Effective albumin concentration and cirrhosis mortality: From concept to reality
Authors: Rajiv Jalan, Mauro Bernardi
Journal: Journal of Hepatology (2013)

Oxidative albumin damage in chronic liver failure: Relation to albumin binding capacity, liver dysfunction and survivalJournal of HepatologyVol. 59Issue 5PreviewImpaired binding function of albumin has been demonstrated in end-stage liver disease. This and other functional disturbances of albumin may be related to oxidative stress which is believed to play an important role in the pathogenesis of liver failure as well as sepsis. The aim of the present study was to relate oxidative modification of albumin to loss of albumin binding function in advanced chronic liver failure and in sepsis. Full-Text PDF Albumin is the most abundant protein in human plasma that is produced exclusively in the liver. Traditionally, albumin has been thought of being important in providing plasma with its oncotic property and therefore its use in clinical practice has been mainly aimed at promoting plasma volume expansion. The past 10 years or so has seen an explosion in the knowledge of albumin biology such that it is now clear that it has multifunctional properties ranging from provision of oncotic pressure, immune regulation and endothelial stabilization to being a molecule that works in the intracellular compartment modifying several key pathophysiological mechanisms (reviewed in Garcie-Martinez et al.) [[1]Garcia-Martinez R. Caraceni P. Bernardi M. Gines P. Arroyo V. Jalan R. Albumin: pathophysiologic basis of its role in the treatment of cirrhosis and its complications.Hepatology. 2013; (in press)Google Scholar]. It has been hypothesized that the pleiotropic effects of albumin in cirrhosis and its proven effectiveness in spontaneous bacterial peritonitis, prevention of post paracentesis circulatory dysfunction, hepatorenal syndrome and hepatic encephalopathy relates to these functional characteristics rather than simple volume expansion [[1]Garcia-Martinez R. Caraceni P. Bernardi M. Gines P. Arroyo V. Jalan R. Albumin: pathophysiologic basis of its role in the treatment of cirrhosis and its complications.Hepatology. 2013; (in press)Google Scholar]. Indeed, albumin administration, but not hydroxyethyl starch, improves systemic hemodynamics in patients with spontaneous bacterial peritonitis by decreasing endothelial activation [[2]Fernández J. Monteagudo J. Bargallo X. Jiménez W. Bosch J. Arroyo V. et al.A randomized unblinded pilot study comparing albumin vs. hydroxyethyl starch in spontaneous bacterial peritonitis.Hepatology. 2005; 42: 627-634Crossref PubMed Scopus (193) Google Scholar]. Moreover, in experimental cirrhosis in rats, albumin exerts a positive cardiac inotropic effect counteracting oxidative stress- and TNF-α-induced impairment of cardiac contractility [[3]Bortoluzzi A. Ceolotto G. Gola E. Sticca A. Bova S. Morando F. et al.Positive cardiac inotropic effect of albumin infusion in rodents with cirrhosis and ascites: molecular mechanisms.Hepatology. 2013; 57: 266-276Crossref PubMed Scopus (94) Google Scholar]. Given the pathophysiological importance of cirrhotic cardiomyopathy in hepatorenal syndrome, it is likely that the beneficial effects of albumin in this setting are mediated, at least in part, by these mechanisms. The paper in the current issue of the Journal of Hepatology by Oettl et al. adds to that growing literature by suggesting that albumin may be irreversibly oxidized in advanced cirrhosis, and this reduction in albumin function predicts mortality [[4]Oettl K. Birner-Gruenberger R. Spindelboeck W. Stueger H.P. Dorn L. Stadlbauer V. et al.Oxidative albumin damage in chronic liver failure: relation to albumin binding capacity, liver dysfunction and survival.J Hepatol. 2013; 56: 978-983Abstract Full Text Full Text PDF Scopus (122) Google Scholar]. Albumin is 67 kDa protein and because of its amino acid composition it has a negative net charge at pH 7, which makes it soluble and imparts its oncotic properties. From the functional perspective, albumin can be considered to have three essential classes of functional domains. It has a metal binding domain, which gives it the property to bind to and remove highly toxic reactive metal species. Assays, which measure binding of cobalt to albumin have been developed and the molecule defective in this function is known as 'ischemia-modified albumin (IMA)'. The second is a group of domains that bind a variety of substances such as long-chain fatty acids, bilirubin, anions, bile acids, endotoxin, hormones, and eicosanoids. The function of these binding domains can be studied using electron paramagnetic resonance spectroscopy (EPR), which essentially measures the binding of and release characteristics of albumin using labeled stearic acid [1Garcia-Martinez R. Caraceni P. Bernardi M. Gines P. Arroyo V. Jalan R. Albumin: pathophysiologic basis of its role in the treatment of cirrhosis and its complications.Hepatology. 