Uremic Solutes Research Articles

Protein-Bound Uremic Solute 3-Carboxy-4-Methyl-5-Propyl-2-Furanpropanoic Acid (CMPF) Induces Eryptosis through a Piezo1-Dependent Pathway

Protein-Bound Uremic Solute 3-Carboxy-4-Methyl-5-Propyl-2-Furanpropanoic Acid (CMPF) Induces Eryptosis through a Piezo1-Dependent Pathway

Assessment of Sodium Butyrate’s Pharmacological Effects on Gentamicin-induced Nephrotic Disorders in a Rat Model

Background Sodium butyrate (SB) used to increase oxidative stress was reduced and the glutathione redox pathway was boosted. It inhibits proliferation, induction of differentiation, and induction or repression of gene expression and it has also anti-inflammatory action. This is endorsed for suppression of nephrotic symptoms. Objectives The present study was carried out to evaluate the nephroprotective activity of SB on gentamicin-induced nephrotoxicity in rats. Materials and Methods Present research work was designed to estimate pharmacological potential of SB in gentamicin-induced nephrotic disorder. The pharmacological prospective was estimated in gentamicin-induced rats through the oral administration of SB formulation 200 mg/kg and 300 mg/kg BW. Results The study observed that the SB significantly protects the kidneys from gentamicin-induced nephrotic disorders. Gentamicin 100 mg/kg for 7 days was used for induction of nephrotoxicity in Wistar rats by intraperitoneal administration. The administration of SB was observed to decrease the occurrence of glomerular congestion, vascular congestion, epithelial desquamation, inflammatory cell build-up, and kidney cell necrosis induced by gentamicin. The dose of SB maintained the gentamicin-induced increased creatinine (0.62±0.05), serum urea (18.10±0.79), uric acid (2.10±0.28), and blood urea nitrogen (BUN) (17.56±0.55) levels. This fact is further sustained by the histopathological examinations. Conclusion Sodium butyrate formulation with antinephrotic activity in maintaining uremic solutes such as urea, uric acid, creatinine, BUN, ketone, urobilinogen, glucose, specific gravity, and urine pH in Wistar rats with gentamicin-induced glomerulonephritis (GN). The SB was administered to the animals at doses of 200 and 300 mg/kg body weight. Pretreatment with the drug showed promising results in maintaining elevated parameters in nephritic rats within a short duration.

Evaluation and Management of Kidney Dysfunction in Advanced Heart Failure: A Scientific Statement From the American Heart Association.

Early identification of kidney dysfunction in patients with advanced heart failure is crucial for timely interventions. In addition to elevations in serum creatinine, kidney dysfunction encompasses inadequate maintenance of sodium and volume homeostasis, retention of uremic solutes, and disrupted endocrine functions. Hemodynamic derangements and maladaptive neurohormonal upregulations contribute to fluctuations in kidney indices and electrolytes that may recover with guideline-directed medical therapy. Quantifying the extent of underlying irreversible intrinsic kidney disease is crucial in predicting whether optimization of congestion and guideline-directed medical therapy can stabilize kidney function. This scientific statement focuses on clinical management of patients experiencing kidney dysfunction through the trajectory of advanced heart failure, with specific focus on (1) the conceptual framework for appropriate evaluation of kidney dysfunction within the context of clinical trajectories in advanced heart failure, including in the consideration of advanced heart failure therapies; (2) preoperative, perioperative, and postoperative approaches to evaluation and management of kidney disease for advanced surgical therapies (durable left ventricular assist device/heart transplantation) and kidney replacement therapies; and (3) the key concepts in palliative care and decision-making processes unique to individuals with concomitant advanced heart failure and kidney disease.

Twice Weekly versus Thrice Weekly Hemodialysis-A Pilot Cross-Over Equivalence Trial.

