Serum Concentrations Of Indoxyl Sulfate Research Articles

#885 Combination of medium cut-off dialyzer membrane and diet modification to reduce serum concentrations of p-cresyl sulfate and indoxyl sulfate

Abstract Background and Aims Online hemodiafiltration (ol-HDF) may be considered currently as the best choice for chronic hemodialysis (HD) treatment, but medium cut-off (MCO) membranes may provide similar efficiency for larger and protein-bound toxin removal. P-cresyl sulfate (PCS) and indoxyl-sulfate (IS) concentration depends mainly on residual renal function and diet. Increased dietary fiber intake and short-chain fatty acid (SCFA) supplementation were associated with decreased production of PCS and IS. The aim of this study was to pragmatically combine MCO use with diet modification in an attempt to reduce serum concentration of PCS and IS and compare this strategy to online HDF use. Method We report preliminary results from a subset of a study sample with 42 chronic prevalent HD patients in a prospective randomized controlled interventional study. Two weeks after standard bicarbonate HD (BHD), patients were randomized to hemodialysis with MCO membrane (EXP group) or ol-HDF with a standard "high-flux" (HF) dialyzer membrane (CON group). After 4 weeks, both groups were supplemented by a change in the diet with increase of the daily intake of dietary fiber to 30g and addition of a SCFA propionate in the dose of 1g daily for 8 weeks. Two main outcomes were serum concentrations of PCS and IS. Measurements were taken at baseline (after 2 week wash-in period), after 4 weeks of the first intervention phase (MCO vs. ol-HDF; T1), after 8 weeks of the second intervention phase (fiber and SCFA supplementation; T2) and finally, after 4 weeks of wash-out period with BHD (T3). Results From 42 participants, 38 completed the study (63.2% men, 66.4±10.9 years, Charlson comorbidity index 5.7±2.3). In the CON group, serum concentrations of PCS (in μg/ml) were 36.8±19.6 (baseline), 36.9±20.8 (T1), 32.4±17.4 (T2) and 38.6±22.1 (T3). Serum concentrations of IS (in μg/ml) were 31.8±13.0 (baseline), 32.9±14.7 (T1), 29.5±9.1 (T2) and 35.5±16.8 (T3). In the EXP group, serum concentrations of PCS were 33.5± 16.6 (baseline), 35.9±17.8 (T1), 34.0±16.1 (T2) and 37.4±21.5 (T3). Serum concentrations for IS were 39.6±12.6 (baseline), 37.3±12.9 (T1), 42.3±13.5 (T2) and 41.0±12.8 (T3). Repeated measures ANOVA revealed no significant time*group interaction for PCS (F(1,33)=0.479, p=0.698, ŋ2=0.014) and IS (F(1,33)=2.392, p=0.073, ŋ2=0.068). There was a significant within-group increase in serum IS concentration between T2 and T3 in CON group (p=0.042). There was a trend towards a decrease in PCS concentration between T1 and T2 and increase between T2 and T3 in CON group. Conclusion The results of the study suggest both hemodialysis methods are equivalent in eliminating PCS and IS. Additional fiber and SCFA propionate supplementation did not decrease serum PCS and IS concentrations significantly, although there was a trend towards lower serums concentration of PCS after diet modification in CON group.

#2224 Higher serum level of indoxyl sulfate is not associated with increased abdominal aorta calcification in patients on maintenance hemodialysis

