Middle Molecular Weight Research Articles

Middle-Molecular Weight Toxin Removal: Asymmetric Cellulose Triacetate vs. Helicoidal Polysulfone during Online Hemodiafiltration (CHORALS Study)

Middle-Molecular Weight Toxin Removal: Asymmetric Cellulose Triacetate vs. Helicoidal Polysulfone during Online Hemodiafiltration (CHORALS Study)

Mn-Fe bimetallic oxide catalytic oxidation combined with ultrafiltration for treating secondary effluent: EfOM molecular transformation and membrane fouling control mechanism

Despite extensive application and research of membrane technology in secondary effluent treatment, fouling induced by effluent organic matter (EfOM) remains a challenge. Herein, a pretreatment strategy involving Mn-Fe bimetallic oxide-activated peroxymonosulfate (MnFeO/PMS) was used before ultrafiltration to decompose EfOM and tackle membrane fouling. Results showed that membrane fouling was effectively alleviated, with reductions in reversible and irreversible fouling resistances of 88.3 % and 58.9 %, respectively. Theoretical analyses illustrated a more pronounced adsorption energy and decomposition tendency of PMS on the MnFeO surface. The generated OH and SO4− played a key role in the degradation of EfOM. The parallel factor analysis along with self-organizing maps revealed the preferential oxidative degradation of humic acids and fulvic acids over tryptophan-like substances, which contributed to the reversible fouling mitigation. In addition, MnFeO/PMS transformed high and middle molecular weight organics into low molecular weight compounds. The primary molecular reactions involved in the degradation of EfOM included oxygen addition, dealkylation and deamination, which changed the properties of EfOM and contributed to the mitigation of membrane fouling. Morphology analysis further indicated that the fouling layer on the membrane surface was inhibited when filtering the MnFeO/PMS-pretreated sample. This study offers revelations for integrating oxidation with ultrafiltration in secondary effluent treatment to alleviate EfOM-induced membrane fouling.

The effect of L-cysteine on starch and protein degradation during barley germination.

In order to investigate the impact of L-cysteine (L-Cys) on starch and protein degradation during barley germination. The amylase activities, degradation of macromolecules during germination were determined in this study. Barley was germinated in petri dish for 0 to 5days with different levels of L-Cys (0mM, 2.5mM, 5mM, 10mM). L-Cys addition increased the total limit dextrinase (LD) activities and decreased the LD inhibitor activities during whole germination stage. The activities of α-amylase, β-amylase and free LD were increased with the addition of 2.5, 5mM L-Cys at germination days 1 to 4. Due to higher amylase in malt with the addition of L-Cys, the non-fermentable sugars were reduced and the glucose, maltotriose were improved. Furthermore, the protein degradation analysis showed that low molecular weight protein increased and middle molecular weight protein decreased obviously in wort from the malt germinated with L-Cys, demonstrating that the L-Cys promote the protein degradation. Lastly, the filtration performance of malt with the addition of L-Cys during malting was better than the control. In conclusion, L-Cys can promote the degradation of storage material (starch, protein) during barley germination, leading to a better green malt quality.

#693 Reversed high-flux SUPRA HFR: a proof of concept pilot study to access the middle molecular weight uremic toxins