The 2015 Update of the Kidney Disease Outcomes Quality Initiative (KDOQI) Guideline for Hemodialysis Adequacy increased the contribution of residual kidney function in calculating standard Kt/Vurea (stdKt/Vurea). However, no study has assessed the effect of prescribing twice weekly hemodialysis according to this guideline on patients' quality of life or uremic solute levels. Twenty six hemodialysis patients with average residual urea clearance (Kru) 4.7±1.8 ml/min and hemodialysis vintage of 12±15 months (range two months to 4.9 years) underwent a cross-over trial comparing four weeks of twice weekly hemodialysis and four weeks of thrice weekly hemodialysis. Twice weekly hemodialysis was prescribed to achieve stdKt/Vurea 2.2 incorporating Kru using the 2015 KDOQI Guideline. Thrice weekly hemodialysis was prescribed to achieve spKt/Vurea 1.3 regardless of Kru. Quality of life and plasma levels of secreted uremic solutes and β2 microglobulin (β2m) were assessed at the end of each period. Equivalence testing between twice and thrice weekly hemodialysis based on the Kidney Disease Quality of Life instrument (primary analysis) was inconclusive. Symptoms as assessed by the secondary outcomes Dialysis Symptom Index and post-dialysis recovery time were not worse with twice weekly hemodialysis. StdKt/Vurea was adequate during twice weekly HD (2.7±0.5), and ultrafiltration rate and plasma potassium were controlled with minimally longer treatment times (twice weekly: 195±20 vs. thrice weekly: 191±17 minutes). Plasma levels of the secreted solutes and β2m were not higher with twice weekly than thrice weekly hemodialysis. Twice weekly hemodialysis can be prescribed using the higher contribution assigned to Kru by the 2015 KDOQI Guideline. With twice weekly hemodialysis, quality of life was unchanged, and the continuous function of the residual kidneys controlled fluid gain and plasma levels of potassium and uremic solutes without substantially longer treatment times.

Allo-Hemodialysis, a Novel Dialytic Treatment Option for Patients with Kidney Failure: Outcomes of Mathematical Modelling, Prototyping, and Ex Vivo Testing.

It has been estimated that in 2010, over two million patients with end-stage kidney disease may have faced premature death due to a lack of access to affordable renal replacement therapy, mostly dialysis. To address this shortfall in dialytic kidney replacement therapy, we propose a novel, cost-effective, and low-complexity hemodialysis method called allo-hemodialysis (alloHD). With alloHD, instead of conventional hemodialysis, the blood of a patient with kidney failure flows through the dialyzer's dialysate compartment counter-currently to the blood of a healthy subject (referred to as a "buddy") flowing through the blood compartment. Along the concentration and hydrostatic pressure gradients, uremic solutes and excess fluid are transferred from the patient to the buddy and subsequently excreted by the healthy kidneys of the buddy. We developed a mathematical model of alloHD to systematically explore dialysis adequacy in terms of weekly standard urea Kt/V. We showed that in the case of an anuric child (20 kg), four 4 h alloHD sessions are sufficient to attain a weekly standard Kt/V of >2.0. In the case of an anuric adult patient (70 kg), six 4 h alloHD sessions are necessary. As a next step, we designed and built an alloHD machine prototype that comprises off-the-shelf components. We then used this prototype to perform ex vivo experiments to investigate the transport of solutes, including urea, creatinine, and protein-bound uremic retention products, and to quantitate the accuracy and precision of the machine's ultrafiltration control. These experiments showed that alloHD performed as expected, encouraging future in vivo studies in animals with and without kidney failure.

#598 Successful application of allo-hemodialysis in a porcine model of embolization-induced acute kidney injury

