Plasma concentrations of molecular lipid species predict long-term clinical outcome in coronary artery disease patients

• DOI: 10.1194/jlr.p081281
• Publication Date: 2018-06-01T20:40:11Z
• Authors: Sharda Anroedh; Mika Hilvo; K. Martijn Akkerhuis; Dimple Kauhanen; Kaisa Koistinen; Rohit Oemrawsingh; Patrick Serruys; Robert-Jan van Geuns; Eric Boersma; Reijo Laaksonen; Isabella Kardys
• Keywords: Male; 0301 basic medicine; ceramides; 610; vascular biology; heart; QD415-436; Coronary Artery Disease; Middle Aged; Prognosis; Biochemistry; Lipids; 3. Good health; 03 medical and health sciences; All institutes and research themes of the Radboud University Medical Center; Radboudumc 16: Vascular damage RIHS: Radboud Institute for Health Sciences; follow-up; lipidomics; Humans; Female; atherosclerosis; EMC COEUR-09; Follow-Up Studies

We investigated the associations of ten previously identified high risk molecular lipid species and three ceramide ratios with occurrence major adverse cardiac events (MACEs) during a median follow-up 4.7 years in patients coronary artery disease (CAD). Between 2008 2011, 581 underwent diagnostic angiography or percutaneous intervention for stable angina pectoris (SAP) acute syndrome (ACS). Blood was drawn prior to index procedure were determined. The primary endpoint MACE, comprising all-cause mortality, nonfatal ACS, unplanned revascularization. secondary comprised mortality ACS. During [IQR: 4.2–5.6] years, 155 (27%) had MACEs. In multivariable analyses, Cer(d18:1/16:0) concentration associated MACEs {hazard ratio 2.32; 95% CI [1.09–4.96] per natural logarithm (ln) (pmol/ml) P = 0.030} after adjustment factors, clinical presentation, statin use at baseline, admission nonHDL cholesterol level. Furthermore, adjustment, concentrations Cer(d18:1/16:0), Cer(d18:1/20:0), Cer(d18:1/24:1), their Cer(d18:1/24:0) composite death data together show circulating lipids we here are outcome long-term independent factors. Established markers such as total cholesterol, LDL triglycerides (TGs), HDL have long formed cornerstone lipid-based stratification (CAD) (1.Tarasov K. Ekroos Suoniemi M. Kauhanen D. Sylvanne T. Hurme R. Gouni-Berthold I. Berthold H.K. Kleber M.E. Laaksonen et al.Molecular identify cardiovascular efficiently lowered by simvastatin PCSK9 deficiency.J. Clin. Endocrinol. Metab. 2014; 99: E45-E52Crossref PubMed Scopus (149) Google Scholar, 2.Alshehry Z.H. Mundra P.A. Barlow C.K. Mellett N.A. Wong G. McConville M.J. Simes J. Tonkin A.M. Sullivan D.R. Barnes E.H. al.Plasma lipidomic profiles improve on traditional factors prediction type 2 diabetes mellitus.Circulation. 2016; 134: 1637-1650Crossref (153) 3.Havulinna A.S. Sysi-Aho Hilvo Salomaa V. Circulating ceramides predict outcomes population-based FINRISK 2002 cohort.Arterioscler. Thromb. Vasc. Biol. 36: 2424-2430Crossref (182) 4.Ekroos Janis Tarasov Lipidomics: tool studies atherosclerosis.Curr. Atheroscler. Rep. 2010; 12: 273-281Crossref (82) Scholar). However these measures alone do not fully capture complexity altered metabolism (2.Alshehry Scholar), this may be reason that they fail substantial proportion