Abstract Background and Aims Supra HFR is an extracorporeal purification system in which convection, diffusion, and adsorption act separately. Thanks to a first hemofilter with a sieving coefficient for albumin of 0.02, the endogenous ultrafiltrate is purified by a neutral styrenic resin, then the reconstituted whole blood is led to a second low-flux filter. Recently, an evolution of Supra HFR, called Supra H, has been produced with the innovative second high-flux filter (Bellco, Mirandola, Italy, Kuf > 50 ml/h/mmHg). Aim of the study is a comparison between Supra HFR, Supra H and Supra H used in a reversed configuration, called “reversed Supra H”. The potential advantage of reversed Supra H is to achieve a fine tuning on the removal of medium molecular weight toxins through adsorption acting after the bulky tuning of these molecules with the new high-flux filter (Fig. 1). Method A pilot study was conducted on 4 patients in a cross-over manner. Each patient underwent one session of conventional Supra HFR, one session of Supra H, and one session of reversed Supra H. The median age of the patients was 61 (58-79) years, dialysis vintage was 113 (76-434) months, median blood flow from the vascular access was 300 (270-300) ml/min, dialysis time was 240 (225-240) minutes, and the amount of the endogenous ultrafiltrate (Qinf) produced was kept constant for each patient during the 3 different treatments and was 9.6 (9.2-12) liters. The medium molecular weight toxins studied were β2 microglobulin (β2M), myoglobin and free light chains κ and λ (FLC κ and λ). Albumin removal by all three methods was also studied. The removal of the studied substances was obtained by Reduction Rate per session (RRs) = [(dialysis start value-dialysis end value) / dialysis start value] *100. The dialysis end value was obtained by the stop dialysate flow method and corrected for hemoconcentration according to Schneditz D et al. ASAIO J 2012. Friedman's test was used to analyze the difference between the three groups. Dunn's post-hoc test was conducted to find differences between two groups at a time. Results Regarding the toxic substances studied, the dialysis start values were represented in Table 1. As to the β2M RRs the comparison of Supra HFR vs Supra H vs reversed Supra H was statistically significant in favor of using reversed Supra H (P=0.037), the same result was obtained for the RRs of FLCs κ (P=0.041). Dunn's test showed a significant increase in RRs of FLC κ with reversed Supra H compared with Supra HFR (P=0.040). No significant differences were found for the RRs of myoglobin and FLCs λ although reversed Supra H showed a better trend in the purification of these molecules (Fig. 2). RRs of albumin were similar with these three dialysis techniques. Conclusion The Supra H technique allows an increase in the RRs of tested uremic toxins compared to the RRs obtained by Supra HFR in the conventional version. The reversed Supra H configuration confirms the hypothesis of an even better depurative trend than the other two techniques. The significance of the results is certainly affected by the small number of patients enrolled.

#2799 Myoglobin removal by hemadsorption: ex-vivo adsorption kinetics

Abstract Background and Aims Myoglobin, due to its molecular weight of 17 kDa and a Einstein-Stoke radius greater than expected, is classed as a middle molecular weight uremic toxin and presents a two-compartment model of distribution in the body. Rhabdomyolysis is a condition in which muscle cell content including myoglobin, are released into blood circulation due to severe injury to the muscle. Circulating myoglobin induces renal injury with several mechanisms: reduced concentration of nitric oxide in medullary renal arterioles, production of free radicals with oxidative stress, tubular necrosis as a consequence of tubular obstruction by myoglobin and cellular debris. A possible therapeutic option is the use of High Cut-off membranes but with the risk of significant loss of albumin and other proteins. The use of adsorption can overcome these limitations. The aim of our study is to assess the adsorption of myoglobin with neutral microporous styrene-divinylbenzene copolymer sorbent (e.g., HA330/380, Jafron Biomedical CO, Lts, Zhuhai City, China). Method Using a downscaled module of HA380 Cartridge, we set up an ex-vivo circulation experiment in which 1 liter of plasma containing 0.1 mg of myoglobin was pumped through the cartridge. We collected samples (3 mL) at different timepoints to measure Myoglobin concentration. We calculated the removal ratio (RR) and mass adsorbed at a given timepoints. Results Initial myoglobin concentration was 99.3 ng/ml. Final myoglobin concentration after 4 hours of experiment was 30.6 ng/ml. Removal Ratio at 4 hour was 69 % and mass adsorption of 0.068 mg of myoglobin. Conclusion To our knowledge, this is the first experiment that assessed the adsorption capacity of polystyrene-divinylbenzene sorbent. Our study, demonstrated high adsorption capacity in the first 4 hours of treatment therefore, confirming the possibility to use adsorption as an effective treatment option in rhabdomyolysis and avoiding complications associated to current dialytic treatment.

#1351 Assessment of safety and efficacy of expanded hemodialysis in comparison to HDF

Abstract Background and Aims In end-stage kidney disease, solutes with a wide spectrum of molecular weight, called uremic toxins, are retained due to impaired kidney cleansing capacity. Particularly, the retention of toxins in the middle molecular weight range 15–45 kDa might negatively affects multiple organ systems and metabolic pathways. The removal of such large uraemic toxins is still a challenge. HDF has produced some results although large convective volume, optimal vascular access to increase the blood flow rate and strict water quality management are required. Medium Cut-Off, high Retention Onset membrane have been recently developed introducing the concept therapy called “expanded haemodialysis” (HDx). Furthermore, vitamin E-coated membrane has potential beneficial effects on inflammation and oxidative stress. Method A prospective longitudinal single-center study was conducted for 3 months among 18 chronic haemodialysis patients. Patients were randomly assigned into either HDF with high flux membrane or HDx with Theranova or ViE-X membrane. The primary goal was to assess albumin loss among the three types of dialyzers. Secondary goals included assessment of depurative efficacy for uraemic toxins and clinical outcomes. Throughout the treatment, a proportional part of the dialysis fluid was collected to quantify albumin loss. Blood samples were taken before and after dialysis session in order to calculate the Removal Ratio and at two different intradialytic timepoints from arterial and venous line of the circuit in order to calculate instantaneous clearance. Results Mean albumin loss was significantly higher in patients undergoing HDx with Theranova membrane (ANOVA, p<0.001) without any difference in serum albumin concentration among the three groups. Instantaneous clearance of small and middle molecules was significantly higher in patients undergoing HDF, but we didn't find differences in removal ratio. Reduction of Erythropoietin resistance index was observed in patients treated with ViE-X membrane but it was related to their lower dialysis vintage. Conclusion HDx with Theranova membrane was associated with higher albumin loss without any effect on pre-dialysis serum albumin. Moreover, patients treated Theranova membrane, in the presence lower Qb, lower convective dose and lower instantaneous clearance reached same dialysis efficacy compared to HDF.