Abstract Background and Aims Annually, millions of kidney patients, primarily in low- and low-middle income nations, die prematurely due to the lack of affordable kidney replacement therapy. We propose a novel, low-cost alternative dialysis modality called allo-hemodialysis (alloHD), where blood of a patient with kidney failure flows counter-current to the blood of a healthy subject (“buddy”) through the dialyzer [1]. Uremic solutes and excess fluid are then excreted by the buddy's healthy kidneys. We have already demonstrated the technical feasibility of alloHD in pigs with acute kidney injury (AKI) induced by renal artery ligation [2]. In this study, we explored alloHD in a less invasive porcine model of AKI induced by renal artery embolization. Method The protocol was approved by the Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA), New Delhi, India. We studied two female Yorkshire pigs (weight 80-90 kg) with central venous catheter as vascular access. AKI was induced by embolization of both kidneys by transcatheter deployment of Contour™ Poly Vinyl Alcohol particles (Boston Scientific, Marlborough, MA, USA). The AKI pig and the buddy pig were connected to the dialysate and blood compartments, respectively, of a Nipro Cellentia 17H dialyzer. The AKI pig was then dialyzed for 4 hours against the healthy buddy pig. During alloHD, both pigs were anticoagulated with heparin (5,000 IU/h) and the buddy pig was sedated with 1-2% isoflurane gas. Blood samples were collected before, during, and after alloHD for measurements of blood urea nitrogen (BUN) and creatinine. Results We performed nine alternate day alloHD sessions with the same pair of pigs. During alloHD, BUN and Creatinine levels declined in the AKI pig (Fig. 1) and increased—as expected—transiently above baseline in the buddy pig and then trended towards baseline post-dialysis (Fig. 1). We found no indication of hemolysis or coagulation of the extracorporeal circuit in any of the nine alloHD sessions. The animals were sacrificed after the 9th session. Conclusion We demonstrated the feasibility of alloHD in a porcine embolization AKI model. This study corroborates and complements our previous experience with alloHD in a porcine model of AKI induced by renal artery ligation. Given these promising results, we are investigating the potential for advancing to a clinical trial.

#1094 Hemodiafiltration improves solute clearance without increasing water consumption: an educational bench model for the dialysis ward

Abstract Background and Aims Recently, the positive results of the CONVICE trial have raised the question whether the improved outcome of postdilution high volume HDF (oHDF-p) compared to high flux HD (hfHD) requires increased water consumption. From mathematical modelling it can be shown that oHDF-p always leads to higher low molecular clearance than hfHD at equal total dialysate consumption Qd_tot = Qd_dial + Qs (Qd_dial: dialysate flow through the dialyzer, Qs: substitution flow). This has also been shown in a clinical trial (Mesic et al, HDI 15, p. 522-529). However, mathematical modelling might not be intuitive for everyone, and clinical trials are always influenced by multiple confounders. To add a more intuitive way of reasoning, a simple educational bench model was designed based only on equipment easily available in a standard dialysis clinic which allows to compare dialysis efficiencies based on the urea concentrations as surrogate parameter. Since the fundamental task of dialysis is to cleanse blood from uremic solutes, the treatment modality which at equal resource utilization reduces solute concentration most is the more efficient one! Method Dialysis treatments were simulated in single pass setup using a Fresenius 5008S machine (Fig. 1). Blood inlet was simulated as a fixed urea concentration of 170 mg/dL in dialysate from a 110 L reservoir “cbi”. Treatment parameters were set such that they correspond to effective values in in-vivo treatments: As the mass transfer coefficient is always much higher in vitro with water than in vivo with blood, a small high-flux dialyzer (Fresenius FX40) was used. Blood flow Qb was set to 300 mL/min, corresponding to the in vivo blood water flow for Qb_set ∼ 350 mL/min. Alternating experiments in oHDF-p and hfHD were carried out in triplicate with Qd_tot=400 mL/min and 300 mL/min. This resulted in the settings HDF400, HD400, HDF300 and HD300 (cf. Fig. 1). Qs was set to 100 ml/min in oHDF-p. In each setting the fluid from blood and dialysate outlet were collected (recipients “cbo” and “cdo”, resp.) for 8 min. To be independent of the machine accuracy of set flow rates, sample volumes were measured by weighing, and effective flow rates of blood Qb_eff and total dialysate Qd_tot_eff were calculated. Samples were taken into monovettes from “cbi”, “cbo” and “cdo”. Urea concentrations cbi, cbo and cdo were determined by sending the samples to two commercial clinical labs who delivered consistent results. Blood and dialysate side urea clearances were calculated as KUB= Qb_eff*(cbi-cbo)/cbi and KUD=Qd_tot_eff*cdo/cbi. Results The table in Fig. 1 includes the measure values of Qb_eff and Qd_tot_eff for each setting. Blood and dialysate outlet concentrations (cbo and cdo, resp.) and calculated blood and dialysate side clearances are shown in Fig. 2. Lab results in repeated settings were highly reproducible with a SD < 2 mg/dL. Switching from hfHD to oHDF-p at equal Qd_tot always decreased cbo, corresponding to a higher clearance in oHDF-p. Reducing total dialysate flow from 400 to 300 ml/min caused a slight decrease in clearance in both modalities. KUB and KUD were in good agreement, showing that urea mass balance was well controlled in the experiment. Conclusion The educational bench model demonstrates the superior low molecular clearance of oHDF-p compared to hfHD at equal Qd_tot, reflected by lower urea cbo values in oHDF-p. Although the clinical outcome advantage of oHDF-p is rather related to middle molecular clearance, the model shows that the improved treatment quality of oHDF-p can be provided with no extra resource utilization compared to hfHD, and that dialysis adequacy as measured by urea clearance is even increased despite of the fact that a part of Qd_tot is deviated as substitution fluid. As moderately reducing Qd_tot in many cases only results in acceptable clearance reductions, the automated adjustment of both dialysate and substitution flow rates for oHDF-p could improve patient outcome at even reduced resource utilization.

