white adipose tissue ( wat ) represents an active endocrine organ that regulates body fat mass and energy balance . there is increasing evidence that wat - derived adipokines may contribute to hepatic damage associated with fatty infiltration , inflammation , and fibrosis [ 1 , 2 ] . adiponectin is secreted exclusively from adipose tissue and circulates in different isoforms : trimers , of low molecular weight ( lmw ) , hexamers ( trimer dimer ) of medium molecular weight ( mmw ) , and multimeric high molecular weight ( hmw ) isoforms . leptin is expressed mainly by adipose tissue , although low levels have been detected in the placenta , skeletal muscle , gastric and mammary epithelium , and the brain . although resistin is secreted by human adipocytes , the most significant source appears to be blood mononuclear cells . plasma concentrations of adipokines have been investigated in patients with different liver disorders , that is , nonalcoholic fatty liver disease [ 46 ] , type 1 autoimmune hepatitis , and viral hepatitis b   and c . alcoholic liver disease ( ald ) is another one where adipokines may play pivotal role and represent a link between inflammation and metabolic state . therefore , the objective of the present study was to determine serum concentrations of total adiponectin ( acrp30 ) , leptin , and resistin in patients with chronic alcohol abuse and different grades of liver dysfunction . we also investigated the adipokine correlation with traditional indicators of inflammation , liver laboratory parameters , and ald complications . furthermore , on the basis of above obtained results , we tried to select noninvasive predictors of the disease severity and ald outcome . the prospective study was conducted in the department of gastroenterology of medical university in lublin , poland . one hundred forty - seven consecutive adult inpatients ( pts ) with ald admitted to hospital due to the disease decompensation were prospectively recruited . the control group consisted of 30 healthy volunteers matched to the study group in terms of age and sex and recruited mostly among academics and trainees of department of gastroenterology , medical university of lublin . the diagnosis of ald was based on clinical criteria such as a detailed patient history , typical symptoms and physical findings of chronic liver disease , laboratory values ( elevated serum aminotransferases activity ( normal range : alt < 31  iu / l ; ast < 34 iu / l ) , the ast / alt ratio higher than 2 ) , and imaging studies , in the setting of excessive alcohol intake ( i.e. , alcohol consumption exceeding 40  g / d for male and 20  g / d for female pts for a minimum of 6 months ) . in order to confirm alcohol misuse , the alcohol use disorders identification test - consumption ( audit - c ) an audit score of 8 for men up to age 60 , or 4 for women , or men over age 60 was considered a positive screening test . positive audit - c , in addition to the amount of alcohol consumption , was an inclusion criterion . based on the audit - c score , the alcohol consumption status was determined in both studied groups . two individuals from the control group , who pledged total abstinence , were categorized as abstainers and eighteen who self - reported their consumption as no more than 20  g ethanol per day ( 1016 drinks per month ; 1 drink refers to 10  g of pure alcohol ) as light drinkers . quantity of drinking among risky , current drinkers from the ald group ranged as follows : in females 40  g / d to more than 100 g / d and in males 50  g / d to more than 100  g / d .   according to the study protocol , patients signed informed consent , completed an anamnesis ( their medical history ) , and they answered the audit - c questionnaire . patients entering the study did not consume alcoholic beverages at least 24 hours prior to obtaining blood samples for laboratory tests . all data necessary for further analysis ( i.e. , the major demographic variables included age , gender , alcohol intake and the period of alcohol abuse , education , family history of alcoholism , employment status , treatment of ald before admission , and comorbidities ) were recorded , and the planned procedures including blood sampling were performed within 48 hours after hospital admission . blood samples from all patients were collected at 07:30  am after a minimum 8-hour overnight fast in order to avoid the circadian and feeding impact on serum adipokine fluctuations . basic laboratory tests included determination of liver function parameters ( alanine aminotransferase - alt , aspartate aminotransferase - ast , alkaline phosphatase - ap , gamma - glutamyl transpeptidase - ggt , total bilirubin - tbil , albumin , inr , and prothrombin time ) ; complete blood count ( hemoglobin - hgb , erythrocytes - rbc , platelets - plt , and leukocytes - wbc ) ; parameters of renal function ( creatinine - crea , serum sodium level - na ) ; the traditional markers of inflammation : neutrophils , neutrophil to lymphocytes count rate ( nlr ) and the level of c - reactive protein - crp ) ; indicators of other etiology of chronic liver disease , that is , hbv and hcv infection ( hbsag , anti - hbc class igm and igg antibodies ; anti - hcv antibodies ) , antinuclear , antismooth muscle , antimitochondrial antibodies , and markers of wilson 's disease and hemochromatosis as appropriate ) . liver function parameters ( alanine aminotransferase - alt , aspartate aminotransferase - ast , alkaline phosphatase - ap , gamma - glutamyl transpeptidase - ggt , total bilirubin - tbil , albumin , inr , and prothrombin time ) ;   complete blood count ( hemoglobin - hgb , erythrocytes - rbc , platelets - plt , and leukocytes - wbc ) ;   parameters of renal function ( creatinine - crea , serum sodium level - na ) ;   the traditional markers of inflammation : neutrophils , neutrophil to lymphocytes count rate ( nlr ) and the level of c - reactive protein - crp ) ;   indicators of other etiology of chronic liver disease , that is , hbv and hcv infection ( hbsag , anti - hbc class igm and igg antibodies ; anti - hcv antibodies ) , antinuclear , antismooth muscle , antimitochondrial antibodies , and markers of wilson 's disease and hemochromatosis as appropriate ) .   based on the lab results , the baseline severity of liver dysfunction was determined according to the child - turcotte - pugh ( ctp )   and the model of end - stage liver disease ( meld )   criteria . for the score calculation , patients enrolled into the study were divided into subgroups according to : gender;age : 50 and <  50 years old ; the severity of liver dysfunction according to the ctp ( classes a , b , c ) and meld ( 20 or <  20 ) scores ; the presence of ald complications at the time of hospital admission , that is , ascites , hepatic encephalopathy ( he ) , oesophageal varices , cholestasis , and renal dysfunction . age : 50 and <  50 years old ;   the severity of liver dysfunction according to the ctp ( classes a , b , c ) and meld ( 20 or <  20 ) scores ;   the presence of ald complications at the time of hospital admission , that is , ascites , hepatic encephalopathy ( he ) , oesophageal varices , cholestasis , and renal dysfunction . pts with any other severe diseases , that is , uncontrolled diabetes , heart failure , pulmonary insufficiency or malignancy at the time of inclusion were excluded . symptoms of overt hepatic encephalopathy ( he ) were classified according to west - heaven criteria .   cholestasis was defined based on the recommendations of the european association for the study of the liver ( easl ) , that is , alkaline phosphatase ( ap ) greater than 1.5 times above the upper limit of normal ( uln ) and the activity of -glutamyl transpeptidase ( ggt ) , more than three times the uln . ultrasonography of the abdomen was performed in order to confirm the presence of ascites and to rule out other causes of cholestasis ( e.g. , choledochal cyst and gallstones ) . tests for antimitochondrial antibodies ( amas ) to exclude the diagnosis of primary biliary cirrhosis ( pbc ) were done , and drugs hepatotoxicity was ruled out . renal dysfunction was determined on the basis of elevated levels of serum creatinine ( above the upper limit of normal , that is , 1.3  mg / dl ) .   in some cases , a ct scan was also performed when any doubts about the nature of the pathology existed . body mass index ( bmi ) and/or the waist - to - hip ratio ( whr ) are commonly used for evaluation of body composition and fat mass . however , we did not apply those indicators due to their bias and inaccuracy of fat content assessment in ascitic cirrhotics . ascites was present in 89 of 147 ( 60.5% ) subjects included in the survey . alcoholics with decompensated liver disease have both high prevalence of malnutrition and high bmi values due to fluid retention at the same time . furthermore , some of our patients could not stand at the time of hospital admission , so the measure of their weight and height was impossible . the applicability of these constants for an estimation of body fat mass has often been questioned [ 15 , 16 ] . the evaluation of plasma levels of selected adipokines ( total adiponectin - acrp30 , leptin , and resistin ) was performed with immunoenzymatic elisa ( enzyme linked immunosorbent assay ) tests using commercially available kits ( quantikine elisa kit , r & d systems , usa ) . the study was conducted according to the procedure recommended by the producer and described in the attached materials . the study protocol conforms to the ethical guidelines of the 1975 declaration of helsinki ( 6th revision , 2008 ) as reflected in a priori approval by the institutional review board of medical university of lublin . statistical analysis was performed using the statistica 10 software package ( statsoft , poland ) . the distribution of the data in the groups was preliminarily evaluated by kolmogorov and smirnov test . the