Abstract Background and Aims Vascular calcification is strongly associated with an increase in all-cause and cardiovascular mortality, especially in patients with and stage renal disease. Results of numerous experimental studies have shown direct toxic effect of microbial-derived uremic toxins on vascular endothelial and smooth muscle cells which may contribute vascular calcification. Meanwhile, the effects of these metabolites on vascular remodeling in patients receiving dialysis treatment remains largely unclear. Here we aimed to investigate the relationship between serum level of the uremic toxin indoxyl sulfate and severity of abdominal aorta calcification (AAC) in patients on maintenance hemodialysis. Method This single-center, cross-sectional study included eighty (40 men, average age – 59,7±11,8 years) clinically stable patients receiving hemodialysis treatment for 52(21,3-120) months. 13(16,3%) participants had history of smoking, 27(33,8%) were diabetic, 63(78,8%) had high blood pressure, 17(21,3%) – previous history of ischemic heart disease, including 5(6,3%) suffered a myocardial infarction. Severity of AAC was determined by lateral lumbar radiography with Kauppila scoring. The total AAC score ranged from 0 to 24 points, and a higher AAC score indicated a more pronounced degree of calcification. Routine laboratory data were collected before hemodialysis session after fasting for 12 hours. Serum concentration of indoxyl sulfate was assessed by ELISA using commercially available kit (Cloud-Clone Corp., USA). Statistical analysis was performed with IBM SPSS Statistics 26 program. Non-parametric Spearman's rank correlation method was used to evaluate the associations between the studied parameters. Value of p <0.05 was considered statistically significant in all tests. Results Median serum indoxyl sulfate was 2,1(1,4-3,0) μmol/l (the same parameter in relatively healthy individuals without impaired renal function is 0,1(0-0,3) μmol/l, р˂0,001). Serum levels of calcium (Ca) was 2,27(2,1-2,5) mmol/l, ionized calcium (Ca++) – 1,15(1,1-1,3) mmol/l, phosphate (P) – 1,6(1,3-2,0) mmol/l, intact parathyroid hormone (iPTH) – 303(162,2-492,8) pg/ml, total cholesterol (TH) – 4,5(3,5-5,4) mmol/l, low-density lipoprotein (LDL) – 2,6(2,0-3,3) mmol/l. AAC score in hemodialysis participants was 4,5(0,0-9,0) and significantly correlated with age (r=0,693; р˂0,001), Ca (r=0,208; р=0,04) and iPTH (r=0,418; р=0,021) but not with other indicators, including indoxyl sulfate concentration (r=0,164; р=0,147). Conclusion Our preliminary results do not allow to consider the elevated level of indoxyl sulfate as a factor contributing to increased AAC in patients on maintenance hemodialysis. Further prospective studies in larger populations should clarify the true role of indoxyl sulfate in vascular remodeling in individuals with chronic kidney disease.

Indoxyl sulfate induced frailty in patients with end-stage renal disease by disrupting the PGC-1α-FNDC5 axis.

Sarcopenia or frailty is common among patients with chronic kidney disease (CKD). The protein-bound uremic toxin indoxyl sulfate (IS) is associated with frailty. IS induces apoptosis and disruption of mitochondrial activity in skeletal muscle. However, the association of IS with anabolic myokines such as irisin in patients with CKD or end-stage renal disease (ESRD) is unclear. This study aims to elucidate whether IS induces frailty by dysregulating irisin in patients with CKD. The handgrip strength of 53 patients, including 28 patients with ESRD, was examined. Serum concentrations of IS and irisin were analyzed. CKD was established in BALB/c mice through 5/6 nephrectomy. Pathologic analysis of skeletal muscle was assessed through haematoxylin and eosin and Masson's trichrome staining. Expression of peroxisome proliferator-activated receptor-gamma coactivator PGC-1α and irisin were analyzed using real-time polymerase chain reaction and Western blotting. Handgrip strength was lower among patients with ESRD than among those without ESRD. In total, 64.3% and 24% of the patients in the ESRD and control groups had low handgrip strength, respectively (p < 0.05). Serum concentrations of IS were significantly higher in the ESRD group than in the control group (222.81 ± 90.67 μM and 23.19 ± 33.28 μM, respectively, p < 0.05). Concentrations of irisin were lower in the ESRD group than in the control group (64.62 ± 32.64 pg/mL vs. 99.77 ± 93.29 pg/mL, respectively, p < 0.05). ROC curves for low handgrip strength by irisin and IS were 0.298 (95% confidence interval (CI): 0.139-0.457, p < 0.05) and 0.733 (95% CI: 0.575-0.890, p < 0.05), respectively. The percentage of collagen was significantly higher in mice with 5/6 nephrectomy than in the control group. After resveratrol (RSV) treatment, the percentage of collagen significantly decreased. RSV modulates TGF-β signaling. In vitro analysis revealed that IS treatment suppressed expression of PGC-1α and FNDC5 in a dose-dependent manner, whereas RSV treatment attenuated IS-induced phenomena in C2C12 cells. IS was positively correlated with frailty in patients with ESRD through the modulation of the PGC-1α-FNDC5 axis. RSV may be a potential drug for reversing IS-induced suppression of the PGC-1α-FNDC5 axis in skeletal muscle.