Lipidomics is systems-based study all (5.Watson A.D. Thematic review series: systems biology approaches metabolic disorders. global approach analysis biological systems.J. Lipid Res. 2006; 47: 2101-2111Abstract Full Text PDF (366) Scholar) has been defined full characterization roles (6.Roberts L.D. McCombie Titman C.M. Griffin J.L. A matter fat: an introduction profiling methods.J. Chromatogr. B Analyt. Technol. Biomed. Life Sci. 2008; 871: 174-181Crossref (102) its most advanced form, lipidomics able quantify hundreds diverse across multiple classes sphingolipids, phospholipids, sterol esters, acylglycerols (7.Jänis M.T. Oresic Metabolomic strategies tissue-specific effects drugs.Expert Opin. Drug Toxicol. 4: 665-680Crossref (13) many which play integral role modulation function formation cellular membranes, energy storage, cell signaling (8.Stock emerging lipidomics.Atherosclerosis. 2012; 221: 38-40Abstract (8) 9.Hu C. van der Heijden Wang Greef Hankemeier Xu Analytical applications biomarker discovery.J. 2009; 877: 2836-2846Crossref (168) Because provides detailed profiles, it further CAD provide novel mechanistic insights into (4.Ekroos line hypothesis, recently performed Ludwigshafen Risk Cardiovascular Health (LURIC) several fatal current study, hypothesized follow-up. design European Collaborative Project Inflammation Vascular Wall Remodeling Atherosclerosis (ATHEROREMO) described elsewhere detail (10.de Boer S.P. Cheng J.M. Garcia-Garcia H.M. Oemrawsingh R.M. Geuns R.J. Regar E. Zijlstra F. Halperin al.Relation genetic profile biomarkers plaque phenotype determined intravascular ultrasound: rationale ATHEROREMO-IVUS study.EuroIntervention. 10: 953-960Crossref (23) brief, from until indication (CAG) and/or (PCI) due (ACS) Erasmus MC, Rotterdam, Netherlands, included. Prior CAG PCI procedure, blood samples collected arterial sheath transported laboratory MC within h collection storage −80°C. All included 18 older. ATHEROREMO approved medical ethics committee accordance criteria Declaration Helsinki. Written informed consent obtained patients. Levels TGs measured serum using Roche/Hitachi cobas c 701/702 analyzer (Roche Diagnostics, Indianapolis, IN) Cobas 8000 modular platform Diagnostics). Molecular found < 0.05 level LURIC selected evaluation These cholesteryl esters (CEs): CE 14:0, 18:3, 20:4, 20:5, 22:5; (Cer): Cer(d18:1/24:0), Cer(d18:1/24:1); ratios: Cer(d18:1/16:0)/Cer(d18:1/24:0), Cer(d18:1/20:0)/Cer(d18:1/24:0), Cer(d18:1/24:1)/Cer(d18:1/24:0)), lactosylceramide (LacCer): LacCer(d18:1/18:0). Plasma measurement available 574 Stored plasma subjected extraction Zora Biosciences, Finland. Briefly, (10 μl) spiked known amounts lipid-class specific, nonendogenous synthetic internal standards, D6-CE 18:0 (C/D/N Isotopes Inc.,Pointe-Claire, Quebec, Canada), Cer(d18:1/17:0) (Avanti Polar Lipids Inc., Alabaster, AL) D3-LacCer(d18:1/16:0) (Matreya LLC, State College, PA). chloroform (HPLC grade) (Rathburn Chemicals Ltd., Walkerburn, Scotland), methanol, acetic acid (both LC-MS (Sigma-Aldrich GmbH, Steinheim, Germany) (11.Heiskanen L.A. Ta H.X. Long-term performance stability shotgun human