#3023 Dietary protein intake modulates the tubular handling of the uremic retention solute indoxyl sulfate

Abstract Background and Aims Nutritional intervention constitutes a key part of the clinical management of individuals with chronic kidney disease (CKD). The gut emerged as an important source of uremic retention solutes, including indoxyl sulfate, p-cresyl sulfate and TMAO among the others. Indoxyl sulfate (IS) is produced through the gut microbial fermentation of tryptophan into indole and its conversion into indoxyl sulfate in the liver. Previous authors hypothesised that higher amounts of gut microbial metabolites can be sensed by kidney's proximal tubule cells through the use of receptors and signaling pathways. Particular evidence points to the involvement of kidney Organic Anion Transporters’(OATs') in controlling the gut-kidney communication. According to the so-called remote sensing hypothesis, kidney tubular cells respond to uremic toxins plasma fluctuations altering the activity of OAT1. Few studies have investigated whether in CKD the protein content in the diet affects gut generation, plasma retention and excretion mechanisms of indolic uremic toxins including kidney cells OAT1 expression. Method Eighteen male Sprague-Dawley rats (Janvier, France) aged 7-8 weeks and weighing between 270 and 388 g were randomly assigned to a low protein (LP) diet (n = 10) or a high protein (HP) diet (n = 8) after undergoing a 5/6 nephrectomy to develop CKD. For each diet, a sham-operated control group (n = 7 and n = 8 correspondingly) was used. After 7 weeks, urine was collected, and 8 weeks after the disease was induced, euthanasia was performed. Samples of kidney and blood were taken. mRNA and protein expression of OAT1 were sought via qPCR and Western blot. Forty patients with CKD were recruited in a cross-sectional study investigating eating habits and uremic toxins levels. Eating habits were searched via a food diary. Protein and energy intake were therefore calculated. Blood and 48 h-urine were collected. Blood creatinine and urea were determined with standard laboratory techniques. Total plasmatic and urinary IS concentrations were measured using LC-MS/MS. The fractional excretion (FE) i.e. the percentage of IS excreted relatively to the kidney filtered load, was calculated to assess IS remote sensing. Results Rats with chronic kidney disease (CKD) had considerably higher plasmatic levels of IS (p < .001) than sham rats. Rats with CKD on an HP diet, however, did not differ in plasma IS from rats fed an LP diet (p = .63). On the other hand, CKD rats fed an HP diet showed a substantial rise in 24-hour urinary IS (p < .001). The FE of IS was significantly higher (p = .005) in CKD rats on a HP diet and correlated with 24 h-urinary IS (Spearman r = .51, p value=.03) and with protein intake (Spearman r = .52, p value=.03) in CKD rats. OAT1 mRNA and protein expression was significantly higher in CKD rats given a HP diet (p = .025 and p = .009, respectively). By dividing the CKD patients into two groups by the median of protein intake, the FE of IS was significantly higher (p = .018) in the patients consuming a larger portion of proteins. Conclusion A diet higher in protein content in 5/6 nephrectomized CKD rats as well as in CKD patients likely leads to an increased production of indole in the colon. This results in increased IS fractional excretion but unchanged IS plasma retention. These findings offer more evidence for the mechanisms behind the remote sensing and signaling of uremic toxins derived from indole. In summary, a low-protein diet is still advised for people with chronic kidney disease.