Clinical significance of uremic toxin indoxyl sulfate and inflammation in the development of vascular calcification and cardiovascular complications in stage C3-C5D chronic kidney disease

To clarify the role of the uremic toxin indoxyl sulfate (IS) and inflammation in the development of vascular calcification and cardiovascular complications in chronic kidney disease (CKD). One hundred fifteen patients aged 25 to 68 years with CKD stage C3-C5D were examined. Serum concentrations of IS, interleukin 6 (IL-6), tumor necrosis factor (TNF-α), troponin I, parathyroid hormone were determined by enzyme immunoassay using kits from BluGene biotech (Shanghai, China), Cloud-Clone Corp. (USA), ELISA Kit (Biomedica, Austria). An increase in the serum concentration of IS, IL-6, TNF-α was revealed, which was significantly associated with a deterioration in renal function and changes in the morphological and functional parameters of the heart and aorta. High concentrations of IS, IL-6, TNF-α, which are closely associated with an increase in renal failure and cardiovascular complications, indicate their significant role in vascular calcification, which underlies the damage to the cardiovascular system in CKD.

The Effect of Synbiotic Supplementation on Uremic Toxins, Oxidative Stress, and Inflammation in Hemodialysis Patients-Results of an Uncontrolled Prospective Single-Arm Study.

Introduction: Numerous studies to date have shown that the development of dysbiotic gut microbiota is a characteristic finding in chronic kidney disease (CKD). A number of uremic toxins progressively accumulate in the course of CKD, some of them generated by the intestinal microbiome, such as indoxyl sulfate (IS) and p-cresyl sulfate (p-CS). They are found to be involved in the pathogenesis of certain complications of uremic syndrome, including low-grade chronic inflammation and oxidative stress. The aim of the present study is to research the serum concentration of IS and p-CS in end stage renal disease (ESRD) patients undergoing conventional hemodialysis, as well as to study the possibilities of influencing some markers of inflammation and oxidative stress after taking a synbiotic. Materials and Methods: Thirty patients with end-stage renal disease (ESRD) undergoing hemodialysis treatment who were taking a synbiotic in the form of Lactobacillus acidophilus La-14 2 × 1011 (CFU)/g and prebiotic fructooligosaccharides were included in the study. Serum levels of total IS, total p-CS, Interleukin-6 (IL-6), and Malondialdehyde (MDA) were measured at baseline and after 8 weeks. Results. The baseline values of the four investigated indicators in the patients were significantly higher-p-CS (29.26 ± 58.32 pg/mL), IS (212.89 ± 208.59 ng/mL), IL-6 (13.84 ± 2.02 pg/mL), and MDA (1430.33 ± 583.42 pg/mL), compared to the results obtained after 8 weeks of intake, as we found a significant decrease in the parameters compared to the baseline-p-CS (6.40 ± 0.79 pg/mL, p = 0.041), IS (47.08 ± 3.24 ng/mL, p < 0.001), IL-6 (9.14 ± 1.67 pg/mL, p < 0.001), and MDA (1003.47 ± 518.37 pg/mL, p < 0.001). Conclusions: The current study found that the restoration of the intestinal microbiota in patients with CKD significantly decreases the level of certain uremic toxins. It is likely that this favorably affects certain aspects of CKD, such as persistent low-grade inflammation and oxidative stress.

Indoxyl sulfate mediates low handgrip strength and is predictive of high hospitalization rates in patients with end-stage renal disease.