2013; (in press)Google Scholar, 5Jalan R. Schnurr K. Mookerjee R.P. Sen S. Cheshire L. Hodges S. et al.Alterations in the functional capacity of albumin in patients with decompensated cirrhosis is associated with increased mortality.Hepatology. 2009; 50: 555-564Crossref PubMed Scopus (177) Google Scholar]. Studies of the functional capacity of albumin in cirrhotic patients using these techniques were described recently. Albumin functional impairment closely correlated with the degree of liver insufficiency. The IMA/total albumin ratio was significantly higher in non-surviving patients with acute-on-chronic liver failure (ACLF) leading to the concept of 'effective albumin concentration', which is dependent upon the functional characteristics of albumin rather than its quantity. Importantly, the investigators showed that albumin dialysis did not restore the functional ability of the native molecule suggesting that albumin in liver failure may be irreversibly destroyed [[5]Jalan R. Schnurr K. Mookerjee R.P. Sen S. Cheshire L. Hodges S. et al.Alterations in the functional capacity of albumin in patients with decompensated cirrhosis is associated with increased mortality.Hepatology. 2009; 50: 555-564Crossref PubMed Scopus (177) Google Scholar]. The third major function of albumin is its thiol function, which is imparted by a free cysteine molecule. It is this property that was the main focus of the article by Oettl et al. [[4]Oettl K. Birner-Gruenberger R. Spindelboeck W. Stueger H.P. Dorn L. Stadlbauer V. et al.Oxidative albumin damage in chronic liver failure: relation to albumin binding capacity, liver dysfunction and survival.J Hepatol. 2013; 56: 978-983Abstract Full Text Full Text PDF Scopus (122) Google Scholar]. The free cysteine (Cys-34) residue provides a single free redox-active thiol (-SH). This is capable of reducing oxidative stress through thiolation, nitrosylation, and oxidation. In healthy individuals, albumin exists mostly in a reduced form (with a free thiol). This is referred to as mercaptoalbumin (HMA). A small proportion of albumin exists as mixed disulfide compounds known as non-mercaptalbumin 1 (HNA1), representing a molecule that has lost its anti-oxidant property but can be returned to the HMA form using a reducing agent. In health, a tiny fraction is found in a highly oxidized form with cysteine 34 (as sulfinic or sulfonic acid) known as non-mercaptalbumin 2 (HMA2). It is the sulfonic form that is thought to constitute an irreversibly damaged albumin [1Garcia-Martinez R. Caraceni P. Bernardi M. Gines P. Arroyo V. Jalan R. Albumin: pathophysiologic basis of its role in the treatment of cirrhosis and its complications.Hepatology. 2013; (in press)Google Scholar, 6Oettl K. Stadlbauer V. Petter F. Greilberger J. Putz-Bankuti C. Hallstrom S. et al.Oxidative damage of albumin in advanced liver disease.Biochim Biophys Acta. 2008; 1782: 469-473Crossref PubMed Scopus (118) Google Scholar]. These three different fractions of albumin can be isolated and quantified by high performance liquid chromatography (HPLC). During aging [[7]Oettl K. Marsche G. Redox state of human serum albumin in terms of cysteine-34 in health and disease.Methods Enzymol. 2010; 474: 181-195Crossref PubMed Scopus (81) Google Scholar] and different diseases, such as end-stage kidney disease [[8]Matsuyama Y. Terawaki H. Terada T. Era S. Albumin thiol oxidation and serum protein carbonyl formation are progressively enhanced with advancing stages of chronic kidney disease.Clin Exp Nephrol. 2009; 13: 308-315Crossref PubMed Scopus (75) Google Scholar] and liver failure [[6]Oettl K. Stadlbauer V. Petter F. Greilberger J. Putz-Bankuti C. Hallstrom S. et al.Oxidative damage of albumin in advanced liver disease.Biochim Biophys Acta. 2008; 1782: 469-473Crossref PubMed Scopus (118) Google Scholar], the fractions of the oxidized forms are increased. The paper by Oettl et al. extends this observation and shows for the first time that the proportion of HNA2 measured in the plasma of patients with advanced cirrhosis may predict mortality [[4]Oettl K. Birner-Gruenberger R. Spindelboeck W. Stueger H.P. Dorn L. Stadlbauer V. et al.Oxidative albumin damage in chronic liver failure: relation to albumin binding capacity, liver dysfunction and survival.J Hepatol. 