Albumin redox state of maintenance haemodialysis patients is positively altered after treatment.

BACKGROUND AND AIM: Maintenance haemodialysis patients have increased morbidity and mortality which is mainly driven by an elevated inflammation level due to the uraemic milieu. A major part of this increased inflammation level is the degree of oxidative stress which can be assessed by albumin redox state (ARS). Aim of this study was to evaluate how the ARS is affected by a haemodialysis treatment and different dialyzer properties. METHODS: ARS was determined before and after haemodialysis treatment by fractionating it into reduced human mercaptalbumin (HMA), reversibly oxidized human non-mercaptalbumin 1 (HNA-1), and irreversibly oxidized human non-mercaptalbumin 2 (HNA-2) by high-performance liquid chromatography. In healthy individuals, albumin circulates in the following proportions: HMA 70-80%, HNA-1 20-30% and HNA-2 2-5%. High flux (FX 100 [Fresenius Medical Care], BG 1.8 [Toray], Xevonta Hi 18 [B. Braun], CTA-2000 [Kawasumi]) and low flux FX10 [Fresenius Medical Care] dialyzers were used. RESULTS: 58 patients (59% male, median age 68 years, median time on haemodialysis 32 month) were included in the study. Before haemodialysis treatment, HMA (median 55.9%, IQR 50.1-61.2%) was substantially lower than in healthy individuals. Accordingly, oxidized albumin fractions were above the level of healthy individuals (median HNA-1 38.5%, IQR 33.3-43.2%; median HNA-2 5.8%, IQR 5.1-6.7%). Before haemodialysis treatment HMA was significantly higher in patients usually treated with high flux membranes (p < 0.01). After haemodialysis treatment there was a significant increase of HMA and a decrease of HNA-1 and HNA-2 (p < 0.01). These effects were more pronounced in patients treated with high flux dialyzers (p < 0.01). There were no differences of ARS alteration with regard to the dialyzer´s sterilization mode or the presence of diabetes. CONCLUSION: The study confirms that the ARS is positively altered by haemodialysis and shows for the first time that this effect depends on dialyzer properties.

Redox state as assessed using the measurement of human non-mercaptalbumin in embryo culture media is associated with successful embryo development in human in vitro fertilization.

The role of oxidative stress in the pathogenesis of various diseases has been attracting attention. We speculated as to whether the redox state of treatment solutions used for various diseases may play a role in treatment success. In the current study, we focused on the human embryo culture medium used for in vitro fertilization (IVF). A total of 173 oocytes from a total of 91 patients treated with IVF were enrolled. The redox state was assessed by measuring the levels of human non-mercaptalbumin (HNA). We analyzed factors related to blastocyst formation on day 5 or 6 after insemination. We also developed a random forest (RF) model for the prediction of blastocyst formation. The variable importance in the predictive model was assessed using the mean decrease in the Gini impurity. Blastocyst formation was observed in 41.04% (71/173) of the oocytes and was associated with a lower %HNA in the culture medium, a younger patient age, and the fertilization method (standard IVF or intracytoplasmic sperm injection). The RF model developed using these factors and 70% of the samples (training set, n = 121) was validated in the remaining testing set (n = 52) and produced an area under the curve of 0.761, where the %HNA in the culture medium was the most important variable for predicting blastocyst formation. In conclusion, lower levels of oxidative stress in embryo culture media were associated with the success of IVF treatment. The redox state of treatment solutions should be considered to support treatment success.

Oxidative albumin damage in chronic liver failure: relation to albumin binding capacity, liver dysfunction and survival.

BACKGROUND & AIMS: Impaired 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. METHODS: Patients with decompensated cirrhosis or sepsis and healthy controls were investigated. Three fractions of albumin were separated by chromatography according to the redox state of cysteine-34: non-oxidized human mercaptalbumin, reversibly oxidized human non-mercaptalbumin-1, and irreversibly oxidized human non-mercaptalbumin-2 (HNA2). Binding properties of albumin site II were measured using dansylsarcosine as a ligand. RESULTS: Both in cirrhotic and septic patients, fractions of oxidized albumin were increased and binding capacity for dansylsarcosine was decreased. Mass spectroscopy confirmed specific oxidation of cysteine-34. In cirrhotic patients, dansylsarcosine binding correlated strongly with liver function parameters and moderately with HNA2. Baseline levels of HNA2 accurately predicted 30-day and 90-day survival in cirrhotic patients and this was confirmed in an external validation cohort. CONCLUSIONS: Our results suggest that oxidative damage impairs binding properties of albumin. In advanced liver disease, reduced binding capacity of albumin site II is mainly related to impaired liver function. The plasma level of HNA2 is closely related to survival and may represent a novel biomarker for liver failure.

A simple colorimetric assay to determine the concentration and proportion of human mercaptalbumin.

Objectives: Human serum albumin can take on two forms, mercaptalbumin (HMA) or non-mercaptalbumin (HNA), depending on the redox status of its Cys34. The ratio of HMA and HNA is considered to be a novel biomarker of oxidative stress. While HPLC and mass spectrometry are established methods to measure HMA and HNA, a simple colorimetric assay was applied to measure this biomarker. Design and methods: Michler's Hydrol (4,4'-Bis(dimethylamino)benzhydrol) is a blue dye with a maximum absorption at 612 nm, and its absorption decreases when it reacts with a thiol group. Concentrations of HMA in serum samples from 36 healthy subjects were measured based on absorption changes of Michler's Hydrol. The proportion of HMA (HMA%) in total albumin was also obtained by dividing the HMA concentration by total albumin concentration, which was obtained by a bromocresol purple (BCP) assay. The proportion of HNA (HNA%) was obtained by subtracting HMA% from 100%. Results: HMA concentrations obtained by Michler's Hydrol assay were highly correlated (r2 = 0.97) with reference values obtained by HPLC (HMA%) and BCP assay (total albumin). The HNA% obtained by Michler's Hydrol and BCP assays combined also gave a good correlation (r2 = 0.96) and a small deviation (average 2.4%) with respect to HPLC as a reference method. Conclusions: A colorimetric assay using Michler's Hydrol was optimized for a 96-well plate format so that it can be easily performed in a standard laboratory setting. This assay gives HMA concentrations and HNA proportions comparable to HPLC.

A Method for Stabilizing the Proportion of the Reduced Form of Albumin During Cell-Free and Concentrated Ascites Reinfusion Therapy in Patients with Malignant Ascites.

Cell-free and concentrated ascites reinfusion therapy (CART) is used to treat malignant ascites. However, the qualities of albumin in malignant ascites, such as antioxidative activity, may decrease owing to oxidative stress caused by cancer cells and inflammatory reactions. We investigated the fraction percentages of mercaptalbumin (HMA%, reduced form) and non-mercaptalbumin (HNA%, oxidized form) in malignant ascites from 21 patients who received CART and compared the HMA% in the malignant ascites and human serum albumin (HSA) preparations. HMA% of albumin in malignant ascites (22.5%) was significantly lower than that in HSA preparation (42.2%). To ensure a high HMA%, we added L-cysteine to the paracentesis-treated ascites followed by dialysis 1 h later. As a result, the HMA% of albumin in malignant ascites was increased to 59.1%. Our results suggest that using this method in CART will improve patient's albumin quality.