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  • The International Journal of Neuropsychopharmacology, Volume 14, Issue 7
  • August 2011, pp. 899-911

Plasma proteomic alterations in non-human primates and humans after chronic alcohol self-administration

  • Willard M. Freeman (a1), Heather D. VanGuilder (a1), Elizabeth Guidone (a2), John H. Krystal (a2) (a3), Kathleen A. Grant (a4) and Kent E. Vrana (a1)
  • DOI:
  • Published online: 08 February 2011

Objective diagnostics of excessive alcohol use are valuable tools in the identification and monitoring of subjects with alcohol use disorders. A number of potential biomarkers of alcohol intake have been proposed, but none have reached widespread clinical usage, often due to limited diagnostic sensitivity and specificity. In order to identify novel potential biomarkers, we performed proteomic biomarker target discovery in plasma samples from non-human primates that chronically self-administer high levels of ethanol. Two-dimensional difference in-gel electrophoresis (2D-DIGE) was used to quantify plasma proteins from within-subject samples collected before exposure to ethanol and after 3 months of excessive ethanol self-administration. Highly abundant plasma proteins were depleted from plasma samples to increase proteomic coverage. Altered plasma levels of serum amyloid A4 (SAA4), retinol-binding protein, inter-alpha inhibitor H4, clusterin, and fibronectin, identified by 2D-DIGE analysis, were confirmed in unmanipulated, whole plasma from these animals by immunoblotting. Examination of these target plasma proteins in human subjects with excessive alcohol consumption (and control subjects) revealed increased levels of SAA4 and clusterin and decreased levels of fibronectin compared to controls. These proteins not only serve as targets for further development as biomarker candidates or components of biomarker panels, but also add to the growing understanding of dysregulated immune function and lipoprotein metabolism with chronic, excessive alcohol consumption.

Corresponding author
Address for correspondence: K. E. Vrana, Ph.D., Department of Pharmacology, Penn State College of Medicine, R130, 500 University Drive, Hershey, PA 17033-0850, USA. Tel.: 717-531-8285Fax: 717-531-0419Email:
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D Alte , J Ludemann , M Piek , C Adam , (2003). Distribution and dose response of laboratory markers to alcohol consumption in a general population: results of the study of health in Pomerania (SHIP). Journal of Studies on Alcohol and Drugs 64, 7582.

NL Anderson , NG Anderson (2002). The human plasma proteome: history, character, and diagnostic prospects. Molecular & Cellular Proteomics 1, 845867.

RF Anton , C Lieber , B Tabakoff (2002). Carbohydrate-deficient transferrin and gamma-glutamyltransferase for the detection and monitoring of alcohol use: results from a multisite study. Alcoholism: Clinical and Experimental Research 26, 12151222.

LL Bausserman , PN Herbert , KP McAdam (1980). Heterogeneity of human serum amyloid A proteins. Journal of Experimental Medicine 152, 641656.

KA Bradley , AF DeBenedetti , RJ Volk , EC Williams , (2007). AUDIT-C as a brief screen for alcohol misuse in primary care. Alcoholism: Clinical and Experimental Research 31, 12081217.

ND Brener , JO Billy , WR Grady (2003). Assessment of factors affecting the validity of self-reported health-risk behaviour among adolescents: evidence from the scientific literature. Journal of Adolescent Health 33, 436457.

K Bush , DR Kivlahan , MB McDonell , SD Fihn , (1998). The AUDIT alcohol consumption questions (AUDIT-C): an effective brief screening test for problem drinking. Ambulatory Care Quality Improvement Project (ACQUIP). Alcohol Use Disorders Identification Test. Archives of Internal Medicine 158, 17891795.

KM Conigrave , P Davies , P Haber , JB Whitfield (2003). Traditional markers of excessive alcohol use. Addiction 98 (Suppl. 2), 3143.

KM Conigrave , LJ Degenhardt , JB Whitfield , JB Saunders , (2002). CDT, GGT, and AST as markers of alcohol use: the WHO/ISBRA collaborative project. Alcoholism: Clinical and Experimental Research 26, 332339.