The unique properties of Big tau in the visual system.

Tau is a microtubule associated protein that plays important roles in regulating the properties of microtubules and axonal transport, as well as tauopathies associated with toxic aggregates leading to neurodegenerative diseases. It is encoded by the MAPT gene forming multiple isoforms (45-60 kDa) by alternative splicing which are developmentally regulated. The high molecular weight (MW) tau isoform of 105 kDa, termed Big tau, was originally discovered in the peripheral nervous system (PNS) but later found in selective CNS areas. It contains an additional large exon 4a generating a long projecting domain of about 250 amino acids. Here we investigated the properties of Big tau in the visual system of rats, its distribution in retinal ganglion cells and the optic nerve as well as its developmental regulation using biochemical, molecular and histological analyses. We discovered that Big tau is expresses as a 95 kDa protein (termed middle MW) containing exons 4a, 6 as well as exon 10 which defines a 4 microtubule-binding repeats (4R). It lacks exons 2/3 but shares the extensive phosphorylation characteristic of other tau isoforms. Importantly, early in development the visual system expresses only the low MW isoform (3R) switching to both the low and middle MW isoforms (4R) in adult retinal ganglion neurons and their corresponding axons. This is a unique structure and expression pattern of Big tau, which we hypothesize is associated with the specific properties of the visual system different from what has been previously described in the PNS and other areas of the nervous system.

Does online high-volume hemodiafiltration offer greater efficiency and sustainability compared with high-flux hemodialysis? A detailed simulation analysis anchored in real-world data.

Recent findings, including the CONVINCE (comparison of high-dose HDF with high-flux HD) study report, suggest the superiority of high-volume hemodiafiltration (HDF) over high-flux hemodialysis (HD) in improving patients' outcomes. Despite positive patient outcomes, concerns have arisen about the potential negative environmental impact of high-volume HDF, as it may lead to increased water and dialysis fluid consumption and higher waste production. In this manuscript, we address the environmental impact of high-volume HDF, focusing on three key factors: water treatment consumption, dialysis fluid consumption, and solute efficiency markers of HD and HDF. By optimizing HDF prescription through adjustments in operational capabilities, while keeping a high blood flow (i.e.,>350ml/min) such as reducing the QD/QB ratio to 1.2 rather than 1.4 or 1.5 and incorporating automated ultrafiltration and substitution control, we demonstrate that HDF delivers a higher dialysis dose for small- and middle-molecule uremic compounds with the same dialysis fluid consumption, and at equal dialysis doses dialysis fluid consumption is reduced. This finding is supported by real-world data from 26031 patients who underwent high-volume postdilution HDF at a reduced dialysis flow (430mL/min) and achieved an effective OCMKt/V of 1.70 (where "OCM" stands for online clearance measurement, "K" represents effective dialysis clearance and "V" denotes total body water measured by multifrequency bioimpedance). In addition, simulation modeling calculations, using blood extraction coefficient, dialysate saturation coefficient and solute clearances with urea (small molecular weight) and β2-microglobulin (middle molecular weight), consistently show the superiority of postdilution HDF to HD. This holds true even with a significant reduction in dialysis flow down to 430mL/min, reflecting QD/QB ratio of 1.2. Postdilution HDF generates high ultrafiltrate flow (up to 35% of blood flow), delivering saturated ultrafiltrate to the lower solute concentration containing effluent dialysate, thus enhancing solute clearance which opens the way to reduce the dialysis flow. In conclusion, our analysis, combining simulation and real-world data, suggests that postdilution HDF could be a more environmentally friendly treatment option compared with conventional HD. Additionally, automated user-friendly functions that minimize dialysis fluid use can further strengthen this environmental benefit while enhancing efficiency.