Sarcopenia has a higher occurrence rate in patients with chronic kidney disease (CKD) and end-stage renal disease (ESRD) than in the general population. Low handgrip strength-and not sarcopenia per se-is associated with clinical outcomes in patients with CKD, including cardiovascular mortality and hospitalization. The factors contributing to low handgrip strength are still unknown. Accordingly, this study aimed to determine whether uremic toxins influence low handgrip strength in patients with CKD. This cohort study lasted from August 2018 to January 2020. The participants were divided into three groups: the control group [estimated glomerular filtration rate (eGFR) ≥ 60 ml/min], an advanced CKD group (eGFR = 15-60 ml/min), and an ESRD group (under maintenance renal replacement therapy). All participants underwent handgrip strength measurement, dual-energy X-ray absorptiometry, and blood sampling for myokines (irisin, myostatin, and interleukin 6) and indoxyl sulfate. Sarcopenia was defined according to the Asian Working Group for Sarcopenia consensus as low appendicular skeletal muscle index (appendicular skeletal muscle/height2 of < 7.0 kg/m2 in men and < 5.4 kg/m2 in women) and low handgrip strength (< 28 kg in men and < 18 kg in women). Among the study participants (control: n = 16; CKD: n = 17; and ESRD: n = 42), the ESRD group had the highest prevalence of low handgrip strength (41.6 vs. 25% and 5.85% in the control and CKD groups, respectively; p < 0.05). The sarcopenia rate was similar among the groups (12.5, 17.6, and 19.5% for the control, CKD, and ESRD groups, respectively; p = 0.864). Low handgrip strength was associated with high hospitalization rates within the total study population during the 600-day follow-up period (p = 0.02). The predictions for cardiovascular mortality and hospitalization were similar among patients with and without sarcopenia (p = 0.190 and p = 0.094). The serum concentrations of indoxyl sulfate were higher in the ESRD group (227.29 ± 92.65 μM vs. 41.97 ± 43.96 μM and 6.54 ± 3.45 μM for the CKD and control groups, respectively; p < 0.05). Myokine concentrations were similar among groups. Indoxyl sulfate was associated with low handgrip strength in univariate and multivariate logistic regression models [univariate odds ratio (OR): 3.485, 95% confidence interval (CI): 1.372-8.852, p = 0.001; multivariate OR: 8.525, 95% CI: 1.807-40.207, p = 0.007]. Handgrip strength was lower in the patients with ESRD, and low handgrip strength was predictive of hospitalization in the total study population. Indoxyl sulfate contributed to low handgrip strength and counteracted the benefits of myokines in patients with CKD.

Older Patients on Hemodiafiltration: Behavior of Uremic Toxins, Inflammation, Endothelium, and Bone Mineral Disorder

Chronic kidney disease (CKD) affects 10% of the world’s population. Uremic toxins, such as indoxyl sulfate (IS), p-Cresylsulfate (PCS) and indole acetic acid (IAA), are not sufficiently removed by conventional hemodialysis (HD) and have been associated with inflammation, poor quality of life, bone mineral disease (BMD) and endothelial injury. Online hemodiafiltration (OL-HDF) may promote greater clearance of uremic toxins than HD. However, there are few studies evaluating the effect of OL-HDF on serum levels of IS, PCS, IAA, and biomarkers associated with inflammatory, endothelial, and bone and mineral disorder in the elderly population. We evaluated the effect of 6 months of OL-HDF on the serum concentration of uremic toxins, biomarkers of inflammation, endothelial and bone mineral disorder in older patients on OL-HDF. IS, PCS, and IAA were measured by high-performance liquid chromatography. We included 31 patients (77.4 ± 7.1 years, 64.5% male, 35.5% diabetic, on maintenance dialysis for 45 ± 20 days). From baseline to 6 months there was a decrease in serum concentration of IS but not PCS and IAA. We found no change in serum concentration of inflammatory, endothelial, or mineral and bone biomarkers. In summary, OL-HDF was capable to reduce IS in older patients. Whether this reduction may have an impact on clinical outcomes deserves further evaluation.

Effect of Unripe Banana Flour on Gut-Derived Uremic Toxins in Individuals Undergoing Peritoneal Dialysis: A Randomized, Double-Blind, Placebo-Controlled, Crossover Trial.

