Uremic Solutes Indoxyl Sulfate Research Articles

P1059REMOVAL OF LOW MOLECULAR WEIGHT AND PROTEIN-BOUND UREMIC RETENTION SOLUTES WITH ALLO-HEMODIALYSIS: RESULTS FROM EX VIVO STUDY

Abstract Background and Aims It is projected that in 2030, 14.5 million people will have end stage kidney disease and need kidney replacement therapy, yet only 5.4 million will receive it due to economic, social, and political factors. We have proposed allo-hemodialysis (alloHD) as a simple and low-cost HD alternative (https://www.kidneynews.org/kidney-news/features/buddy-dialysis-probed-hemodialysis-alternative). In alloHD, the patient’s blood is dialyzed against the blood of a healthy subject (‘buddy’), who receives the excess fluid and uremic solutes and excretes them via his/her healthy kidneys. Method We conducted ex vivo experiment with bovine whole blood, 4L in a patient and buddy bucket, respectively. In this setup, buddy blood flows through the dialyzer fiber lumen, while patient blood flows in the dialysate space (Nipro Cellentia 17H dialyzer). The patient blood was spiked with urea, creatinine, potassium chloride, indoxyl sulfate (IS), and p-cresyl sulfate (pCS); solute levels on the buddy side were not altered. The alloHD session lasted for 3 hours, the ultrafiltration (UF) volume was 750 mL. The blood flow rates were kept constant at 150 and 200 mL/min on the patient and buddy side, respectively. Heparin (5000 IU/L) was added to either bucket. The alloHD machine prototype comprises 2 pumps on the buddy side and one on patient side (Figure 1A). UF is controlled by the blood flow rate differential between the buddy-sided arterial and venous pumps, respectively. Results The levels of small unbound solutes (urea, creatinine, potassium) equilibrate quickly between patient and buddy buckets (Fig. 1B, top panel). Of note, the buddy-sided potassium equilibrium concentration is lower, because ongoing UF dilutes albumin in the buddy bucket and concentrates it in the patient bucket, resulting in a Gibbs-Donnan potential. The protein-bound uremic solutes IS and pCS equilibrate towards lower total concentrations on the buddy side due to the earlier mentioned UF-induced albumin dilution. Of note, the levels of free IS and pCS, respectively, converge on both sides of the dialyzer membrane (Fig 1B, bottom panel). Conclusion Our ex vivo data suggest that alloHD can be used to treat hyperkalemia, a major cause of death in acute and chronic kidney failure. Both unbound and protein-bound low molecular weight uremic solutes are also removed. With elimination of “classical” dialysate, we simplify the HD procedure and reduce machine size and costs significantly, making it an affordable HD alternative. Feasibility and efficacy studies in animal models are the next step.

Serum Levels and Removal by Haemodialysis and Haemodiafiltration of Tryptophan-Derived Uremic Toxins in ESKD Patients.

Tryptophan is an essential dietary amino acid that originates uremic toxins that contribute to end-stage kidney disease (ESKD) patient outcomes. We evaluated serum levels and removal during haemodialysis and haemodiafiltration of tryptophan and tryptophan-derived uremic toxins, indoxyl sulfate (IS) and indole acetic acid (IAA), in ESKD patients in different dialysis treatment settings. This prospective multicentre study in four European dialysis centres enrolled 78 patients with ESKD. Blood and spent dialysate samples obtained during dialysis were analysed with high-performance liquid chromatography to assess uremic solutes, their reduction ratio (RR) and total removed solute (TRS). Mean free serum tryptophan and IS concentrations increased, and concentration of IAA decreased over pre-dialysis levels (67%, 49%, −0.8%, respectively) during the first hour of dialysis. While mean serum total urea, IS and IAA concentrations decreased during dialysis (−72%, −39%, −43%, respectively), serum tryptophan levels increased, resulting in negative RR (−8%) towards the end of the dialysis session (p < 0.001), despite remarkable Trp losses in dialysate. RR and TRS values based on serum (total, free) and dialysate solute concentrations were lower for conventional low-flux dialysis (p < 0.001). High-efficiency haemodiafiltration resulted in 80% higher Trp losses than conventional low-flux dialysis, despite similar neutral Trp RR values. In conclusion, serum Trp concentrations and RR behave differently from uremic solutes IS, IAA and urea and Trp RR did not reflect dialysis Trp losses. Conventional low-flux dialysis may not adequately clear Trp-related uremic toxins while high efficiency haemodiafiltration increased Trp losses.

Indolic uremic solutes enhance procoagulant activity of red blood cells through phosphatidylserine exposure and microparticle release.

Increased accumulation of indolic uremic solutes in the blood of uremic patients contributes to the risk of thrombotic events. Red blood cells (RBCs), the most abundant blood cells in circulation, may be a privileged target of these solutes. However, the effect of uremic solutes indoxyl sulfate (IS) and indole-3-acetic acid (IAA) on procoagulant activity (PCA) of erythrocyte is unclear. Here, RBCs from healthy adults were treated with IS and IAA (mean and maximal concentrations reported in uremic patients). Phosphatidylserine (PS) exposure of RBCs and their microparticles (MPs) release were labeled with Alexa Fluor 488-lactadherin and detected by flow cytometer. Cytosolic Ca2+ ([Ca2+]) with Fluo 3/AM was analyzed by flow cytometer. PCA was assessed by clotting time and purified coagulation complex assays. We found that PS exposure, MPs generation, and consequent PCA of RBCs at mean concentrations of IS and IAA enhanced and peaked in maximal uremic concentrations. Moreover, 128 nM lactadherin, a PS inhibitor, inhibited over 90% PCA of RBCs and RMPs. Eryptosis or damage, by indolic uremic solutes was due to, at least partially, the increase of cytosolic [Ca2+]. Our results suggest that RBC eryptosis in uremic solutes IS and IAA plays an important role in thrombus formation through releasing RMPs and exposing PS. Lactadherin acts as an efficient anticoagulant in this process.

Indolic uremic solutes increase tissue factor production in endothelial cells by the aryl hydrocarbon receptor pathway

In chronic kidney disease (CKD), uremic solutes accumulate in blood and tissues. These compounds probably contribute to the marked increase in cardiovascular risk during the progression of CKD. The uremic solutes indoxyl sulfate and indole-3-acetic acid (IAA) are particularly deleterious for endothelial cells. Here we performed microarray and comparative PCR analyses to identify genes in endothelial cells targeted by these two uremic solutes. We found an increase in endothelial expression of tissue factor in response to indoxyl sulfate and IAA and upregulation of eight genes regulated by the transcription factor aryl hydrocarbon receptor (AHR). The suggestion by microarray analysis of an involvement of AHR in tissue factor production was confirmed by siRNA inhibition and the indirect AHR inhibitor geldanamycin. These observations were extended to peripheral blood mononuclear cells. Tissue factor expression and activity were also increased by AHR agonist dioxin. Finally, we measured circulating tissue factor concentration and activity in healthy control subjects and in patients with CKD (stages 3-5d), and found that each was elevated in patients with CKD. Circulating tissue factor levels were positively correlated with plasma indoxyl sulfate and IAA. Thus, indolic uremic solutes increase tissue factor production in endothelial and peripheral blood mononuclear cells by AHR activation, evoking a 'dioxin-like' effect. This newly described mechanism of uremic solute toxicity may help understand the high cardiovascular risk of CKD patients.