Prevalence of dysmagnesemia among CKD patients in North India

Prevalence of dysmagnesemia among CKD patients in North India

Post-translational modifications in kidney diseases and associated cardiovascular risk.

Patients with chronic kidney disease (CKD) are at an increased cardiovascular risk compared with the general population, which is driven, at least in part, by mechanisms that are uniquely associated with kidney disease. In CKD, increased levels of oxidative stress and uraemic retention solutes, including urea and advanced glycation end products, enhance non-enzymatic post-translational modification events, such as protein oxidation, glycation, carbamylation and guanidinylation. Alterations in enzymatic post-translational modifications such as glycosylation, ubiquitination, acetylation and methylation are also detected in CKD. Post-translational modifications can alter the structure and function of proteins and lipoprotein particles, thereby affecting cellular processes. In CKD, evidence suggests that post-translationally modified proteins can contribute to inflammation, oxidative stress and fibrosis,and induce vascular damage or prothrombotic effects, which might contribute to CKD progression and/or increase cardiovascular risk in patients with CKD. Consequently, post-translational protein modifications prevalent in CKD might be useful as diagnostic biomarkers and indicators of disease activity that could be used to guide and evaluate therapeutic interventions, in addition to providing potential novel therapeutic targets.

Fabrication and characterization of TiO2‐hydroxyapatite composite‐loaded Polysulfone membranes with integrated biocompatibility for dialysis application

AbstractBackgroundThe use of Polysulfone (PSf) as membrane material for dialysis therapy is becoming more common. However, the PSf membrane can cause the adsorption of blood proteins followed by the adhesion of platelets, destruction of red blood cells, and thrombus formation. One of the biggest challenges in creating membranes for hemodialysis (HD) applications is ensuring biocompatibility. Therefore, to maintain the biocompatibility of PSf membranes, further modifications are necessary.MethodsIn this study, the performance and biocompatibility of PSf membranes were improved by the impregnation of TiO2‐hydroxyapatite composite as an additive. The TiO2‐hydroxyapatite composite was first synthesized via sintering at 1200°C, and then a dope solution composed of pristine PSf polymer solution in N‐Methyl‐2‐pyrrolidone solvent (NMP) and different wt% ratios of the synthesized composite were prepared, and resultant membranes were fabricated using a film applicator via a phase inversion scheme using water as a non‐solvent exchange medium. The synthesized composite was characterized using SEM, EDX, and XRD techniques.ResultsThe FTIR analysis confirmed the weak interaction of PSf polymer with the hydroxyapatite moiety of the composite. The interaction of the synthesized composite within the membrane matrix was evaluated via leaching ratio results. The hydrophilicity, pore profile, and pure water permeation flux rates were also determined. The results of the biocompatibility study showed that membranes based on TiO2‐hydroxyapatite composite had reduced adsorption of BSA protein by 34.41% as well as less than 5% hemolysis ratio. Additionally, the study displayed thrombus formation ranging from 2.08% to 8.69%, respectively. The dialysis results indicated that compared with the original PSf membrane, the TiO2‐hydroxyapatite composite showed high clearance rates for urea (73.81%) and creatinine (72.13%) solutes, respectively.ConclusionTherefore, blending TiO2‐hydroxyapatite composite in the PSf membrane significantly improved the biocompatibility and removal ability of uremic solutes.

Mixed matrix membranes with incorporated glycosaminoglycans have good blood biocompatibility combined to high toxin removal