2013; 56: 978-983Abstract Full Text Full Text PDF Scopus (122) Google Scholar]. The authors prospectively studied 67 patients who were admitted to the hospital with acute decompensation of cirrhosis of whom 18 patients were in the intensive care unit. In addition to the clinical parameters, they measured albumin function, specifically, HMA, HNA1, and HNA2 using HPLC, i.e., looking at the free cysteine status and they also measured the binding characteristics of albumin to danylsarcosine, which looks at the functional activity of binding site 2 using HPLC as well. They found that the concentration of both oxidized forms were significantly increased in the cirrhotic patients and the value of HNA2, in particular correlated closely with the MELD score, bilirubin, INR, and CRP, confirming that the values were reflected by the severity of liver disease and associated inflammation, both of which have been identified as predictors of ACLF [9Moreau R. Jalan R. Gines P. Pavesi M. Angeli P. Cordoba J. et al.Acute-on-chronic liver failure is a distinct syndrome that develops in patients with acute decompensation of cirrhosis.Gastroenterology. 2013; 144: 1426-1437Abstract Full Text Full Text PDF PubMed Scopus (1894) Google Scholar, 10Jalan R. Stadlbauer V. Sen S. Cheshire L. Chang Y.M. Mookerjee R.P. Role of predisposition, injury, response and organ failure in the prognosis of patients with acute-on-chronic liver failure: a prospective cohort study.Crit Care. 2012; 16: R227Crossref PubMed Scopus (92) Google Scholar]. Therefore, it was not surprising and important that they found a high predictive ability of HNA2 in determining 30- and 90-day mortality. In fact, the prediction of 30-day mortality was possibly higher than MELD score. A cut-off of 12% was considered optimal and identified a cohort with a very low survival rate. They observed a correlation between this and danylsarcosine binding which may suggest that the structural change due to oxidation of albumin may have non-specific effects on other functional characteristics of albumin. They then validated these data in a second cohort of 40 similar patients from Innsbruck. Finally, in a small sample of 3 patients, they extended their findings using mass spectrometry and confirmed that the HNA2 fraction represented an irreversible damage to albumin as the modification of the Cys-34 was in the sulfonic acid form [[4]Oettl K. Birner-Gruenberger R. Spindelboeck W. Stueger H.P. Dorn L. Stadlbauer V. et al.Oxidative albumin damage in chronic liver failure: relation to albumin binding capacity, liver dysfunction and survival.J Hepatol. 2013; 56: 978-983Abstract Full Text Full Text PDF Scopus (122) Google Scholar]. The data from this study significantly adds to the weight of the argument that albumin dysfunction is likely to be pathophysiologically important [1Garcia-Martinez R. Caraceni P. Bernardi M. Gines P. Arroyo V. Jalan R. Albumin: pathophysiologic basis of its role in the treatment of cirrhosis and its complications.Hepatology. 2013; (in press)Google Scholar, 4Oettl K. Birner-Gruenberger R. Spindelboeck W. Stueger H.P. Dorn L. Stadlbauer V. et al.Oxidative albumin damage in chronic liver failure: relation to albumin binding capacity, liver dysfunction and survival.J Hepatol. 2013; 56: 978-983Abstract Full Text Full Text PDF Scopus (122) Google Scholar]. It would be useful to know in a larger cohort of patients whether HNA2 is a surrogate marker of ACLF or independent of it and whether changes in its concentration occur in the patients who deteriorate in the hospital (Fig. 1) . These data provide a possible explanation why the current albumin dialysis systems, such as the molecular adsorbents recirculating system (MARS) or Prometheus, have not been shown to improve the survival of patients with liver failure [11Banares R. Nevens F. Larsen F.S. Jalan R. Albillos A. Relief Study group. Extracorporeal liver support with the molecular adsorbent recirculating system (MARS) in patients with acute-on-chronic liver failure (AOCLF). The RELIEF trial.J Hepatol. 2010; 52: S459Abstract Full Text PDF Google Scholar, 12Kribben A. Gerken G. Haag S. Herget-Rosenthal S. Treichel U. Betz C. et al.Effects of fractionated plasma separation and adsorption on survival in patients with acute-on-chronic liver failure.Gastroenterology. 