care surgical procedures. Urine samples were collected from patients' Foley catheters placed for standard of care procedure. Liver samples were obtained from the excess tissue collected as part of the standard of care liver biopsies performed at the time of surgery that would otherwise be discarded. Subsequently, studies at University of California, San Diego were conducted under further auspices of University of California, San Diego Internal Review Board #121220.
Sample Type:Urine

Treatment:

Treatment ID:TR000969
Treatment Summary:-

Sample Preparation:

Sampleprep ID:SP000962
Sampleprep Summary:Urine sample extraction. Five hundred microliters of thawed urine containing 4 µl of internal standard mix were extracted with 500 µl cold (-20°C) CH3OH by incubation in an ice bath for 30 min. After mixing by vortex at 4°C for 1 min and centrifugation (4°C, 18,000 g, 10 min), the solvent was evaporated from the supernatant and the residue was dissolved in 400 µl of resuspension solvent, vortexed to mix (1 min at 4°C), and centrifuged (4°C, 18,000 g, 10 min) to remove any insoluble material. GPLs: GPLs from liver samples were extracted and analyzed by MS essentially as described in (20, 21). Extraction and analysis of plasma samples was according to previously published procedures (22). Cardiolipin, coenzyme Q, and dolichol: Lipid extractions were performed based on the Bligh and Dyer method with minor modifications (23-25). FAs and eicosanoids: FFAs were extracted essentially as previously described after supplementation with deuterated internal standards (Cayman Chemicals) (26, 27). Eicosanoids were isolated via solid phase extraction, utilizing 25 deuterated internal standards (28, 29). Sterols and oxysterols: Sterols and oxysterols were extracted using previously described methods (30). Neutral lipids: Cholesteryl esters (CEs), TAGs, and DAGs were extracted from weighed liver tissue (0.5-1 mg) suspended in 0.5 ml PBS that had been homogenized by sonication. Extractions of plasma (0.05 ml diluted to 0.1 ml with PBS), urine (1 ml), and tissue sonicates were carried out using 1 ml hexane:methyl t-butyl ether (1:1, v/v), essentially as previously described (31). Sphingolipids: Sphingolipids from liver, plasma, and urine were extracted following previously published procedures (32, 33), with the exception that methylene chloride was substituted for chloroform for the single-phase extraction of sphingoid bases.
20. Ivanova P. T., Milne S. B., Byrne M. O., Xiang Y., Brown H. A. 2007. Glycerophospholipid identification and quantitation by electrospray ionization mass spectrometry. Methods Enzymol. 432: 21-57.
21. Myers D. S., Ivanova P. T., Milne S. B., Brown H. A. 2011. Quantitative analysis of glycerophospholipids by LC-MS: acquisition, data handling, and interpretation. Biochim. Biophys. Acta. 1811: 748-757.
22. Quehenberger O., Armando A. M., Brown H. A., Milne S. B., Myers D. S., Merrill A. H., Jr, Bandyopadhyay S., Jones K. N., Kelly S., Shaner R. L., et al. 2010. Lipidomics reveals a remarkable diversity of lipids in human plasma. J. Lipid Res. 51: 3299-3305.
23. Guan Z., Li S., Smith D., Shaw W., Raetz C. 2007. Identification of N-acylphosphatidylserine molecules in eukaryotic cells. Biochemistry. 46: 14500-14513.
24. Tan B. K., Bogdanov M., Zhao J., Dowhan W., Raetz C. R., Guan Z. 2012. Discovery of cardiolipin synthase utilizing phosphatidylethanolamine and phosphatidylglycerol as substrates. Proc. Natl. Acad. Sci. USA. 109: 16504-16509.
25. Wen R., Lam B., Guan Z. 2013. Aberrant dolichol chain lengths as biomarkers for retinitis pigmentosa caused by impaired dolichol biosynthesis. J. Lipid Res. 54: 3516-3522.
26. Quehenberger O., Armando A., Dumlao D., Stephens D. L., Dennis E. A. 2008. Lipidomics analysis of essential fatty acids in macrophages. Prostaglandins Leukot. Essent. Fatty Acids. 79: 123-129.
27. Quehenberger O., Armando A. M., Dennis E. A. 2011. High sensitivity quantitative lipidomics analysis of fatty acids in biological samples by gas chromatography-mass spectrometry. Biochim. Biophys. Acta. 1811: 648-656.
28. Deems R., Buczynski M. W., Bowers-Gentry R., Harkewicz R., Dennis E. A. 2007. Detection and quantitation of eicosanoids via high performance liquid chromatography-electrospray ionization-mass spectrometry. Methods Enzymol. 432: 59-82.
29. Dumlao D. S., Buczynski M. W., Norris P. C., Harkewicz R., Dennis E. A. 2011. High-throughput lipidomic analysis of fatty acid derived eicosanoids and N-acylethanolamines. Biochim. Biophys. Acta. 1811: 724-736.
30. McDonald J. G., Smith D. D., Stiles A. R., Russell D. W. 2012. A comprehensive method for extraction and quantitative analysis of sterols and secosteroids from human plasma. J. Lipid Res. 53: 1399-1409.
31. Hutchins P. M., Barkley R. M., Murphy R. C. 2008. Separation of cellular nonpolar neutral lipids by normal-phase chromatography and analysis by electrospray ionization mass spectrometry. J. Lipid Res. 49: 804-813.
32. Shaner R. L., Allegood J. C., Park H., Wang E., Kelly S., Haynes C. A., Sullards M. C., Merrill A. H., Jr 2009. Quantitative analysis of sphingolipids for lipidomics using triple quadrupole and quadrupole linear ion trap mass spectrometers. J. Lipid Res. 50: 1692-1707.
33. Sullards M. C., Liu Y., Chen Y., Merrill A. H., Jr 2011. Analysis of mammalian sphingolipids by liquid chromatography tandem mass spectrometry (LC-MS/MS) and tissue imaging mass spectrometry (TIMS). Biochim. Biophys. Acta. 1811: 838-853.