Most patients in need of renal replacement therapy use peritoneal or hemodialysis (HD) therapy. Major drawbacks of HD are the incomplete removal of uremic solutes (especially middle-sized uremic solutes and protein-bound uremic solutes, PBUTs) as well as the non-continuous therapy (3 times per week for 4 h), causing large fluctuations in water balance and uremic waste, potassium, and phosphate. For achieving better patient outcomes, more continuous therapies are required including the application of home HD or portable HD. The latter require membranes with excellent long-term biocompatibility. Heparin-modified dialysis membranes can offer improved biocompatibility, but systemic anticoagulation is still needed, whereas potential heparin-induced thrombocytopenia (HIT) may occur.Glycosaminoglycans (GAGs) contribute to the kidney glomerular filtration barrier and provide natural anti-fouling properties. Here, we hypothesized that the incorporation of GAGs within polymeric membranes, either via membrane coating, post membrane fabrication, or via blending GAGs into the membrane forming polymer matrix, would provide membranes with improved hemocompatibility. We implemented these strategies to mixed matrix membranes (MMMs) which combine diffusion and adsorption for removing a broad range of uremic solutes. Firstly, we fabricated flat sheet MMMs with various GAG sources, including heparan sulphate from bovine kidney (HSBK), heparinase III-digested HSBK, HS isolated from cultured glomerular endothelial glycocalyx, GAGs from porcine intestinal mucosa (danaparoid, DA), or heparin. The flat sheet MMMs with DA blended within the membrane selective layer have excellent blood compatibility, based on a panel of anti-coagulation and immune activation assays, combined to high water transport and very low albumin leakage. Besides, these MMMs had low clot formation, did not activate immune cells or the complement system and had low platelet adhesion. Based on these findings, we also developed first hollow fiber MMMs with DA blended in the selective layer, which also had high water transport, no leakage of protein, and achieved very good removal of various uremic solutes, comparable to commercial HD membranes.

Effect of time-dependent dialysate bicarbonate concentrations on acid–base and uremic solute kinetics during hemodialysis treatments

Recent studies have suggested benefits for time-dependent dialysate bicarbonate concentrations (Dbic) during hemodialysis (HD). In this clinical trial, we compared for the first time in the same HD patients the effects of time-dependent changes with constant Dbic on acid–base and uremic solute kinetics. Blood acid–base and uremic solute concentration were measured in twenty chronic HD patients during 4-h treatments with A) constant Dbic of 35 mmol/L; B) Dbic of 35 mmol/L then 30 mmol/L; and C) Dbic of 30 mmol/L then 35 mmol/L (change of Dbic after two hours during Treatments B and C). Arterial blood samples were obtained predialysis, every hour during HD and one hour after HD, during second and third treatments of the week with each Dbic concentration profile. Blood bicarbonate concentration (blood [HCO3]) during Treatment C was lower only during the first three HD hours than in Treatment A. Overall blood [HCO3] was reduced during Treatment B in comparison to Treatment A at each time points. We conclude that a single change Dbic in the middle of HD can alter the rate of change in blood [HCO3] and pH during HD; time-dependent Dbic had no influence on uremic solute kinetics.

Associations of Indoxyl Sulfate and p-cresyl Sulfate with Serum Uncarboxylated Matrix γ-carboxyglutamate Protein in Chronic Kidney Disease Patients.

Indoxyl sulfate (IS) and p-cresyl sulfate (PCS) are two important protein-bound uremic retention solutes. Increased serum levels of IS and PCS are associated with cardiovascular calcification. Matrix γ-carboxyglutamate protein (MGP) is a potent inhibitor of vascular calcification and inactivated uncarboxylated MGP (ucMGP) is related to vascular calcification. Nevertheless, whether serum levels of IS and PCS are associated with the serum ucMGP level in chronic kidney disease (CKD) patients with different stages is unknown. This cross-sectional study enrolled 90 patients in different stages of chronic kidney disease. Serum levels of IS and PCS were determined. The serum concentration of ucMGP was measured with an enzyme-linked immunosorbent assay. Independent associations between serum total IS and PCS with ucMGP were evaluated. The mean serum level of ucMGP in participants of this study is 10.78±5.22 μg/mL. Serum levels of the two above-mentioned uremic toxins and ucMGP were elevated commensurately with deteriorating renal function. The serum level of ucMPG was associated with total IS (r = 0.456, p < 0.001) and total PCS (r =0.413, p < 0.001) levels. Multiple linear regression analysis showed that ucMGP was significantly related to levels of IS (β = 0.442, p <0.001), but not the level of PCS concentrations after adjusting for other confounding variables. Our study showed that a higher serum IS level was independently associated with ucMGP in deteriorating CKD. Therefore, it would be worthwhile to investigate the effect of IS on ucMGP in the pathogenesis of vascular calcification in future studies.

Donor-Derived Engineered Microvessels for Cardiovascular Risk Stratification of Patients with Kidney Failure.