Sustainable production of multimeric and functional recombinant human adiponectin using genome-edited chickens

BackgroundAdiponectin (ADPN) plays a critical role in endocrine and cardiovascular functions, but traditional production methods, such as Escherichia coli and mammalian systems, have faced challenges in generating sufficiently active middle molecular weight (MMW) and high molecular weight (HMW) forms of recombinant human ADPN (hADPN). In our previous study, we proposed genome-edited chickens as an efficient platform for producing multimeric hADPN. However, the consistency of multimeric hADPN expression in this system across generations had not been further investigated.ResultsIn this study, subsequent generations of ovalbumin (OVA) ADPN knock-in chickens showed stable multimeric hADPN production, yielding ~ 26% HMW ADPN (0.59 mg/mL) per hen. Comparative analysis revealed that egg white (EW)-derived hADPN predominantly consisted of hexameric and HMW forms, similar to serum-derived hADPN. In contrast, hADPN obtained from human embryonic kidney (HEK) 293 and High-Five (Hi-5) cells also exhibited the presence of trimers, indicating variability across different production systems. Furthermore, transcriptional expression analysis of ADPN multimerization-associated endoplasmic reticulum chaperone genes (Ero1-Lα, DsbA-L, ERP44, and PDI) indicated upregulation in the oviduct magnum of ADPN KI hens, suggesting the chicken oviduct magnum as the optimal site for HMW ADPN production. Lastly, the functional analysis demonstrated that EW-derived hADPN significantly reduced lipid droplets and downregulated lipid accumulation-related genes (LOX-1, AT1R, FAS, and FABP4) in human umbilical vein endothelial cells (HUVECs).ConclusionIn summary, stable and functional multimeric hADPN can be produced in genome-edited chickens even after generations. This highlights the potential of using chicken bioreactor for producing various high-value proteins.

Changes in Endogenous Intoxication and Antioxidant Defense System Markers in Patients with Tumor-Induced Acute Colonic Obstruction

A comprehensive evaluation of clinical and biochemical markers defining the status of endotoxemia in patients diagnosed with acute colonic obstruction (ACO) was conducted. The findings revealed that advancing stages of ACO corresponded to an increase in serum levels of endogenous intoxication markers and a simultaneous decrease in antioxidant defense system (ADS) markers.
 The aim of the study was to analyze changes in endogenous intoxication and ADS markers in patients with tumor-induced ACO throughout the disease progression and evaluate the outcomes.
 Materials and Methods. The study is based on examining serum levels of endogenous intoxication markers, including middle weight molecules (MWM), products of lipid peroxidation (LPO) - conjugated dienes (CD), malonaldehyde (MA), protein oxidative modification (POM), and key ADS markers, specifically superoxide dismutase (SOD), catalase (CAT), and ceruloplasmin (CP), in 155 patients with ACO throughout the disease progression and 20 apparently healthy individuals.
 Results. Upon patient admission to the hospital, the duration of ACO was found to correlate with an increase in endogenous intoxication markers and a decrease in antioxidant defense markers. Depending on ACO stage, MWM levels were 0.42 (0.38; 0.43) IU, 0.56 (0.51; 0.58) IU, and 0.72 (0.69; 0.73) IU, respectively (р1<0.01, р2<0.01, р3<0.01), with a normal range of 0.23 (0.18; 0.25) IU. In the compensated stage of ACO, MA levels significantly increased compared to the control group, reaching 6.39 (6.25; 6.42) nmol/mL (р1<0.001, р2<0.01, р3<0.05). Additionally, with the progression of the disease, a gradual increase in MA levels up to 6.39 (6.25; 6.42) nmol/mL, 7.88 (7.62; 7.92) nmol/mL, and 8.11 (8.05; 8.15) nmol/mL, respectively, was noted (р1<0.001, р2<0.01, р3<0.05), with a normal range of 3.14 (2.58; 3.17) nmol/mL. Similar trend was observed in serum CD levels - 3.11 (3.02; 3.18) IU, 3.69 (3.52; 3.72) IU, and 4.69 (4.53; 4.82) IU, respectively, with a normal range of 1.41 (1.32; 1.46) IU (р1<0.01, р2<0.01, р3<0.01). The study of LPO intensity via the optical density of aldehyde and ketone derivatives of neutral and basic dinitrophenylhydrazones (POM 356, POM 370, POM 430, POM 530) revealed an increase in POM product levels with the disease progression compared to the control group (р1<0.001, р2<0.01, р3<0.01). The analysis of the ADS showed an opposite trend, characterized by a significant decrease in marker levels. In patients with different ACO stages, serum CAT levels were 8.08 (7.95; 8.12) Н2О2/mL, 6.32 (6.25;6.41) Н2О2/mL, and 5.67 (5.59; 5.75) Н2О2/mL, respectively, with a normal range of 12.36 (12.21; 12.55) Н2О2/ml (р1<0.001, р2<0.01, р3<0.01), while SOD concentration decreased with the disease progression as well - 42.43 (31.75; 51.84) IU, 35.65 (24.72; 45.53) IU, and 32.18 (18.72; 41.15) IU, respectively, with a normal range of 63.60 (54.41; 74.56) IU (р1<0.001, р2<0.01, р3<0.01). In the compensated stage of ACO, CP level exceeded the normal value - 39.21 (38.92; 39.41) IU, with a normal range of 28.9 (28.4; 29.1) IU, indicating a compensatory defensive response of the body to a dramatic increase in LPO products. CP reserves, however, were depleted with the disease progression and a consistent decrease in this marker was subsequently observed: 19.10 (18.91; 19.23) IU in the subcompensated stage and 16.12 (16.01; 16.15) IU in the decompensated stage (р1<0.001, р2<0.01, р3<0.01).
 Conclusions. As a systemic process that induces various metabolic homeostasis disorders, endogenous intoxication serves as the pathogenetic foundation for the development of multiple organ failure in ACO patients. With ACO progression, endogenous intoxication caused an increase in LPO and POM activity, resulting in elevation of highly toxic middle-molecular weight substances, concurrently accompanied by a decline in the AOS activity.