samples.Anal. Chem. 2013; 85: 8757-8763Crossref (61) After extraction, reconstituted chloroform-methanol (1:2, v/v) sphingolipids analysis, extracts diluted containing 5 mM ammonium acetate. Quality control prepared along actual analyses monitor MS performance. intra-day (n 3) average coefficient variation CEs less than equal 6% inter-day [n 24 Cer LacCer; n 23 except CE(22:5) 22] 21% both CE. Sphingolipids analyzed QTRAP® 5500 mass spectrometer (AB SCIEX, Concord, Canada) equipped ultra-high pressure liquid chromatography (UHPLC) system CTC PAL autosampler (Leap Technologies) Accela 1250 Pump (Thermo Fisher Scientific, Agawam, MA). Chromatographic separation Acquity BEH C18, 2.1 × 50 mm column particle size 1.7 μm (Waters, Milford, Mobile phases 10 acetate water 0.1% formic (solvent A) acetonitrile-isopropanol (4:3, B). separated linear gradient 75% 100% 15 min. Flow rate 500 µl/min temperature 60°C. Data reaction monitoring positive ion mode (12.Merrill Jr., A.H. Sullards M.C. Allegood J.C. Kelly S. Sphingolipidomics: high-throughput, structure-specific, quantitative tandem spectrometry.Methods. 2005; 207-224Crossref (468) Curtain gas set 25, spray voltage 5000, source heated 400°C. Collision optimized each class. LacCer 40 45, respectively. Shotgun SCIEX) robotic nanoflow NanoMate HD (Advion, Ithaca, NY) precursor scanning 369.35 collision 30 (13.Ejsing C.S. Duchoslav Sampaio Simons Bonner Thiele Shevchenko A. Automated identification quantification glycerophospholipid scanning.Anal. 78: 6202-6214Crossref (329) Mass spectrometry files processed MultiQuant™ 2.0.1 LipidView™ 1.0 Identified quantified normalizing against respective standard volume used extraction. limit (LOQ) Cer, LacCer, extract 0.0004 µM, 0.0016 0.012 monitored LOQ. LOQ lowest point calibration curve signal-to-noise greater 10. Clinical vital status charts, civil registries, written telephone contacts relatives. living participating received questionnaire consisting queries regarding readmissions. For events, hospital discharge letters treating physicians institutions contacted if necessary additional information. MACE ACS diagnosis ST-segment elevation myocardial infarction (STEMI), nonSTEMI, unstable guidelines Society Cardiology (14.Roffi Patrono Collet J.P. Mueller Valgimigli Andreotti Bax J.J. Borger M.A. Brotons Chew D.P. al.ESC Guidelines management syndromes presenting without persistent elevation: Task Force Management Acute Coronary Syndromes Patients Presenting Persistent ST-Segment Elevation (ESC).Eur. Heart 37: 267-315Crossref (4341) 15.Ibanez B. James Agewall Antunes Bucciarelli-Ducci Bueno H. Caforio A.L.P. Crea Goudevenos J.A. Halvorsen task force 2018; 39: 119-177Crossref (5409) Unplanned revascularization repeated bypass grafting (CABG). endpoints adjudicated according definitions blinded data. Categorical variables presented numbers percentages. distributions continuous variables, including ratios, examined normality visual inspection histogram. Normally distributed mean ± SD. Nonnormally (which ratios) (interquartile range [IQR]) analyses. lost considered date last contact, time-point censored. Cox