Safety and performance of the Clearum™ high flux hemodialyzer.

BACKGROUND: Clearum™ is a high flux steam sterilized dialyzer for patients with hemodialysis or hemodiafiltration. This study evaluated the safety and performance of the Clearum high flux steam sterilized hemodialyzer in the removal of small and middle-sized toxins. METHODS: A prospective, interventional, nonrandomized study enrolled twenty end-stage renal disease patients undergoing hemodialysis. The Clearum high flux steam sterilized dialyzer was compared to Fresenius FX dialyzers for baseline comparison. The duration of the trial was 2 weeks for the FX dialyzer and 6 weeks with the Clearum high flux steam sterilized dialyzer. In vitro studies with dextrans of varying sizes were performed to compare the membrane characteristics and sieving coefficient curves for the two dialyzers. RESULTS: The primary objective of a mean urea reduction ratio >65% was met, with no significant difference in mean urea reduction ratio between the Clearum high flux steam sterilized and Fresenius FX-series of dialyzers (p = 0.86). No dialyzer-related adverse events were reported in the study. ß-2-microglobulin reduction with the Clearum high flux steam sterilized dialyzer was statistically higher than the FX-series dialyzer (66.5% vs. 53.6%; p < 0.0001). Predialysis interleukin-6 and C-reactive protein concentrations, blood-rest scores (residual blood after blood restitution), and thrombin-anti-thrombin values were comparable. Albumin remained stable during the 6 weeks of Clearum high flux steam sterilized dialyzer use, with no appreciable differences compared to the Fresenius FX-series. CONCLUSION: The Clearum high flux steam sterilized dialyzer showed good mid-term effectivity for small and middle molecule removal with no reported dialyzer-related adverse events.

Development of a degenerated TaqMan real-time Q-PCR for detection of bacteria-free DNA in dialysis fluid.

Bacterial-derived DNA fragments (BDNAs) have been shown to be present in a dialysis fluid, to pass through dialyzer membranes, and to induce interleukin 6 (IL-6) in mononuclear cells. DNA fragments are thought to be derived from microorganisms inhabiting hemodialysis water and fluid. The primary aim of the present study was to develop two degenerated TaqMan real-time quantitative-PCR (Q-PCR) for detection of a broad range of bacterial DNA that specifically detect 16S ribosomal DNA (rDNA) (862 and 241 bp) and evaluate the efficiency of the Bellco Selecta resin to capture the BDNAs in the dialysis fluid. For this purpose, we decided to compare measurements of unfragmented samples (9.8 × 105 Escherichia coli genome) with artificially fragmented DNA samples. We assessed two broad-range real-time PCR targeting bacterial 16S rRNA genes for detection of fragmented and unfragmented bacterial DNA in the dialytic fluid and demonstrated that Bellco Selecta resin is capable of retaining these types of bacterial DNA.

Direct electrical control of IgG conformation and functional activity at surfaces.

We have devised a supramolecular edifice involving His-tagged protein A and antibodies to yield surface immobilized, uniformly oriented, IgG-type, antibody layers with Fab fragments exposed off an electrode surface. We demonstrate here that we can affect the conformation of IgGs, likely pushing/pulling electrostatically Fab fragments towards/from the electrode surface. A potential difference between electrode and solution acts on IgGs' charged aminoacids modulating the accessibility of the specific recognition regions of Fab fragments by antigens in solution. Consequently, antibody-antigen affinity is affected by the sign of the applied potential: a positive potential enables an effective capture of antigens; a negative one pulls the fragments towards the electrode, where steric hindrance caused by neighboring molecules largely hampers the capture of antigens. Different experimental techniques (electrochemical quartz crystal microbalance, electrochemical impedance spectroscopy, fluorescence confocal microscopy and electrochemical atomic force spectroscopy) were used to evaluate binding kinetics, surface coverage, effect of the applied electric field on IgGs, and role of charged residues on the phenomenon described. These findings expand the concept of electrical control of biological reactions and can be used to gate electrically specific recognition reactions with impact in biosensors, bioactuators, smart biodevices, nanomedicine, and fundamental studies related to chemical reaction kinetics.

Radical improvement of signs and symptoms in systemic lupus erythematosus when treated with hemodiafiltration with endogenous reinfusion dialysis.

Lupus nephritis is one of the most serious complications of systemic lupus erythematosus (SLE). In the kidney, immune complexes and autoantibodies activate mesangial cells that secrete cytokines that can further amplify inflammatory processes. We present the case of a 42-year-old woman with lupus nephritis accompanied by periods of exacerbation of SLE, with necrotic-like skin lesions, psoriatic arthritis without skin psoriasis, purpura of the lower limb, petechial rash, joint pain, fever, eyelid edema with bilateral conjunctival hyperemia and itching. The patient underwent a dialytic treatment of hemodiafiltration with endogenous reinfusion. The technique uses the super-high-flux membrane Synclear 02 (SUPRA treatment) coupled with an adsorbent cartridge that has affinity for many toxins and mediators. Fever and joint pain were immediately reduced after treatment and, subsequently, there was a notable reduction of the skin damage. Prednisone and immunosuppressive drugs were gradually reduced until complete suspension. High-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometer was performed for identification of proteins captured by a resin bed during a dialysis session of the patient. This technique identified several biomarkers of kidney injuries, uremic toxins, fragments of immunoglobulins, antigens involved in antiphospholipid syndrome and a new marker (α-defensin) that correlated significantly with disease activity. The removal of these different proteins could possibly provide an explanation of the improvement in the patient's symptoms and the normalization of her SLE. SUPRA coupled with an adsorption may be a promising new technique for the treatment of lupus nephritis.