Assessment of the Influence of Postdilution Online Hemodiafiltration on the Rate of Removal of Middle Molecular Weight Uremic Toxins

Abstract Hemodiafiltration is a method of treatment used to replace kidney function, which effectively removes uremic toxins of middle molecular weight from the blood of patients with the end-stage of chronic kidney disease. The aim of this study was to examine the effect of postdilution online hemodiafiltration on the degree of β2- microglobulin removal. Thirty patients treated with postdilution online hemodiafiltration were examined. The main parameter for assessing the removal efficiency of uremic toxins of middle molecular weight was the concentration of β2-microglobulin in the serum before and after a single session of postdilution online hemodiafiltration. The following tests were used for statistical analysis: Shapiro-Wilk test, Student’s T test for bound samples and Wilcoxon test. The average total convective volume is 21.38 ± 2.97 liters per session. The reduction index of β2-microglobulin during a single session of postdilution online hemodiafiltration is 70.86 ± 6.87%. The average loss of albumin during a single postdilution online hemodiafiltration is 2.50 ± 0.92 g/4h, and the albumin reduction index is 6.20 ± 2.12%. Postdilution online hemodiafiltration effectively removes β2-microglobulin from the blood of patients with end-stage chronic kidney disease. The reduction index of β2-microglobulin is ˜ 71.00% and the loss of albumin is less than 4.0 g/4h. This dialysis modality prevents the development of dialysis-related amyloidosis and atherosclerotic cardiovascular diseases in the population of patients treated with regular hemodiafiltration.

Hemodiafiltration for children with stage 5 chronic kidney disease: technical aspects and outcomes

Despite significant medical and technical improvements in the field of dialysis, the morbidity and mortality among patients with chronic kidney disease (CKD) stage 5 on dialysis remains extremely high. Hemodiafiltration (HDF), a dialysis method that combines the two main principles of hemodialysis (HD) and hemofiltration—diffusion and convection—has had a positive impact on survival when delivered with a high convective dose. Improved outcomes with HDF have been attributed to the following factors: HDF removes middle molecular weight uremic toxins including inflammatory cytokines, increases hemodynamic stability, and reduces inflammation and oxidative stress compared to conventional HD. Two randomized trials in adults have shown improved survival with HDF compared to high-flux HD. A large prospective cohort study in children has shown that HDF attenuated the progression of cardiovascular disease, improved bone turnover and growth, reduced inflammation, and improved blood pressure control compared to conventional HD. Importantly, children on HDF reported fewer headaches, dizziness, and cramps; had increased physical activity; and improved school attendance compared to those on HD. In this educational review, we discuss the technical aspects of HDF and results from pediatric studies, comparing outcomes on HDF vs. conventional HD. Convective volume, the cornerstone of treatment with HDF and a key determinant of outcomes in adult randomized trials, is discussed in detail, including the practical aspects of achieving an optimal convective volume.

Predictive value of serum β2-microglobulin in cardiac valve calcification in maintenance hemodialysis patients.

