Proteomic Analysis of Polypeptides Captured from Blood during Extracorporeal Albumin Dialysis in Patients with Cholestasis and Resistant Pruritus

Marina Gay,Joaquin Abian,Marina Gorga,Montserrat Carrascal,Pau Bosch-I-Crespo,Antoni Mas,Albert Pares

doi:10.1371/journal.pone.0021850

Abstract

Albumin dialysis using the molecular adsorbent recirculating system (MARS) is a new therapeutic approach for liver diseases. To gain insight into the mechanisms involved in albumin dialysis, we analyzed the peptides and proteins absorbed into the MARS strong anion exchange (SAX) cartridges as a result of the treatment of patients with cholestasis and resistant pruritus. Proteins extracted from the SAX MARS cartridges after patient treatment were digested with two enzymes. The resulting peptides were analyzed by multidimensional liquid chromatography coupled to tandem mass spectrometry. We identified over 1,500 peptide sequences corresponding to 144 proteins. In addition to the proteins that are present in control albumin-derived samples, this collection includes 60 proteins that were specific to samples obtained after patient treatment. Five of these proteins (neutrophil defensin 1 [HNP-1], secreted Ly-6/uPAR-related protein 1 [SLURP1], serum amyloid A, fibrinogen alpha chain and pancreatic prohormone) were confirmed to be removed by the dialysis procedure using targeted selected-reaction monitoring MS/MS. Furthermore, capture of HNP-1 and SLURP1 was also validated by Western blot. Interestingly, further analyses of SLURP1 in serum indicated that this protein was 3-fold higher in cholestatic patients than in controls. Proteins captured by MARS share certain structural and biological characteristics, and some of them have important biological functions. Therefore, their removal could be related either to therapeutic or possible adverse effects associated with albumin dialysis.

Highlights

Extracorporeal albumin dialysis (ECAD) using the molecular adsorbent recirculating system (MARS) has been used in patients with acute-on-chronic liver failure or acute liver failure, and in the treatment of primary graft dysfunction after liver transplantation with favorable, indefinite, results in terms of survival [1], [2], [3], [4], [5], [6], [7], [8], [9]. Another application of MARS is the treatment of resistant pruritus in chronic cholestatic diseases and in patients with pruritus resulting from liver graft rejection and severe cholestasis
Parallel analysis by MDLC-ESIMS/MS of tryptic and GluC (Staphylococcus aureus protease V8) digests of the MARS strong anion exchange (SAX) extracts led to the identification of a total of 1,726 distinct peptides (FDR,1%)
There is still little knowledge of the proteins and other biological compounds that are removed from blood during albumin dialysis

Summary

Introduction

Extracorporeal albumin dialysis (ECAD) using the molecular adsorbent recirculating system (MARS) has been used in patients with acute-on-chronic liver failure or acute liver failure, and in the treatment of primary graft dysfunction after liver transplantation with favorable, indefinite, results in terms of survival [1], [2], [3], [4], [5], [6], [7], [8], [9] Another application of MARS is the treatment of resistant pruritus in chronic cholestatic diseases and in patients with pruritus resulting from liver graft rejection and severe cholestasis. None of these molecules have been shown to be pruritogens and the pathogenesis of pruritus in cholestasis is still not firmly established [10], [11], [12], [13]

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Methods

Results