analysis showed a skewed distribution of all values ; therefore , continuous variables were described as median with interquartile range , its 95% confidence interval ( 95%  ci ) , and compared using mann - whitney u - test . differences between categorical variables were assessed by fisher 's exact test or the  test with yates correction for continuity , as appropriate . the differences of studied adipokine levels between ctp classes were analyzed using kruskal - wallis and multiple comparisons post hoc tests . spearman 's rank correlation test was used for the assessment of association between parameters of liver function , traditional indicators of inflammation , and studied biomarker serum levels . the receiver operating curves ( rocs ) for significant adipokines , that is , acrp30 and resistin were constructed to assess their areas under the curve ( aucs ) and the best threshold values for predicting ald complications . the method of delong et al .   for the calculation of the standard error of the auc was used . the youden index and its associated cut - off point aucs of significant variables were compared to assess their accuracy in predicting the severity of liver dysfunction and the development of ald complications . multivariable logistic regression was applied in order to select independent predictors of advanced liver dysfunction and the occurrence of the disease complications . a two - sided p value of less than 0.05 was considered to be associated with statistical significance . the survey population included 147 patients ( 107 males ( 72.8% ) and 40 females ( 27.2% ) ) . the baseline characteristics of the study cohort are shown in tables 1 and 2 . initially we performed an assessment of the adipokine serum levels in patients with ald in comparison with healthy controls . the results indicated that serum acrp30 and resistin levels were significantly higher in patients with ald as compared to healthy controls . the next step was to compare the levels of studied biomarkers inside the ald and control group according to gender . serum acrp30 ( p = 0.004 ) and leptin ( p = 0.03 ) concentrations were significantly higher in females compared to males from the control group . on the other hand , none of studied adipokines showed a significant difference according to gender in ald group . furthermore , significantly higher levels of acrp30 in both sexes were found in ald patients as compared to the control group . resistin serum levels were significantly higher in both sexes in ald patients as compared to the control group . the results are presented in table 4 . since aging alters body fat mass and its function , we evaluated serum adipokine levels in two age subgroups : 50 and <  50 years old . as expected , serum leptin concentration was significantly higher in the older patients in comparison with the younger subgroup . it was found that only the level of acrp30 among studied adipokines significantly increased with the severity of liver dysfunction classified according to both ctp and meld scores . the analysis of the association of adipokine serum levels and liver function parameters showed a significant , positive correlation between leptin and both aminotransferases and bilirubin serum levels in males . there was an inverse correlation of acrp30 and resistin with serum albumin levels and a positive correlation of acrp30 with ap and bilirubin levels in men . we found a significant positive correlation of serum resistin level and both the white blood cells count and crp level . the next step of the study was carried out to compare the level of studied adipokines in ald patients divided according to the presence of the disease complications . for adipokines which serum concentrations differed significantly in subgroups of patients selected according to the severity of liver dysfunction ( meld  20 ) and the complications of ald , the areas under the curve ( aucs ) were checked , and the diagnostic accuracy of studied variables for the association with complications of the disease was compared . serum acrp30 levels were identified as an independent predictor of advanced liver dysfunction ( meld  20 ) and the development of ascites and he . none of the studied adipokines was independently associated with cholestasis and esophageal varices . above results excessive and chronic alcohol consumption leads to inflammation in adipose tissue , insulin resistance , and hepatic steatosis . therefore , the exploration of the pathogenic mechanisms of ald should include the role of adipose tissue secretion and adipokines . in addition , alcohol abuse is associated with impaired energy intake and expenditure , as well as increased catabolism . the recent study of zhong et al .   