Cardiac valve calcification (CVC) is associated with adverse cardiovascular events. We studied the risk factors of CVC in maintenance hemodialysis (MHD) patients and the value of serum β2-microglobulin (β2-MG) levels in predicting the incidence of CVC. β2-MG is a middle molecular weight toxin. In recent years, researchers found that elevated blood β2-MG was associated with coronary, thoracic, and abdominal aortic calcifications with significant correlations. β2-MG has been emerging as a strong biomarker for cardiovascular mortality in uremic patients but its role in CVC is not well studied. This study looked specifically at CVC occurrence in relation to β2-MG for MHD patients. Patients who underwent MHD for more than 3 months in the First People's Hospital of Nantong City from November 2012 to November 2019 with complete available data were included in the study. The patients were divided into the CVC group and the non-CVC group. The general information and clinical laboratory indicators of the patients were collected in a retrospective manner. We analyzed the risk factors for developing CVC in MHD patients using binary logistic regression method. Receiver operating characteristic (ROC) curves were used to calculate the cut-off value of β2-MG for predicting CVC. The decision tree (DT) method was used to classify and explore the probability of CVC in patients with MHD. The β2-MG in the CVC group was significantly higher than that in the non-CVC group (t=6.750, P<0.001). Multivariate binary logistic regression analysis showed that gender, age, serum β2-MG, and hemodialysis (HD) adequacy (Kt/V urea) were independent risk factors for CVC in MHD patients. ROC analysis showed that a β2-MG value of 25 µg/L was the best cut-off point for predicting CVC in MHD patients. According to binary logistic regression analysis, the β2-MG ≥25 µg/L group was 3.39 times more likely to develop CVC than the β2-MG <25 µg/L group [odds ratio (OR), 3.39; 95% confidence interval (CI), 1.63-7.06; P=0.001]. The DT model determined that serum β2-MG ≥25 µg/L and age >69 years were important determinants for predicting CVC in MHD patients. Serum β2-MG in MHD patients has a positive correlation with the severity and occurrence of CVC.

Performance analysis and design for an asymmetric membrane of a hollow fiber hemodiafilter with a two-layer membrane model

A hemodiafilter model including the two-layer membrane model based on the irreversible thermodynamics was first introduced, which considers the convective solute flux besides the diffusive solute one. This model makes it possible to analyze the solute transfer of an asymmetric membrane which is assumed to be equivalent to the two-layer membrane consisting of a skin layer and a porous support layer. Verification was performed with in-vitro data on commercial hemodiafilters for solutes of middle molecular weight, and the results were good in solute clearances. In order to increase the clearance of middle molecular weight solutes in high-performance membranes, it is necessary to increase the solute diffusive permeability in the support layer. Through the model simulation, the author was able to quantitatively grasp the relationship between myoglobin clearance versus membrane microstructure parameters and membrane thickness and to clarify the direction of increase in solute transfer amount. In addition, it made clear that the convective solute transfer is more predominant than the diffusive solute one in ultrafiltration effect of the hemodiafilter with the asymmetric membrane.

#3205 A 2-YEAR STUDY FOR CHANGE OF PLASMA BIOMARKERS INCLUDING LARGER MIDDLE MOLECULES IN DIALYTIC PATIENTS USING DIALYZERS WITH MEDIUM CUT-OFF MEMBRANES