Combined analysis:

Analysis ID AN001497 AN001499 AN001500 AN001501
Analysis type MS MS MS MS
Chromatography type Core G Core K Core E Core I
Chromatography system Multiple Multiple Multiple Multiple
Column Multiple Multiple Multiple Multiple
MS Type Other Other Other Other
MS instrument type - - - -
MS instrument name
Ion Mode UNSPECIFIED UNSPECIFIED UNSPECIFIED UNSPECIFIED
Units pmol/ml pmol/ml pmol/ml pmol/ml

Chromatography:

Chromatography ID:CH001054
Chromatography Summary:FFAs were analyzed by stable isotope dilution GC-MS after derivatization, essentially as described previously (26, 27). This method quantifies 33 FAs including all major and minor saturated FAs, monounsaturated FAs, and PUFAs containing 12 to 26 carbons. Eicosanoids were analyzed by a stable isotope dilution LC/MS method utilizing 26 deuterated internal standards (28, 29). The metabolites were quantified after separation by reverse phase chromatography on a 2.1 × 100 mm BEH Shield column, 1.7 µM (Waters, Milford, MA) employing an Acquity UPLC system (Waters). Detection and quantification were performed on an AB SCIEX 6500 QTrap mass spectrometer equipped with an IonDrive Turbo V source (AB SCIEX, Framingham, MA), operated in negative ionization mode via MRM, using standard curves generated from 145 authentic quantification standards (34). The method analyzes an additional 13 metabolites based on authentic primary standards, but which cannot be quantified due to the lack of appropriate internal standards. Data analysis was performed using MultiQuant 2.1 software (AB SCIEX). 26. Quehenberger O., Armando A., Dumlao D., Stephens D. L., Dennis E. A. 2008. Lipidomics analysis of essential fatty acids in macrophages. Prostaglandins Leukot. Essent. Fatty Acids. 79: 123-129. 27. Quehenberger O., Armando A. M., Dennis E. A. 2011. High sensitivity quantitative lipidomics analysis of fatty acids in biological samples by gas chromatography-mass spectrometry. Biochim. Biophys. Acta. 1811: 648-656.
Instrument Name:Multiple
Column Name:Multiple
Chromatography Type:Core G
  