2012; 142: 782-789Abstract Full Text Full Text PDF PubMed Scopus (292) Google Scholar]. The concept these devices are based upon is the potential reversibility of albumin dysfunction in liver failure as the damaged albumin is not removed and remains in the circulation. In MARS, the patient's blood is dialysed against albumin across a membrane with a 55 kDa cut-off [[11]Banares R. Nevens F. Larsen F.S. Jalan R. Albillos A. Relief Study group. Extracorporeal liver support with the molecular adsorbent recirculating system (MARS) in patients with acute-on-chronic liver failure (AOCLF). The RELIEF trial.J Hepatol. 2010; 52: S459Abstract Full Text PDF Google Scholar]. Therefore, if the patient's albumin is irreversibly destroyed, it cannot be regenerated. In Prometheus, the patient's is simply cleaned by filters in the secondary circuit [[12]Kribben A. Gerken G. Haag S. Herget-Rosenthal S. Treichel U. Betz C. et al.Effects of fractionated plasma separation and adsorption on survival in patients with acute-on-chronic liver failure.Gastroenterology. 2012; 142: 782-789Abstract Full Text Full Text PDF PubMed Scopus (292) Google Scholar]. An irreversibly destroyed albumin will therefore not be regenerated. By contrast, plasma exchange, which allows removal of the damaged molecule, has been recently shown to improve survival in acute liver failure patients but is limited by non-selectivity [[13]Larsen F.S. Schimdt L.E. Wendon J. Hockerstedt K. Isoniemi H. Bernal W. et al.Liver assisting with high-volume plasma exchange in patients with acute liver failure.Hepatology. 2010; 52: 376AGoogle Scholar]. Based upon these data, newer devices that address albumin exchange are in development [[14]Davies N. Proven A. Leckie P. Storr M. Flieg R. Schenk M. et al.The development and testing of the UCL liver support device (UCL-ARSENEL): improvement of survival in paracetamol-induced acute liver failure pigs.Hepatology. 2010; 52: 1092AGoogle Scholar]. The data in this study generates numerous new questions. It is not clear whether the increased concentration of HNA2 detected in the patients who will die is only a marker or whether this may actually be harmful to cell function. The mechanism of formation of HNA2 remains to be defined. In the test tube, HNA2 can be generated by the addition of hydrogen peroxide. Therefore, it is likely that inflammation and the associated oxidative stress can produce HNA2 in vivo. An alternative question is whether the stressed hepatocyte is producing a functionally damaged albumin due to cellular stress leading to alterations in the folding of the albumin molecule. This will need to be investigated in future studies. The authors have clearly identified a potential biomarker. Larger numbers of patients will be required to confirm their data and it will be important to simultaneously study all the known albumin functions to tease out whether more than one function will further improve its prognostic ability. In order for this to happen, a simpler bedside test will need to be developed. Finally, it will be important to answer whether the concentration of HNA2 changes with either deterioration or improvement of liver function and whether instead of aiming at a target concentration of albumin, one should be aiming at a 'functional concentration' of albumin, i.e., a goal directed approach [[1]Garcia-Martinez R. Caraceni P. Bernardi M. Gines P. Arroyo V. Jalan R. Albumin: pathophysiologic basis of its role in the treatment of cirrhosis and its complications.Hepatology. 2013; (in press)Google Scholar]. In conclusion, on-going research in the field of albumin biology in liver failure has opened up new translational possibilities which will impact on finding new indications for albumin use, functional characterization to define 'effective albumin concentration', and the use of these concepts to design new albumin dialysis systems. Rajiv Jalan has ongoing research collaboration with Grifols, a company that manufactures albumin and with GAMBRO a company that makes an albumin dialysis device, MARS. He has received lecture fees from Grifols. UCL have filed patents around the UCL-ARSENEL device. Mauro Bernardi has received fees, as consultant and lecturer, by CLS Behring GmH, a Company that manufactures albumin, as lecturer, by the Plasma Protein Therapeutics Association (PPTA) Europe, which represents the private sector manufacturers of plasma-derived and recombinant analog therapies, and as lecturer, by Grifols Italia S.p.A.