demonstrated a significant loss of white adipose tissue ( wat ) in a mouse model of alcoholic steatosis . it suggests that wat dysfunction can directly impact hepatic lipid homeostasis by reverse triglyceride transport . clinical studies have shown that lower fat mass is associated with higher liver fat content in alcoholics . those reports prompted us to design a study in order to explain the association of serum adipokine concentrations with the severity of liver dysfunction and ald complications . results obtained in the present study confirmed the crucial role of wat endocrine secretion in the pathogenesis of ald . significantly higher levels of two adipokines , acrp30 and resistin were found in patients with ald compared to the control subjects . unexpectedly , there were no differences in leptin levels in both studied groups ( table 3 ) . the analysis of data by gender showed significantly higher levels of acrp30 and leptin in females from the control group in comparison with the level of both adipokines in males ( table 4 ) . our results are consistent with other reports regarding gender - related differences of serum adipokine concentrations . however , no significant difference in the level of three studied adipokines in relation to sex in the ald group was observed . in contrast , a significantly lower leptin level was found in females with ald in comparison with healthy controls . when comparing the severity of liver dysfunction in both sexes , females showed significantly higher scores of the ctp and meld ( table 2 ) . we are tempted to speculate that metabolic alterations caused by ethanol in the course of ald , by modulating secretion of leptin , might be responsible for different clinical presentation of the disease in females and males . that ingestion of moderate amounts of alcohol had an inhibitory effect on leptin secretion in normal subjects . the effect might be direct rather than indirect , since several factors known to affect leptin were not influenced by alcohol in their study . furthermore , leptin levels increase during abstinence , and this may be related to a reduction of dopaminergic action in mesolimbic system . gender - related differences observed in our study are also consistent with the results obtained by dammann et al .   who observed no significant effect of acute moderate alcohol intake on leptin levels in healthy male volunteers . some of them pointed out increased [ 28 , 29 ] , and others lowered leptin concentrations in the peripheral blood . greco et al .   also observed higher serum leptin levels in healthy females compared to healthy males in the control group . however , unlike in the present work , they reported elevated levels of leptin in women with alcoholic cirrhosis child class c. the above mentioned study revealed a significant reduction in leptin serum levels in posthepatitis cirrhotic patients . the discrepancy between our results could be likely explained by evaluation of patients in different stages of ald . the level of leptin in our cohort was assessed during an acute phase of liver disease with coexistence of signs of hypermetabolism and systemic inflammatory activation . in contrast to present report , the above - cited study was performed in patients with a stable chronic phase of the disorder . de timary et al .   proposed a dual model for regulation of energy intake in alcohol - dependent subjects . they showed that alcohol accelerates metabolism and decreases fat mass and leptin levels in individuals consuming above 12.5  kcal / kg / day of alcohol ( lower leptin level was observed in females with ald in our study ) . for individuals consuming below 12.5  kcal / kg / day of alcohol , alcohol intake is compensated for by a decrease in nonalcoholic nutrient intakes , probably due to changes in metabolic and satiety factors .   on the other hand , campillo et al . observed an increase in leptin levels in the blood after nutritional therapy for patients with alcoholic cirrhosis in parallel to the liver function improvement . similar profile of serum adipokine concentrations that is elevated acrp30 and resistin , and decreased leptin was reported in patients with inflammatory bowel disease . we investigated the levels of adipokines in subgroups of ald patients with varying severity of hepatic dysfunction defined according to the criteria of the ctp and meld . we found that serum acrp30 levels significantly rose with the degree of liver function impairment ( tables 6 and 7 ) . multivariable analysis confirmed its independent impact on the severity of liver dysfunction ( meld  20 ) ( table 12 ) . it is in contrast to nonalcoholic fatty liver disease ( nafld ) , where serum adiponectin concentrations were reported to be low and negatively related to necroinflammatory injury [ 34 , 35 ] . on the other hand , a great body of evidence suggests a beneficial and protective effect of acrp30 in liver injury in mice [ 36 , 37 ] . its administration reduced ros production stimulated by lps and expression of tumor necrosis factor  ( tnf  ) in kupffer cells . the mechanism by which adiponectin levels and action differ between humans and rodents is still unknown . correlations were found in men between serum acrp30 levels and the parameters of liver function : positive with the level of bilirubin , inverse with albumin level ( table 8) . these results are consistent with previous indices of positive acrp30 association with the severity of liver dysfunction ( ctp and meld ) . in addition , a weak inverse correlation between serum resistin concentrations and the level of albumin in ald group was observed . furthermore , the results of our study indicate the presence of adipokine association with major complications of the disease ( table 10 ) . multivariable analysis confirmed independent impact of acrp30 on both complications ( table 12 ) .   