In chronic kidney disease (CKD), the accumulation of gut-derived metabolites, such as indoxyl sulfate (IS), p-cresyl sulfate (pCS), and indole 3-acetic acid (IAA), has been associated with the burden of the disease. In this context, prebiotics emerge as a strategy to mitigate the accumulation of such compounds, by modulating the gut microbiota and production of their metabolites. The aim of this study was to evaluate the effect of unripe banana flour (UBF—48% resistant starch, a prebiotic) on serum concentrations of IS, pCS, and IAA in individuals undergoing peritoneal dialysis (PD). A randomized, double-blind, placebo-controlled, crossover trial was conducted. Forty-three individuals on PD were randomized to sequential treatment with UBF (21 g/day) and placebo (waxy corn starch—12 g/day) for 4 weeks, or vice versa (4-week washout). The primary outcomes were total and free serum levels of IS, pCS, and IAA. Secondary outcomes were 24 h urine excretion and dialysis removal of IS, pCS, and IAA, serum inflammatory markers [high-sensitivity C-reactive protein (hsCRP), interleukin-6 (IL-6), interleukin-10 (IL-10), and tumor necrosis factor-α (TNF-α)], serum lipopolysaccharide LPS, and dietary intake. Of the 43 individuals randomized, 26 completed the follow-up (age = 55 ± 12 years; 53.8% men). UBF did not promote changes in serum levels of IS (p = 0.70), pCS (p = 0.70), and IAA (p = 0.74). Total serum IS reduction was observed in a subgroup of participants (n = 11; placebo: median 79.5 μmol/L (31–142) versus UBF: 62.5 μmol/L (31–133), p = 0.009) who had a daily UBF intake closer to that proposed in the study. No changes were observed in other secondary outcomes. UBF did not promote changes in serum levels of IS or pCS and IAA; a decrease in IS was only found in the subgroup of participants who were able to take 21g/day of the UBF.

Pro-Inflammatory Effects of Indoxyl Sulfate in Mice: Impairment of Intestinal Homeostasis and Immune Response.

The intestines are recognized as the main source of chronic inflammation in chronic kidney disease (CKD) and, among other cells, macrophages are involved in modulating this process as well as in the impaired immune response which also occurs in CKD patients. In this study, we evaluated the effect of Indoxyl Sulfate (IS), a protein bound uremic toxin poorly eliminated by hemodialysis, on inflammatory, oxidative stress and pro-apoptotic parameters, at the intestinal level in mice, on intestinal epithelial cells (IEC-6) and on primary murine peritoneal macrophages. C57BL/6J mice were treated with IS (800 mg/kg i.p.) for 3 or 6 h and histopathological analysis showed that IS induced intestinal inflammation and increased cyclooxygenase-2 (COX-2), nitrotyrosine and Bax expression in intestinal tissue. In IEC-6 cells, IS (125–1000 µM) increased tumor necrosis factor-α levels, COX-2 and inducible nitric oxide synthase expression and nitrotyrosine formation. Moreover, IS increased pro-oxidant, pro-inflammatory and pro-apoptotic parameters in peritoneal macrophages from IS-treated mice. Also, the serum concentration of IS and pro-inflammatory levels of cytokines resulted increased in IS-treated mice. Our results indicate that IS significantly contributes to affect intestinal homeostasis, immune response, and to induce a systemic pro-inflammatory state thus highlighting its potential role as therapeutic target in CKD patients.