Cardiovascular disease is the cause of death in ≈50% of hemodialysis patients. Accumulation of uremic solutes in systemic circulation is thought to be a key driver of the endothelial dysfunction that underlies elevated cardiovascular events. A challenge in understanding the mechanisms relating chronic kidney disease to cardiovascular disease is the lack of in vitro models that allow screening of the effects of the uremic environment on the endothelium. Here, a method is described for microfabrication of human blood vessels from donor cells and perfused with donor serum. The resulting donor-derived microvessels are used to quantify vascular permeability, a hallmark of endothelial dysfunction, in response to serum spiked with pathophysiological levels of indoxyl sulfate, and in response to serum from patients with chronic kidney disease and from uremic pigs. The uremic environment has pronounced effects on microvascular integrity as demonstrated by irregular cell-cell junctions and increased permeability in comparison to cell culture media and healthy serum. Moreover, the engineered microvessels demonstrate an increase in sensitivity compared to traditional 2D assays. Thus, the devices and the methods presented here have the potential to be utilized to risk stratify and to direct personalized treatments for patients with chronic kidney disease.

In vitro Removal of Protein-Bound Retention Solutes by Extracorporeal Blood Purification Procedures

Introduction: When the kidneys or liver fail, toxic metabolites accumulate in the patient's blood, causing cardiovascular and neurotoxic complications and increased mortality. Conventional membrane-based extracorporeal blood purification procedures cannot remove these toxins efficiently. The aim of this in vitro study was to determine whether commercial hemoperfusion adsorbers are suitable for removing protein-bound retention solutes from human plasma and whole blood as well as to compare the removal to conventional hemodialysis. Methods: For in vitro testing of the removal of protein-bound substances, whole blood and plasma were spiked with uremic retention solutes (homocysteine, hippuric acid, indoxyl sulfate, 3-carboxy-4-methyl-5-propyl-2-furanpropionic acid) and the toxins of liver failure (bilirubin, cholic acid, tryptophan, phenol). Subsequently, the protein binding of each retention solute was determined. The adsorption characteristics of the hemoperfusion adsorbers, Jafron HA and Biosky MG, both approved for the adsorption of protein-bound uremic retention solutes and Cytosorb, an adsorber recommended for adsorption of cytokines, were tested by incubating them in spiked whole blood or plasma for 1 h. Subsequently, the adsorption characteristics of the adsorbers were tested in a dynamic system. For this purpose, a 6-h in vitro hemoperfusion treatment was compared with an equally long in vitro hemodialysis treatment. Results: Hippuric acid, homocysteine, indoxyl sulfate, and tryptophan were most effectively removed by hemodialysis. Bilirubin and cholic acid were removed best by hemoperfusion with Cytosorb. A treatment with Jafron HA and Biosky MG showed similar results for the adsorption of the tested retention solutes and were best for removing phenol. 3-Carboxy-4-methyl-5-propyl-2-furanpropionic acid could not be removed with any treatment method. Discussion/Conclusion: A combination of hemodialysis with hemoperfusion seems promising to improve the removal of some toxic metabolites in extracorporeal therapies. However, some very strongly protein-bound metabolites cannot be removed adequately with the adsorbers tested.

Microbe-derived uremic solutes enhance thrombosis potential in the host.

Alterations in gut microbial composition and function have been linked to numerous diseases. Identifying microbial pathways responsible for producing molecules that adversely impact the host is an important first step in the development of therapeutic interventions. Here, we first use large-scale clinical observations to link blood levels of defined microbial products to cardiovascular disease risks. Notably, the previously identified uremic toxins p-cresol sulfate and indoxyl sulfate were shown to predict 5-year mortality risks. After identifying the microbes and microbial enzymes involved in the generation of these uremic toxins, we used bioengineering technologies coupled with colonization of germ-free mice to show that the gut microbial genes that generate p-cresol and indole are sufficient to confer p-cresol sulfate and indoxyl sulfate formation, and a pro-thrombotic phenotype in vivo. The findings and tools developed serve as a critical step in both the study and targeting of these gut microbial pathways in vivo.

Incorporating Uremic Solute-mediated Inhibition of OAT1/3 Improves PBPK Prediction of Tenofovir Renal and Systemic Disposition in Patients with Severe Kidney Disease.