Chromatography ID:CH001056
Chromatography Summary:Cardiolipin analysis was achieved with normal phase LC coupled with high-resolution MS performed on a TripleTOF 5600 system (AB SCIEX, Foster City, CA) (24). Dolichol and coenzyme Q were analyzed by reverse phase LC coupled with multiple reaction monitoring (MRM) MS utilizing a 4000 Q-Trap hybrid triple quadrupole linear ion-trap mass spectrometer (AB SCIEX) (22, 23). For dolichol analysis, MRM was performed in negative ion mode, with the precursor ions being the (M+acetate)- adduct ions and the product ions being the acetate ions (m/z 59). For coenzyme Q analysis, MRM was carried out in positive ion mode, with ammonium adducts (M+NH4)+ as precursor ions and the proton adducts of the quinone ring of coenzyme Q (m/z 197) as product ions. For quantitation, an internal standard mixture composed of a cardiolipin mix (Avanti Polar Lipids, Inc.), nor-dolichol (13-22) (Avanti Polar Lipids, Inc.), and yeast coenzyme Q6 (Sigma) was added during the first step of lipid extraction (22). 22. Quehenberger O., Armando A. M., Brown H. A., Milne S. B., Myers D. S., Merrill A. H., Jr, Bandyopadhyay S., Jones K. N., Kelly S., Shaner R. L., et al. 2010. Lipidomics reveals a remarkable diversity of lipids in human plasma. J. Lipid Res. 51: 3299-3305. 23. Guan Z., Li S., Smith D., Shaw W., Raetz C. 2007. Identification of N-acylphosphatidylserine molecules in eukaryotic cells. Biochemistry. 46: 14500-14513. 24. Tan B. K., Bogdanov M., Zhao J., Dowhan W., Raetz C. R., Guan Z. 2012. Discovery of cardiolipin synthase utilizing phosphatidylethanolamine and phosphatidylglycerol as substrates. Proc. Natl. Acad. Sci. USA. 109: 16504-16509.
Instrument Name:Multiple
Column Name:Multiple
Chromatography Type:Core K
  
Chromatography ID:CH001057
Chromatography Summary:The organic solvent extraction layer containing CEs, TAGs, and DAGs was taken to dryness, then derivatized with 2,5-difluorophenylisocyanate to convert DAGs to urethane derivatives (35). The derivatized extract was separated by normal phase LC as previously described (35). The CEs eluting first from the LC column were detected by 20 specific MRM transitions corresponding to each [M+NH4]+ ion being collisionally activated to m/z 369.3. During elution of TAGs, the mass spectrometer was set to carry out full mass scanning from m/z 400-1,000. The DAGs were detected by neutral loss scanning of 190 Da. 35. Leiker T. J., Barkley R. M., Murphy R. C. 2011. Analysis of diacylglycerol molecular species in cellular lipid extracts by normal-phase LC-electrospray mass spectrometry. Int. J. Mass Spectrom. 305: 103-109
Instrument Name:Multiple
Column Name:Multiple
Chromatography Type:Core E
  
Chromatography ID:CH001058
Chromatography Summary:Sphingolipids were analyzed by LC-MS/MS essentially as described in (32, 33) with minor modifications to include the 1-deoxy-sphingolipids as generally described in (36). 32. Shaner R. L., Allegood J. C., Park H., Wang E., Kelly S., Haynes C. A., Sullards M. C., Merrill A. H., Jr 2009. Quantitative analysis of sphingolipids for lipidomics using triple quadrupole and quadrupole linear ion trap mass spectrometers. J. Lipid Res. 50: 1692-1707. 33. Sullards M. C., Liu Y., Chen Y., Merrill A. H., Jr 2011. Analysis of mammalian sphingolipids by liquid chromatography tandem mass spectrometry (LC-MS/MS) and tissue imaging mass spectrometry (TIMS). Biochim. Biophys. Acta. 1811: 838-853. 36. Zitomer N. C., Mitchell T., Voss K. A., Bondy G. S., Pruett S. T., Garnier-Amblard E. C., Liebeskind L. S., Park H., Wang E., Sullards M. C., et al. 2009. Ceramide synthase inhibition by fumonisin B1 causes accumulation of 1-deoxysphinganine: a novel category of bioactive 1-deoxysphingoid bases and 1-deoxydihydroceramides biosynthesized by mammalian cell lines and animals. J. Biol. Chem. 284: 4786-4795.
Instrument Name:Multiple
Column Name:Multiple
Chromatography Type:Core I

MS:

MS ID:MS001380
Analysis ID:AN001497
Instrument Type:-
MS Type:Other
Ion Mode:UNSPECIFIED
  
MS ID:MS001382
Analysis ID:AN001499
Instrument Type:-
MS Type:Other
Ion Mode:UNSPECIFIED
  
MS ID:MS001383
Analysis ID:AN001500
Instrument Type:-
MS Type:Other
Ion Mode:UNSPECIFIED
  
MS ID:MS001384
Analysis ID:AN001501
Instrument Type:-
MS Type:Other
Ion Mode:UNSPECIFIED
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