Abstract Background and Aims Uremic toxins with middle molecular weight usually develop uremic symptoms. The larger middle molecules are regarded as causative molecules of uremic symptoms. Medium cut-off (MCO) membrane newly developed can remove larger middle molecules which can be not removed through high flux membrane and hemodiafiltration. Larger middle molecules are mainly related to cardiovascular complications. Therefore, long-term use of MCO membrane may have beneficial effects in dialytic patients, in terms of lowering concentrations of larger middle molecules associated with cardiovascular complications. Now, we report interesting 2-year results of long-term use of MCO membrane in dialytic patients. Method Thirty-four patients undergoing hemodialysis were prospectively analyzed during 24 months. We randomly divided enrolled patients into two groups: control and MCO group. Patients in the control group used dialyzer with high flux membrane and patients in MCO group used MCO membrane. We measured plasma levels of biomarkers with 3 or 6-month to investigate efficacy of the MCO membrane for small (3-month interval) to larger middle molecules (6-month interval). Particularly, we performed comparative analysis with larger middle molecules in both groups. We calculated reduction ratio of biomarkers per one session at 24-month. Results The levels of larger middle molecules at baseline did not show significant difference between two groups. However, compared with the control group, plasma sclerostin levels at 6-month and 12-month in MCO group did not increased significantly compared to baseline levels. Plasma sclerostin were lower in MCO group throughout 2 year, but the significant difference of plasma sclerostin levels between two groups unfortunately disappeared after 12 months. The levels of other biomarkers including FGF23, leptin and GDF15, also did not show significant difference between two groups. However, β2MG levels in MCO group compared to control group decreased significantly throughout 2 years (MCO group: 18-month, p = 0.01 and 24-month, p &amp;lt;0.001, respectively; control group: 18-month, p = 0.003 and 24-month, p &amp;lt;0.067, respectively). Interestingly, even in MCO group, there was no statically significant decrease in total protein and albumin levels after 24 months (p = 0.429 and p = 0.101, respectively) (Figure 1). The reduction ratio of larger middle molecules compared to small molecules per one session prominently increased in MCO group (sclerostin, p &amp;lt; 0.001; GDF15, p &amp;lt;0.001; FGF23, p &amp;lt;0.01; RBP4, p &amp;lt;0.01; phosphate, p = 0.821; calcium, p = 0.827; CRP, p = 0.729 and β2MG, p = 0.286, respectively) (Figure 2). Conclusion Conclusively, in MCO group, the significant change of plasma sclerostin associated with cardiovascular complications after 12 months disappeared. However, in MCO group compared to control group, there was a continuous decrease of β2MG and RBP4 throughout study period. In addition, the clearance of larger middle molecules is also prominently high in MCO group. Interestingly, there were also no significant differences of albumin between 24-month and baseline in MCO group. Therefore, the MCO membrane can be a good tool for controlling uremic symptoms or sings without protein loss even when used for a relatively long time in dialytic patients.

#3595 DIALYSIS MODALITY AFFECTS SENESCENCE-RELATED PHENOTYPE OF T AND B LYMPHOCYTES

Abstract Background and Aims Hemodiafiltration (HDF) is characterized by enhanced clearance of middle molecular weight toxins and therefore, it is considered superior to hemodialysis (HD), especially in higher hemofiltration volumes. Although proinflammatory cytokines are lower in HDF, its impact on immune phenotype has not been studied, so far. In this study we compared the lymphocytes’ senescent phenotype between HD and HDF patients. Method Senescent and exhausted related lymphocyte markers, including CD45RA, CCR7, CD28, CD57 and PD1 on T cells and CD27 and IgD on B cells, were assessed by flow cytometry in 35 ESRD patients on HD and 25 ESRD patients on HDF of similar age, sex, and ethnicity. Moreover, we evaluated the association of these markers with hemofiltration volume. Results Patients on HDF had significantly reduced CD4+CD28- T lymphocytes, 31(18-74) vs 50(36-131)cells/μL, respectively, p = 0.019, which was restricted only in the CD28-CD57- T cells compartment, [17(9-25) vs 31(19-50)cells/μL, p = 0,002], while terminally differentiated CD28-CD57+ did not differ between the two groups [14(4-49) vs 23(7-52)cells/μL, p = 0,22]. Expression of CD57 molecule on CD4+ T cells depended on dialysis vintage (r = 0.27, p = 0.03). Moreover, HDF patients had lower percentage of CD4+PD1+ exhausted T cells [8.5(6.8-11.7) vs 13.7(8.7-19.5) % for HDF and HD respectively, p = 0.006]. CD8+PD1+ T cells were also lower in HDF patients, without reaching statistical significance [17.4(8.5-41) vs 34(12,7-52.9) % for HDF and HD respectively, p = 0.06]. Hemofiltration volume was negatively correlated to effector memory CD8+ T cells re-expressing CD45RA count (CD8+ EMRA), (r = -0.46, p = 0.027) and positively correlated to total B cells count (r = 0.46, p = 0.025). However, not all B cell subsets were equally affected, with only naïve IgD+CD27- B cells and switched memory IgD-CD27+ B cells count showing positive correlation with hemofiltration volume (r = 0.53, p = 0.008 and r = 0.5, p = 0.015, respectively). Conclusion Hemodiafiltration and high filtration volumes may have beneficial effects on adaptive immunity, predisposing to less senescent and exhausted T and B cell phenotypes.