the borderline statistical significance ( p = 0,054 ) of leptin level differences in subgroups with and without ascites was found and might suggest its negative influence on the evolution of ald ( table 10 ) . nevertheless , the multivariable analysis failed to prove its impact on the development of the above complication . as observed in our study , also brennan et al .   did not find any significant association of leptin concentration with morbidity and mortality for cardiovascular disease in women with diabetes . conversely , reports of other authors indicate that leptin deficiency may enhance the sensitivity to the toxic effects of inflammatory factors , including endotoxin and tnf  [ 4042 ] . serum resistin levels were significantly increased in the subgroups with symptoms of cholestasis and renal dysfunction ( table 9 ) . the presence of elevated levels of resistin was previously reported in the course of chronic kidney disease [ 4345 ] . on the other hand , menzaghi et al .   demonstrated that resistin may play an important role in modulating the kidney function in healthy subjects . in the present study , the multivariable analysis failed to confirm its independent effect on both the above - mentioned complications . suggestions about the prognostic value of adiponectin and resistin in the assessment of severity and outcome of inflammatory diseases and cancer appeared in several recent publications [ 4751 ] . we measured the plasma levels of total adiponectin ( acrp30 ) , which has been reported to be more useful than hmw for assessing mortality risk . the high plasma concentration of acrp30 was an independent prognostic predictor in chronic heart failure patients with normal bmi . higher concentrations of total adiponectin were also associated with heart failure and mortality among patients with existing ischemic heart disease . the development of ald is believed to combine metabolic and inflammatory activity , so the assessment of the relationship between serum levels of selected adipokines and inflammatory markers was of vast interest . we speculate that significantly increased serum levels of acrp30 and resistin in the studied cohort may be an indicator of systemic inflammatory activation in the course of the disease . the data from the literature suggest that the levels of both adipokines rise in chronic inflammation [ 5457 ] . we found its weak inverse correlation with the white blood cells count .   on the other hand , an increase in acrp30 concentration in the blood of patients with ald may reflect the defense mechanism rather than inflammatory response due to its known anti - inflammatory properties . furthermore , the level of acrp30 in the studied group did not show any correlation with the traditional parameters of inflammation ( table 9 ) . it was reported that acrp30 deficiency leads to persistent subclinical inflammation in the course of obesity , nonalcoholic fatty liver disease ( nafld ) , ischemic heart disease , and type 2 diabetes [ 5963 ] . in contrast , increased levels of serum adiponectin have been observed in inflammatory and/or autoimmune disorders which show no association with obesity and positive energy balance . those include rheumatoid arthritis , lupus erythematosus , and inflammatory bowel diseases [ 64 , 65 ] . it appears that secretion of acrp30 may be differently regulated depending on the pathogenesis of underlying disease , in particular on its association with energy balance ( overweight or obesity ) . furthermore , behre   hypothesized that in subjects with an energy deficit in the course of anorexia , cachexia , type 1 diabetes , and renal failure , when the blood level of adiponectin increased , it might have a protective role and adapt the body during fasting . the results obtained in the present study are consistent with such an interpretation of the facts . the patients with ald , regardless of their gender , had significantly elevated plasma levels of acrp30 compared with the control group ( table 4 ) . it may be explained by malnutrition frequently occurring in the course of the disease . proinflammatory properties of resistin in our study were confirmed by a positive correlation of its serum concentration with the white blood cells count and crp level ( table 9 ) . reported that the plasma levels of resistin and leptin positively correlated with the crp level in patients with rheumatoid arthritis . the level of crp was significantly associated with an increased level of resistin in that study . the results of trzeciak - ryczek et al .   