FT Crews , R Bechara , LA Brown , DM Guidot , (2006). Cytokines and alcohol. Alcoholism: Clinical and Experimental Research 30, 720730.

SK Das , L Dhanya , DM Vasudevan (2008). Biomarkers of alcoholism: an updated review. Scandinavian Journal of Clinical & Laboratory Investigation 68, 8192.

JA Ewing (1984). Detecting alcoholism. The CAGE questionnaire. Journal of the American Medical Association 252, 19051907.

D Falk (1961). Production of polydipsia in normal rats by an intermittent food schedule. Science 133, 195196.

D Frank , AF DeBenedetti , RJ Volk , EC Williams , (2008). Effectiveness of the AUDIT-C as a screening test for alcohol misuse in three race/ethnic groups. Journal of General Internal Medicine 23, 781787.

WM Freeman , RS Gooch , ME Lull , TJ Worst , (2006 a). Apo-aii is an elevated biomarker of chronic non-human primate ethanol self-administration. Alcohol and Alcoholism 41, 300305.

WM Freeman , ME Lull , MT Guilford , KE Vrana (2006 b). Depletion of abundant proteins from non-human primate serum for biomarker studies. Proteomics 6, 31093113.

WM Freeman , AC Salzberg , SW Gonzales , KA Grant , (2010). Classification of alcohol abuse by plasma protein biomarkers. Biological Psychiatry 68, 219222.

WM Freeman , KE Vrana (2010). Future prospects for biomarkers of alcohol consumption and alcohol-induced disorders. Alcoholism: Clinical and Experimental Research 34, 946954.

P Ghosh , EA Hale , MR Lakshman (2001). Plasma sialic-acid index of apolipoprotein J (SIJ): a new alcohol intake marker. Alcohol 25, 173179.

K Golka , A Wiese (2004). Carbohydrate-deficient transferrin (CDT) – a biomarker for long-term alcohol consumption. Journal of Toxicology and Environmental Health Part B: Critical Reviews 7, 319337.

KA Grant , AJ Bennett (2003). Advances in nonhuman primate alcohol abuse and alcoholism research. Pharmacology & Therapeutics 100, 235255.

KA Grant , X Leng , HL Green , KT Szeliga , (2008 a). Drinking typography established by scheduled induction predicts chronic heavy drinking in a monkey model of ethanol self-administration. Alcoholism: Clinical and Experimental Research 32, 18241838.

KL Green , KT Szeliga , CA Bowen , MA Kautz , (1999). Comparison of ethanol metabolism in male and female cynomolgus macaques (Macaca fascicularis). Alcoholism: Clinical and Experimental Research 23, 611616.

ML Hannuksela , MK Liisanantti , AE Nissinen , MJ Savolainen (2007). Biochemical markers of alcoholism. Clinical Chemistry and Laboratory Medicine 45, 953961.

H Harwood (2000). Updating Estimates of the Economic Costs of Alcohol Abuse in the United States: Estimates, Update Methods, and Data. Rockville, MD: National Institute on Alcohol Abuse and Alcoholism.

A Helander , CJ Eriksson (2002). Laboratory tests for acute alcohol consumption: results of the WHO/ISBRA Study on State and Trait Markers of Alcohol Use and Dependence. Alcoholism: Clinical and Experimental Research 26, 10701077.

B Hock , M Schwarz , I Domke , VP Grunert , (2005). Validity of carbohydrate-deficient transferrin (%CDT), gamma-glutamyltransferase (gamma-GT) and mean corpuscular erythrocyte volume (MCV) as biomarkers for chronic alcohol abuse: a study in patients with alcohol dependence and liver disorders of non-alcoholic and alcoholic origin. Addiction 100, 14771486.

AM Hoyumpa (1986). Mechanisms of vitamin deficiencies in alcoholism. Alcoholism: Clinical and Experimental Research 10, 573581.

P Ivester , LJ Roberts , T Young , D Stafforini , (2007). Ethanol self-administration and alterations in the livers of the cynomolgus monkey, Macaca fascicularis. Alcoholism: Clinical and Experimental Research 31, 144155.