P1509SEVERITY OF PRURITUS IN HEMODIALYSIS PATIENTS: RELATIONSHIP TO UREMIC TOXINS AND DIALYSIS MODALITY

Abstract Background and Aims Pruritus is a complication that can impair the quality of life of patients with chronic kidney disease (CKD). A previous report has suggested that the severity of pruritus is correlated with high serum concentrations of β2-microglobulin, calcium, and phosphorus, however recent developments in hemodialysis (HD), especially use of high-flux membrane and online hemodiafiltration (OLHDF), may improve the severity of pruritus. Protein-bound uremic toxins (PBUTs), such as indoxyl sulfate and p-cresyl sulfate, are known to be factors associated with atherosclerosis and impaired bone metabolism in CKD and may also be associated with pruritus, especially in those undergoing dialysis treatments. The aim of our study was to assess the severity of pruritus using a new tool, the 5D-itch scale (5D), as well as its relation to circulating PBUTs and dialysis modality in patients with end-stage kidney disease. Method This was a prospective cohort multicenter study, wherein we analyzed the baseline data. The 5D score (5–25 points) was measured in maintenance dialysis patients. The serum concentrations of indoxyl sulfate, p-cresyl sulfate, phenyl sulfate and hippuric acid were measured with mass spectrometry before a dialysis session. The correlations of the 5D scores with serum PBUTs, dialysis modality (HD or OLHDF) and the other background factors were assessed with Spearman correlation coefficients and the generalized linear model. Results This study included 135 dialysis patients (98 HD and 37 OLHDF). The mean 5D score was 6.84 (±SD 3.14). The mean serum concentrations of indoxyl sulfate, p-cresyl sulfate, indole acetic acid, phenyl sulfate, and hippuric acid were 2.99 ± 1.40, 2.05 ± 1.54, 0.11 ± 0.08, 0.94 ± 0.71, and 4.52 ± 3.17 mg/dL, respectively. There was a significant correlation between the 5D score and dialysis modality (correlation coefficient = 0.24, p &amp;lt; 0.01). Dialysis method was associated with the 5D score by the generalized linear model, and OLHDF was associated with a lower 5D score. There were no significant associations between the 5D score and serum concentrations of the various PBUTs (e.g., for indoxyl sulfate, the correlation coefficient was 0.11, p = 0.20) Conclusion Pruritus, as assessed by the 5D score, was associated with dialysis modality, but not with the serum concentration of PBUTs. Further studies will be needed to understand the mechanism of the effect of OLHDF on pruritus as well as the role of uremic toxins in the itchy skin in CKD patients.

Preliminary evaluation of fecal fatty acid concentrations in cats with chronic kidney disease and correlation with indoxyl sulfate and p-cresol sulfate.

BackgroundStraight‐ and branched‐chain (BCFA) short‐chain fatty acids (SCFAs) are produced by colonic microbiota and have both beneficial and deleterious effects in humans with chronic kidney disease (CKD). Fecal SCFAs in cats with CKD have not been described.ObjectiveTo characterize fecal SCFA concentrations in cats with CKD as compared to healthy geriatric cats and correlate SCFA to serum indoxyl sulfate (IS) and p‐cresol sulfate (pCS) concentrations.AnimalsTwenty‐eight cats with CKD (International Renal Interest Society [IRIS] stages 2, 3, and 4) and 11 older (≥ 8 years) healthy geriatric cats.MethodsProspective, cross‐sectional study. Voided feces were analyzed using stable isotope dilution gas chromatography‐mass spectrometry to determine fecal concentrations of SCFAs. Serum concentrations of IS and pCS were measured using liquid chromatography tandem mass spectrometry.ResultsFecal isovaleric acid concentrations were significantly higher in CKD cats(P = .02) Cats with IRIS CKD stage 3 and 4 had significantly higher fecal isovaleric acid concentrations compared to healthy geriatric cats (P = .03), but not compared to IRIS CKD stage 2 cats. Total fecal concentrations of BCFAs were found to correlate weakly with serum creatinine concentration (rho, 0.33; P = .05), blood urea nitrogen concentration (rho, 0.40; P = .01), and pCS concentration (rho, 0.35; P = .04).Conclusions and Clinical ImportanceFecal isovaleric acid concentrations were higher in CKD cats, particularly in late stage disease, compared to healthy geriatric cats. Fecal BCFA concentrations correlated with pCS and were higher in cats with muscle wasting, providing evidence for malassimilation of protein in CKD cats.