proportional hazards models evaluate between endpoints. who experienced more one event, first considered. results hazard (HRs) unit increase (ln-transformed) CIs. First, univariably. gender, age, hypertension, hypercholesterolemia, mellitus, potential confounders entered covariates. covariates chosen etiologic reasons based existing literature (16.Stone G.W. Maehara Lansky A.J. de Bruyne Cristea Mintz G.S. Mehran McPherson Farhat N. Marso al.A prospective natural-history atherosclerosis.N. Engl. Med. 2011; 364: 226-235Crossref (2291) To whether levels levels, baseline additionally (and consecutively) added models. Serum calculated subtracting cohort, (ACS versus SAP) also covariate. Interaction terms model account possible effect modification CAG. Subsequently, stratified SPSS software (SPSS 23.0 IBM Corp., Armonk, NY). statistical tests two-tailed P-values statistically significant. characteristics summarized Table 1 2. total, age 61.5 men. 55% diagnosed (28% STEMI 26% nonSTEMI) 46% SAP. 88% procedure. 2.71 2.12–3.54] mmol/l, 1.04 0.87–1.29] 3.23 2.54–4.00] TG 1.27 0.88–1.83] mmol/l cohort. significantly higher {(median: 3.10 2.32–3.87] mmol/l) 0.001}, 3.56 [2.81–4.36]mmol/l) 0.001} lower {(median 1.15 0.77–1.77] 0.001)} compared SAP (median: 2.37 1.94–2.99] 2.83 [2.35–3.56] 1.41 1.05–1.94] respectively). addition, other different (Table 1). At time admission, 89% cohort statins.TABLE 1Clinical characteristicsClinical characteristicsTotal 574)ACS 313)SAP 261)PAge, SD61.5 11.359.7 11.963.6 10.3<0.001Male, (%)432 (75)230 (74)202 (77)0.279Diabetes (%)97 (17)40 (13)57 (22)0.004Hypertension, (%)298 (52)137 (44)161 (62)<0.001Hypercholesterolemia, (%)318 (55)138 (44)180 (69)<0.001Smoking, (%)166 (29)116 (37)50 (19)<0.001Positive family history CAD, (52)145 (46)153 (59)0.004Previous MI, (%)184 (32)80 (26)104 (40)<0.001Previous PCI, (32)57 (18)127 (49)<0.001Previous CABG, (%)18 (3)7 (2)11 (4)0.176Previous stroke, (%)26 (5)11 (4)15 (6)0.200Peripheral disease, (%)35 (6)11 (4)24 (9)0.005History heart failure, (%)19 (3)6 (2)13 (5)0.041Serum mmol/L2.71 [2.12–3.54]3.10 [2.32–3.87]2.37 [1.94–2.99]<0.001Serum mmol/L1.04 [0.87–1.29]1.05 [0.87–1.27]1.03 [0.86–1.30]0.80Serum mmol/L3.23 [2.54–4.00]3.56 [2.81–4.36]2.83 [2.35–3.56]<0.001Serum TG, mmol/L1.27 [0.88-1.83]1.15 [0.77–1.77]1.41 [1.05–1.94]<0.001Statin (%)508 (89%)308 (98%)235 (90%)0.499Procedural characteristicsIndication CAGACS, (%)313 (55)313 (100)0 (0)STEMI, (%)162 (28)162 (52)0 (0)Non-ST-elevation, (%)151 (26)151 (48)0 (0)Stable pectoris, (%)261 (46)0 (0)261 (100)PCI performed, (%)505 (88)291 (93)214 (82)aA significant stenosis ≥ 50% vessel diameter assessment angiogram.No stenosis, (%)42 (7)18 (6)24 (9)1-vessel (%)304 (53)172 (55)132 (51)2-vessel (%)167 (29)88 (28)79 (30)3-vessel (%)61 (11)35 (11)26 (10)Continuous (SD) [IQR]. (n) percentages (%). P-value Student's t-test Chi square test. syndrome; grafting; disease; CAG, angiography; IQR, interquartile range; infarction; intervention; SAP, pectoris; triglyceride.a angiogram. Open table new tab TABLE 2Lipid follow-up, follow-upLipid cohortTotal 261)PCE pmol/µl21.7 [15.9–28.1]22.9 [16.5–30.5]21.2 [15.4–26.8]0.008CE pmol/µl70.3 [51.8–90.7]72.3 [53.6–99.5]66.1 [50.3–85.2]0.003CE pmol/µl386 [317–457]394 [324–453]374 [307–471]0.31CE pmol/µl49.1 [36.3–72.6]49.2 [36.4–72.1]49.0 [35.9–74.7]0.69CE 22:5, pmol/µl2.65 [2.00–3.62]2.81 [2.12–3.77]2.53 [1.90–3.40]0.037Cer(d18:1/16:0) pmol/µl0.12 [0.10–0.15]0.13 [0.11–0.17]0.11 [0.09–0.13]<0.001Cer(d18:1/20:0) pmol/µl0.11 [0.09–0.15]0.12 [0.10–0.16]0.11 [0.08–0.13]<0.001Cer(d18:1/24:0) pmol/µl5.98 [4.72–7.49]6.43 [5.00–8.07]5.65 [4.49–6.61]<0.001Cer(d18:1/24:1) pmol/µl1.79 [1.42–2.25]1.89 [1.52–2.44]1.67 [1.35–2.05]<0.001LacCer(d18:1/18:0) pmol/µl0.13 [0.10–0.16]0.13 [0.11–0.16]0.12 [0.10–0.15]0.001Cer(d18:1/16:0)/Cer(d18:1/24:0) pmol/µl0.020 [0.018–0.024]0.021 [0.018–0.025]0.020 [0.017–0.023]0.001Cer(d18:1/20:0)/Cer(d18:1/24:0) pmol/µl0.019 [0.016–0.024]0.019 [0.015–0.024]0.019 [0.016–0.023]0.62Cer(d18:1/24:1)/Cer(d18:1/24:0) pmol/µl0.31 [0.26–0.36]0.31 [0.26–0.36]0.65Lipid those MACEsaInformation 566.Total 155)ACS 65)SAP 90)PCE pmol/µl22.6 [15.7–27.1]21.7 [15.5–30.3]22.7 [15.7–26.6]0.67CE pmol/µl67.9 [51.2–90.3]70.3 [52.8–103]66.9 [50–84.1]0.17CE pmol/µl381 [310–445]381 [310–432]379 [310–447]0.95CE pmol/µl52.4 [37.4–74.5]52.6 [38.4–76.3]50.9 [36.6–74.4]0.73CE pmol/µl2.57 [1.86–3.71]2.61 [2.04–3.97]2.51 [1.80–3.45]0.065Cer(d18:1/16:0) [0.10–0.16]0.15 [0.09–0.16]0.13 [0.10–0.17]0.11 [0.08–0.14]0.011Cer(d18:1/24:0) pmol/µl5.86 [4.65–7.48]6.44 [5.02–8.10]5.66 [4.54–6.58]0.063Cer(d18:1/24:1) pmol/µl1.78 [1.35–2.36]2.10 [1.66–2.84]1.62 [1.33–2.13]0.008LacCer(d18:1/18:0) [0.10–0.16]0.14 [0.10–0.18]0.13 [0.10–0.16]0.137Cer(d18:1/16:0)/Cer(d18:1/24:0) pmol/µl0.021 [0.018–0.025]0.022 [0.019–0.027]0.019 [0.017–0.024]0.006Cer(d18:1/20:0)/Cer(d18:1/24:0) [0.016–0.025]0.020 [0.016–0.024]0.020 [0.015–0.025]0.504Cer(d18:1/24:1)/Cer(d18:1/24:0) [0.27–0.37]0.33 [0.27–0.36]0.31 [0.26–0.37]0.222Lipid 411)ACS 242)SAP 169)PCE pmol/µl21.5 [15.8–28.7]23 [16.5–30.5]20.7 [15.3–27]0.007CE pmol/µl70.5 [51.7–91.3]73.8 [53.5–99]66 [50.3–86.4]0.011CE pmol/µl391 [321–467]397 [310–432]374 [304–476]0.272CE [35.6–70.5]49.2 [35.5–70.7]47.5 [35.6–70]0.575CE pmol/µl2.69 [2.04–3.58]2.79 [2.12–3.70]2.54 [1.92–3.40]0.041Cer(d18:1/16:0) [0.11–0.16]0.11 [0.09–0.14]0.12 [0.09–0.15]0.11 pmol/µl6 [4.75–7.47]6.39 [4.97–8.07]5.64 [4.49–6.64]<0.001Cer(d18:1/24:1) [1.35–2.36]1.86 [1.51–2.33]1.68 [1.35–2.04]<0.001LacCer(d18:1/18:0) [0.018–0.025]0.021 [0.018–0.024]0.020 [0.017–0.023]0.017Cer(d18:1/20:0)/Cer(d18:1/24:0) [0.016–0.025]0.019 [0.016–0.023]0.60Cer(d18:1/24:1)/Cer(d18:1/24:0) [0.27–0.37]0.30 [0.25–0.36]0.31 [0.26–0.36]0.77Concentrations μM difference ln-transformed concentration. CE, ester; ceramide; lactosylceramide; event; pectoris.a Information 566. Continuous triglyceride. Concentrations pectoris. As shown 2, LacCer(d18:1/18:0) Cer(d18:1/16:0)/Cer(d18:1/24:0) patients, remained free LacCer(d18:1/18:0), above-mentioned tended (P 0.054) 574), acquired 572 (99.7%). assessing completed 99% (IQR: [4.2–5.6]) least (primary endpoint). group, 65 (21%) follow-up; 90 (34%). depicted Fig. supplemental S1a. {HR: 2.14; [1.22–3.76] ln(pmol/ml) 0.008} Cer(d18:1/24:1) 1.64; [1.00–2.68] 0.049} associ