A novel option for reducing free light chains in myeloma kidney: supra-hemodiafiltration with endogenous reinfusion (HFR).

BACKGROUND: In myeloma cast nephropathy, fast reduction of serum free light chain (FLC) levels correlates with renal recovery. Recently, extracorporeal treatments using filters with a high-molecular weight cut-off have been successfully used for FLC removal. However, using these new filters, high cost and elevated albumin leakage are common drawbacks. We studied a new and cheaper therapeutic approach with adsorbent resins to evaluate its efficacy. METHODS: We treated four patients, affected by dialysis-dependent acute kidney injury (AKI) due to biopsy proven de novo FLC myeloma cast nephropathy. Each patient underwent bortezomib chemotherapy and extracorporeal treatment with the supra-hemodiafiltration with endogenous reinfusion (HFR) technique (Supra-HFR, Bellco Mirandola, Modena, Italy). Supra-HFR is a kind of hemodiafiltration that utilizes separated convection, diffusion and adsorption. The sorbent cartridge has a high affinity for FLC (both κ and λ) but is able to re-infuse albumin, avoiding the need for albumin perfusions. Supra HFR treatments (4 h each) were carried out for eight consecutive days and then every other day. RESULTS: All patients showed a significant reduction of serum FLC, whereas serum albumin concentration remained unchanged. Renal function recovered in three out of four patients. CONCLUSIONS: FLC removal with adsorbent resins represents an effective therapeutic strategy that does not require replacement with albumin .

Proteomic analisys of protein extraction during hemofiltration with on-line endogenous reinfusion (HFR) using different polysulphone membranes.

In end-stage renal disease patients, extracorporeal dialytic therapy is not able to prevent the accumulation of toxins related to the uremic syndrome, a severe complication that increases morbidity and mortality rate. In this paper, hemoFiltration with on-line Reinfusion (HFR) architecture is used to evaluate the effect of a more permeable membrane on the extraction of medium-high molecular weight molecules. The aim of this study was to compare two polysulphone membranes for convective chamber: polyphenylene High Flux (pHF) and polyphenylene Super High-Flux (pSHF). Fourteen patients were subjected to HFR with pHF and pSHF membranes and ultra filtrate (UF) samples were collected to evaluate molecular weight cut-off (MWCO) and to identify extracted proteins. Furthermore, image analysis software was used in order to evaluate change in protein extraction during the dialysis. The quantification of four proteins by immunoassay demonstrates a higher permeability of pSHF membrane. Two-dimensional electrophoresis (2-DE) gels showed, for both membranes, the greater number of protein spots at 235 min. Some of the identified proteins, involved in nephropathic disease complications, were compared to assess differences in extraction during dialytic treatment by PDQuest analysis. UF proteomic analysis demonstrated a different behavior for the two membranes; pHF membrane was more permeable at the beginning of HFR treatment (15 min), while pSHF membrane at the end of treatment (235 min). Proteomic analysis is a suitable approach to investigate the behavior of different membranes during dialysis. Results indicated that pSHF membrane offers the higher permeability, and showed higher efficiency in removal of middle molecules related to uremic syndrome.

Quantification of p-cresol sulphate in human plasma by selected reaction monitoring.

Chronic renal failure patients accumulate in the blood molecules that are normally excreted into the urine. p-Cresol Sulphate (pCS), the most representative retained toxin, shows a high level of toxicity. Therefore, its quantification could represent a prediction factor to determine the risk of endothelial dysfunction and cardiovascular complication and response to the haemodialysis treatment. The aim of this study was to evaluate the suitability of the multiple reaction monitoring (MRM) technique in order to improve the sensibility, the selectivity and the timing of pCS detection in a small amount of plasma. Deproteinized plasma of uremic patients was concentrated and dissolved in liquid chromatography (LC) mobile phase solution. pCS was quantified by LC coupled to tandem mass spectrometry (LC-MS/MS) on a triple-quadrupole mass spectrometer. Selective and sensitive detection of pCS was achieved by selecting the specific parent ion and monitoring two specific fragment ions. The MRM assay was carried out using the following transitions: m/z 187 → 80.00 and m/z 187 → 107.00. A good linearity was observed for each calibration curve. The intra-day and inter-day results showed a good precision and repeatability. The percentage recoveries indicate an optimal selectivity of the analytical method. The MRM assay to quantify pCS in a small amount of human plasma is rapid, highly sensitive, selective and with a good repeatability.

A highly selective, biofunctional surface for molecule/cell sorting.

We report in this paper an approach to the effective capture of IgM antibodies from antisera and solutions based on the formation of a carpet of molecules exposing thiols off a surface. Surfaces of different nature, such as OH-exposing (glass, SiO(2), metal oxides, etc.) and noble metal ones (Au, Ag, etc.), have been first functionalized in the liquid phase by suitable chemistry [3-(mercaptopropyl)trimethoxysilane or 1,4-benzenedimethanethiol]. The resulting exposed SH moieties have been further used for binding anti-A, -B, and -D IgM molecules from goat sera via a thiol exchange reaction involving the J chain and other disulfide bonds present in the IgM molecular structure. Antibodies preserve their functional activity at the surface and appear to be able to bind specifically erythrocytes of the proper group in a fast and reliable way. These results can be generalized to the use of any kind of IgM antibody and can be valuable in surface biofunctionalization in the fields of biosensors and immunoassays.