Serum Concentration of Indoxyl Sulfate in Peritoneal Dialysis Patients and Low-Flux Hemodialysis Patients

Background/Aims: To compare the serum concentration of Indoxyl sulfate (IS) in patients on continuous ambulatory peritoneal dialysis (CAPD) and low-flux hemodialysis (HD), and analyze the risk factors associated with IS. Methods: We performed a single-center, cross-sectional observational study including 169 patients on CAPD and 115 patients on low-flux HD. Patients were divided into the anuric HD group, anuric peritoneal dialysis (PD) group, and non-anuric PD group on the basis of dialysis modality and residual urinary output. Serum concentration of IS was determined by high-performance liquid chromatography electrospray tandem mass spectrometry. Results: After matching the urinary volume and dialysis vintage, 58 anuric patients on PD and 58 anuric patients on HD were enrolled. The serum level of IS was significantly lower in patients on PD than that in those on HD (28.05 ± 13.98 vs. 39.64 ± 18.25 μg/mL; p < 0.001). This result persisted even after adjustment for confounding risk factors including nutritional status (β = 0.338, p < 0.001). In addition, the serum level of IS was significantly lower in non-anuric PD patients than that anuric PD patients (18.70 ± 11.21 vs. 28.05 ± 13.98 μg/mL; p < 0.001). After the adjustment for risk factors such as dialysis vintage, IS serum concentration in patients on PD was still significantly correlated with residual renal function (RRF; β = –0.355, p < 0.001). Conclusions: Dialysis modality is the independent risk factor of IS serum concentration and it is substantially lower in patients on CAPD than that in those on low-flux HD. Additionally, RRF was independently associated with IS serum concentration in CAPD patients, and the better the RRF is, the lower IS serum concentration.

Prognostic Value Serum Concentration of Indoxyl Sulfate During Acute Kidney Injury in Septic Shock Patients.

Prognostic Value Serum Concentration of Indoxyl Sulfate During Acute Kidney Injury in Septic Shock Patients.

Neutrophil/lymphocyte ratio elevation in renal dysfunction is caused by distortion of leukocyte hematopoiesis in bone marrow

Objective: We investigate the mechanism of neutrophil/lymphocyte ratio (NLR) elevation, a useful prognostic marker in patients with cardiovascular diseases (CVDs).Methods: In this clinical study, we retrospectively searched for factors associated with NLR elevation in cardiovascular outpatients. In animal experiments using mice with adenine-induced nephropathy, we further examined the hematopoietic process in bone marrow and explored the mechanism of NLR elevation.Result: In patients with CVDs or their risk factors, multiple regression analysis revealed that decrease in estimated glemerular filtration rate and increase in white blood cell count were significantly associated with increase in NLR. In mice with adenine-induced nephropathy, NLR and serum indoxyl sulfate (IS) levels were increased. Fluorescence-activated cell sorting revealed the increase in the number of myeloid progenitors and decrease in the number of common lymphoid progenitors, suggesting biased granulocyte side in the hematopoietic process in bone marrow. Treatment with oral charcoal adsorbent AST-120 decreased serum concentration of IS and normalized NLR and bone marrow abnormalities in mice with adenine-induced nephropathy.Conclusion: Renal function was a strong determinant of NLR in cardiovascular outpatients. NLR elevation due to renal impairment is caused by distortion of the hematopoietic process in bone marrow. IS plays a significant role in these processes.

The fecal microbiome and serum concentrations of indoxyl sulfate and p-cresol sulfate in cats with chronic kidney disease.

BackgroundIntestinal dysbiosis has been documented in humans with chronic kidney disease (CKD) and is thought to contribute to production of the uremic toxins indoxyl sulfate (IS) and p‐cresol sulfate (pCS). Characteristics of the fecal microbiome in cats with CKD and correlation to serum concentrations of uremic toxins are unknown.ObjectivesTo characterize the fecal microbiome and measure serum IS and pCS concentrations of cats with CKD in comparison to healthy older cats.AnimalsThirty client‐owned cats with CKD (International Renal Interest Society stages 2‐4) and 11 older (≥8 years) healthy control cats.MethodsProspective, cross‐sectional study. Fecal samples were analyzed by sequencing of 16S rRNA genes and Escherichia coli quantitative PCR (qPCR). Serum concentrations of IS and pCS measured using liquid chromatography tandem mass spectrometry.ResultsCats with CKD had significantly decreased fecal bacterial diversity and richness. Escherichia coli qPCR showed no significant difference in bacteria count between control and CKD cats. Cats with stage 2 (P = .01) and stages 3 and 4 (P = .0006) CKD had significantly higher serum IS concentrations compared to control cats. No significant difference found between stage 2 and stages 3 and 4 CKD. The pCS concentrations were not significantly different between CKD cats and control cats.Conclusions and Clinical ImportanceDecreased fecal microbiome diversity and richness is associated with CKD in cats. Indoxyl sulfate concentration is significantly increased with CKD, and cats with stage 2 CKD may suffer from a similar uremic toxin burden as do cats with later stage disease.