from in vitro studies revealed that macrophage stimulation with lipopolysaccharide or pro - inflammatory cytokines ( il-1 , il-6 and tnf ) considerablly increased resistin production during infection . the presented data support the hypothesis that resistin may become a new marker of inflammation . it was a single - center trial , and it should be emphasized that due to the insufficient sample size , our results before their generalization need to be confirmed in prospective , multicenter studies . such validation may help to eliminate possible errors resulting from research techniques and subjective differences in the selection of the study population . another limitation of the study was lack of estimation of the body fat content by objective and validated measures . the newest diagnostic option is dual - energy x - ray absorptiometry ( dexa ) , which may be used as a gold standard with high accuracy . nevertheless , the procedure was unavailable during our study . furthermore , the control group of healthy abstainers and light drinkers ( 20  g / day ) was used as the reference category for alcohol consumption in the study . more often disease - free drinkers are included as a comparison cohort to control for the possible effect of alcohol consumption . participants had to determine the amount of alcohol consumed in drinks per day , week , and/or month . since these values ( i.e. , the volume and alcohol concentration of a beverage ) are not easy to estimate , the real ethanol intake may lack precision . a study by stockwell and stirling   also showed that most people are not able to accurately assess the volume and power of a drink . in countries like poland , alcohol intake patterns vary considerably by regions and beverage type , so researchers should pay special attention to careful assessment of drink types and sizes for accurate alcohol consumption estimation . in conclusion , the results obtained in the present study confirmed the active participation of wat in the pathogenesis of ald . we speculate that gender - related differences in serum leptin concentrations may influence the ald course , different in females compared to males . it seems that , beyond their metabolic influence , adipokines may play a role of inflammatory modulators . an independent association of the high serum acrp30 level with advanced liver dysfunction , as well as the development of ald complications ( i.e. , ascites and hepatic encephalopathy ) may indicate its potential role as an ald prognostic indicator . in addition , due to the inverse serum concentration , acrp30 may become a relevant marker for differentiation between ald and nafld . the association of resistin levels with renal dysfunction in ald patients should be further elucidated . good availability of serum biomarkers makes them a promising , minimally invasive diagnostic tool for the possible widespread use in the clinical practice . it appears that the determination of molecules involved in different stages of the disease evolution may help to generate a reliable diagnostic algorithm in the future . whether combination therapy with adipokines may have a role in ald patients the modulation of their secretion might be considered as a tempting therapeutic procedure but needs to be investigated in detail .
background and aims . 
 there is growing evidence that white adipose tissue is an important contributor in the pathogenesis of alcoholic liver disease ( ald ) . 
 we investigated serum concentrations of total adiponectin ( acrp30 ) , leptin , and resistin in patients with chronic alcohol abuse and different grades of liver dysfunction , as well as ald complications . 
 materials and methods . 
 one hundred forty - seven consecutive inpatients with ald were prospectively recruited . 
 the evaluation of plasma adipokine levels was performed using immunoenzymatic elisa tests . 
 multivariable logistic regression was applied in order to select independent predictors of advanced liver dysfunction and the disease complications . 
 results . acrp30 and resistin levels 
 were significantly higher in patients with ald than in controls . 
 lower leptin levels in females with ald compared to controls , but no significant differences in leptin concentrations in males , were found . 
 high serum acrp30 level revealed an independent association with advanced liver dysfunction , as well as the development of ald complications , that is , ascites and hepatic encephalopathy . 
 conclusion . 
 gender - related differences in serum leptin concentrations may influence the ald course , different in females compared with males . 
 serum acrp30 level may serve as a potential prognostic indicator for patients with ald .