Dose modification of renally secreted drugs in patients with chronic kidney disease (CKD) has relied on serum creatinine concentration as a biomarker to estimate glomerular filtration (GFR) under the assumption that filtration and secretion decline in parallel. A discrepancy between actual renal clearance and predicted renal clearance based on GFR alone is observed in severe CKD patients with tenofovir, a compound secreted by renal OAT1/3. Uremic solutes that inhibit OAT1/3 may play a role in this divergence. To examine the impact of transporter inhibition by uremic solutes on tenofovir renal clearance, we determined the inhibitory potential of uremic solutes hippuric acid, indoxyl sulfate, and p-cresol sulfate. The inhibition parameters (IC50) were incorporated into a previously validated mechanistic kidney model; simulated renal clearance and plasma PK profile were compared to data from clinical studies. Without the incorporation of uremic solute inhibition, the PBPK model failed to capture the observed data with an absolute average fold error (AAFE) > 2. However, when the inhibition of renal uptake transporters and uptake transporters in the slow distribution tissues were included, the AAFE value was within the pre-defined twofold model acceptance criterion, demonstrating successful model extrapolation to CKD patients. A PBPK model that incorporates inhibition by uremic solutes has potential to better predict renal clearance and systemic disposition of secreted drugs in patients with CKD. Ongoing research is warranted to determine if the model can be expanded to include other OAT1/3 substrate drugs and to evaluate how these findings can be translated to clinical guidance for drug selection and dose optimization in patients with CKD.

Induction of Vesicular Trafficking and JNK-Mediated Apoptotic Signaling in Mononuclear Leukocytes Marks the Immuno-Proteostasis Response to Uremic Proteins

Introduction: Uremic retention solutes have been alleged to induce the apoptotic program of different cell types, including peripheral blood mononuclear leukocytes (PBL), which may contribute to uremic leukopenia and immune dysfunction. Methods: The molecular effects of these solutes were investigated in uremic PBL (u-PBL) and mononuclear cell lines (THP-1 and K562) exposed to the high molecular weight fraction of uremic plasma (u-HMW) prepared by in vitro ultrafiltration with 50 kDa cut-off microconcentrators. Results: u-PBL show reduced cell viability and increased apoptotic death compared to healthy control PBL (c-PBL). u-HMW induce apoptosis both in u-PBL and c-PBL, as well as in mononuclear cell lines, also stimulating cellular H2O2 formation and secretion, IRE1-α-mediated endoplasmic reticulum stress signaling, and JNK/cJun pathway activation. Also, u-HMW induce autophagy in THP-1 monocytes. u-PBL were characterized by the presence in their cellular proteome of the main proteins and carbonylation targets of u-HMW, namely albumin, transferrin, and fibrinogen, and by the increased expression of receptor for advanced glycation end-products, a scavenger receptor with promiscuous ligand binding properties involved in leukocyte activation and endocytosis. Conclusions: Large uremic solutes induce abnormal endocytosis and terminal alteration of cellular proteostasis mechanisms in PBL, including UPR/ER stress response and autophagy, ultimately activating the JNK-mediated apoptotic signaling of these cells. These findings describe the suicidal role of immune cells in facing systemic proteostasis alterations of kidney disease patients, a process that we define as the immuno-proteostasis response of uremia.

Application of the second-order fluid model to estimate factors affecting clearance of small and middle uremic molecules using combined ultrafiltration and diffusion processes

Uremic solute transport in a hollow fiber dialyzer is investigated theoretically with combined ultrafiltration and diffusion with the assumption of uniform ultrafiltration velocity and other standard assumptions. A numerical solution for concentration profiles in the blood compartment is obtained by using an implicit finite difference scheme implemented with MATLAB R2018a. The resulting system of linear equations is solved using another Matlab script implementing the Gauss–Seidel method. Uremic clearance of three solutes viz. urea, creatinine, and vitamin B-12 are considered. Different factors affecting Cl were observed and discussed in detail. Those factors include operating conditions (inlet blood flow rate, ultrafiltration velocity, etc.), dialyzer geometry (length, radius, number of fibers), and fluid parameters. The solute clearance can be maximized for a suitable set of values of these parameters.