J Junge , KD Bentsen , P Christoffersen , M Orholm , (1988). Fibronectin as predictor of cirrhosis in men who abuse alcohol. British Medical Journal (Clinical Research Edition) 296, 16291630.

MJ Kip , CD Spies , T Neumann , Y Nachbar , (2008). The usefulness of direct ethanol metabolites in assessing alcohol intake in nonintoxicated male patients in an emergency room setting. Alcoholism: Clinical and Experimental Research 32, 12841291.

JC Kissack , J Bishop , AL Roper (2008). Ethylglucuronide as a biomarker for ethanol detection. Pharmacotherapy 28, 769781.

H Koch , GJ Meerkerk , JO Zaat , MF Ham , (2004). Accuracy of carbohydrate-deficient transferrin in the detection of excessive alcohol consumption: a systematic review. Alcohol and Alcoholism 39, 7585.

A Korzec , C de Bruijn , M van Lambalgen (2005). The Bayesian alcoholism test had better diagnostic properties for confirming diagnosis of hazardous and harmful alcohol use. Journal of Clinical Epidemiology 58, 10241032.

S Korzec , A Korzec , K Conigrave , J Gisolf , (2009). Validation of the Bayesian alcoholism test compared to single biomarkers in detecting harmful drinking. Alcohol and Alcoholism 44, 398402.

O Niemela (2007). Biomarkers in alcoholism. Clinica Chimica Acta 377, 3949.

M Pineiro , MA Alava , N Gonzalez-Ramon , J Osada , (1999). ITIH4 serum concentration increases during acute-phase processes in human patients and is up-regulated by interleukin-6 in hepatocarcinoma HepG2 cells. Biochemical and Biophysical Research Communications 263, 224229.

BS Pruett , SB Pruett (2006). An explanation for the paradoxical induction and suppression of an acute phase response by ethanol. Alcohol 39, 105110.

LC Sobell , MB Sobell (1992). Timeline follow-back: a technique for assessing self-reported alcohol consumption. In: R Litten , J Allen (Eds), Measuring Alcohol Consumption (pp. 4172). New York: Humana Press.

RL Spitzer , JBW Williams (2002). Structured Clinical Interview for the DSM-IV Axis I Disorders-Patient Edition (SCID-I/P, Version 2.0). New York: Biometrics Research Department, New York State Psychiatric Institute.

DL Steinweg , H Worth (1993). Alcoholism: the keys to the CAGE. American Journal of Medicine 94, 520523.

E Taracha , B Habrat , P Wozniak , J Walkowiak , (2001). The activity of beta-hexosaminidase (uHex) and gamma-glutamyl-transferase (uGGT) in urine as non-invasive markers of chronic alcohol abuse: I. Alcohol-dependent subjects. World Journal of Biological Psychiatry 2, 184189.

TM Umstead , CJ Lu , WM Freeman , JL Myers , (2010). Dual-platform proteomics study of plasma biomarkers in pediatric patients undergoing cardiopulmonary bypass. Pediatric Research 67, 641649.

HD VanGuilder , GV Bixler , L Kutzler , RM Brucklacher , (2011). Multi-modal proteomic analysis of retinal protein expression alterations in a rat model of diabetic retinopathy. PLoS ONE 6, e16271.

G Vendemiale , I Grattagliano , E Altomare , G Serviddio , (2001). Mitochondrial oxidative damage and myocardial fibrosis in rats chronically intoxicated with moderate doses of ethanol. Toxicology Letters 123, 209216.

JA Vivian , HL Green , JE Young , LS Majerksy , (2001). Induction and maintenance of ethanol self-administration in cynomolgus monkeys (Macaca fascicularis): long-term characterization of sex and individual differences. Alcoholism: Clinical and Experimental Research 25, 10871097.

Q Yang , TE Graham , N Mody , F Preitner , (2005). Serum retinol binding protein 4 contributes to insulin resistance in obesity and type 2 diabetes. Nature 436, 356362.

N Zolotarjova , P Mrozinski , H Chen , J Martosella (2008). Combination of affinity depletion of abundant proteins and reversed-phase fractionation in proteomic analysis of human plasma/serum. Journal of Chromatography A 1189, 332338.

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