Evaluation of Function and Features of Human Induced Pluripotent Stem Cell-Derived Small Intestinal Epithelial Cells for Analyzing Peptide Drug Intestinal Absorption Profiles

Caco-2 cell monolayers are widely employed as an in vitro model of the intestinal barrier, capable of accurately predicting the absorption of conventional small-molecule drugs. However, this model may not be applicable to all drugs, and the accuracy of absorption prediction is typically poor for high molecular weight drugs. Recently, human induced pluripotent stem (iPS) cell-derived small intestinal epithelial cells (hiPSC-SIECs), exhibiting properties similar to those of the small intestine when compared with Caco-2 cells, have been developed and are considered a novel candidate model for in vitro evaluation of intestinal drug permeability. Therefore, we evaluated the utility of human hiPSC-SIECs as a new in vitro model to predict the intestinal absorption of middle-molecular weight drugs and peptide drugs. Firstly, we showed that the hiPSC-SIEC monolayer allowed faster transport of peptide drugs (insulin and glucagon-like peptide-1) than the Caco- 2 cell monolayer. Second, we revealed that hiPSC-SIECs require divalent cations (Mg2+ and Ca2+) to maintain barrier integrity. Third, we demonstrated that experimental conditions established for Caco-2 cells are not persistently applicable to hiPSC-SICEs when analyzing absorption enhancers. Comprehensively clarifying the features of hiPSC-SICEs is essential to establish a new in vitro evaluation model.

Effects of Intradermal Administration Volume Using a Hollow Microneedle on the Pharmacokinetics of Fluorescein Isothiocyanate Dextran (M.W. 4,000).

Hollow microneedles (hMNs) have been gaining attention as a tool to enable the intradermal (i.d.) administration of pharmaceutical products. However, few reports have examined the effect of administration volume on distribution in the skin and pharmacokinetics parameters after i.d. injection. In the present study, a model middle molecular weight compound, fluorescein isothiocyanate dextran (M.W. 4,000, FD-4), was selected, and blood concentration-time profiles after i.d. and subcutaneous (s.c.) injections with different administration volumes were compared. FD-4 solution was injected i.d. using a hMN or injected s.c. with a 27 G needle. Pharmacokinetics and dermatokinetics of FD-4 were analyzed using a compartment model. The skin distribution of iodine, as an X ray tracer, was used to evaluate drug disposition. With the administered drug assumed to be absorbed from the broad injection site into blood vessels in the upper and lower dermis by rapid (krapid) and slow (kslow) first-order absorption rate constants, respectively, better agreement of observed and theoretical values was obtained. Furthermore, the fraction, F, of the administered dose absorbed with krapid decreased with the increase in injection volume after i.d. injection, although the pharmacokinetics parameters were almost the same regardless of administration volume after s.c. injection. The drug distribution in the skin may be related to the obtained pharmacokinetics parameters suggested that the number of needles in the MN system and the total administration volume should be considered in designing hMN systems. The present results provide useful information that may support effective drug delivery with hMNs.

On the balance between albumin loss and removal of middle molecules in dialyzers

Tuning the pore size distribution of hemodialysis membranes is essential for the membrane’s selectivity and significantly affects the quality of the dialysis treatment. Tailoring the membrane’s molecular weight cut-off appropriately balances the removal of middle-molecular-weight uremic toxins while avoiding albumin loss. This undesirable albumin loss is a potential side effect and concern for clinical use when aiming at increased removal of middle molecular weight molecules (middle molecules). It is hypothesized that control of the position of a narrow pore size distribution allows middle molecule removal while simultaneously counteracting the unwanted albumin loss. This study presents a comprehensive ex-vivo methodology and novel data on the balance of the clearance of middle molecules and albumin loss at different blood and dialysate flow rates using novel dialyzers. The outcomes hold significance for dialysis therapy, while the insights acquired have broader implications for the selectivity of ultrafiltration membranes.The Theranova dialyzer shows the largest clearance for small-middle molecules. Phylther stands out with higher removal of the middle molecule YKL-40 than the other dialyzers but exhibits a significant albumin loss. Theranova demonstrates the best compromise between low albumin loss and good clearances of middle molecules.

An extended kinetic model-based correction factor equation to account hemodialysis post-treatment hemoconcentration.

The hemoconcentration effect for middle weight solutes in hemodialysis is corrected by oversimplified methods based on hematocrit changes or distribution volume variations. Here we implemented a variable volume dual pool kinetic model targeted at obtaining a precise correction factor equation for extracellularly distributed solutes based on relevant kinetic parameters such as the ultrafiltration to dry weight ratio UF/DW, the dialyzer clearance, Kd, the intercompartment mass-transfer coefficient, Kc, and the central compartment to extracellular volume ratio, α. More than 300,000 solutions of the model were computed, performing a sweep among physiological values of the proposed kinetic parameters, resulting in a linear regression denoted by the expression fcorr = 1.0707 - 5.2246 (UF/DW) - 0.0005 Kd - 0.0004 Kc - 0.0007 α, with an excellent coefficient of determination R2 = 0.983. The presented fcorr provides a substantial extension of the currently implemented methods to estimate the hemoconcentration factor for middle and high molecular weight extracellular distributed solutes in hemodialysis.