Albumin is the main plasma binding protein for indoxyl sulfate and p‐cresyl sulfate

Indoxyl sulfate and p-cresyl sulfate are two uremic retention solutes implicated in the uremic syndrome. Removal during dialysis is limited, mainly due to protein binding. Binding characteristics to healthy albumin have recently been characterized. Whether uremia alters the binding characteristics of albumin is currently unknown. Moreover, protein binding values previously determined with ultrafiltration are in sharp contrast to recently reported values based on microcalorimetry. In the present study, indoxyl sulfate and p-cresyl sulfate binding were therefore quantified using both equilibrium dialysis and ultrafiltration. Deming regression demonstrated good agreement between equilibrium dialysis and ultrafiltration. Free serum concentrations of indoxyl sulfate (+26.6%) and p-cresyl sulfate (+19.7%) were slightly higher at body temperature compared with at room temperature. To investigate binding kinetics, the plasma of healthy individuals or hemodialysis patients was titrated with albumin solutions. Theoretical models of protein binding were fitted to observed titration curves. Binding coefficients of both toxins were highest in purified albumin, and were reduced from healthy to uremic plasma. In conclusion, the ultrafiltration-HPLC technique reliably measures free serum concentrations of indoxyl sulfate and p-cresyl sulfate. Albumin is the main binding protein, both in health and in advanced stages of chronic kidney disease. Modeling suggests that albumin contains two binding sites for both toxins, a single high affinity binding site and a second low affinity binding site. The high affinity binding site accounts for at least 90% of overall binding. Competition for this binding site could be used to augment free solute concentrations during dialysis, thus improving epuration.

Indoxyl Sulfate Serum Level in Chronic Renal Failure Patients detected using Fluorescence-HPLC

Indoxyl sulfate (IS) is a protein-bound typical uremic toxin that accumulates in patients with impaired kidney function. The laboratory methods used to quantify serum concentrations of IS require improvement. We report an optimal, cost-effective alternative to the methods currently used.The methods is as follows: serum samples were extracted and deproteinized using acetonitrile and then further purified using dichloromethane. The samples were then analyzed using high-performance liquid chromatography (HPLC) with a fluorescence detector at a flow rate of 1.0 mL/min. The isocratic mobile phase consisted of acetonitrile−0.2% (V/V) trifluoroacetic acid in phosphate buffered saline, pH 2.5 (8:92, V/V). Detector settings were λex 280 nm/λem 390 nm. Under the indicated conditions, IS was well separated. The retention time was approximately 6.59±0.11 min. The results of the precision and reproducibility tests were In conclusion, HPLC with a fluorescence detector is a simple, sensitive, and reliable method for quantifying IS concentration in the serum of patients with chronic renal failure.

Protein-bound uremic toxins in hemodialysis patients measured by liquid chromatography/tandem mass spectrometry and their effects on endothelial ROS production

Cardiovascular disease (CVD) is prevalent in patients with chronic kidney disease (CKD). In hemodialysis (HD) patients, some protein-bound uremic toxins are considered to be associated with CVD. However, it is not yet known which uremic toxins are important in terms of endothelial toxicity. Serum samples were obtained from 45 HD patients before and after HD. Total and free serum concentrations of indoxyl sulfate, indoxyl glucuronide, indoleacetic acid, p-cresyl sulfate, p-cresyl glucuronide, phenyl sulfate, phenyl glucuronide, phenylacetic acid, phenylacetyl glutamine, hippuric acid, 4-ethylphenyl sulfate, and 3-carboxy-4-methyl-5-propyl-2-furanpropionic acid (CMPF) were simultaneously measured by liquid chromatography/electrospray ionization-mass spectrometry/mass spectrometry (LC/ESI-MS/MS). The effects of these solutes at their pre-HD mean and maximum serum concentrations on reactive oxygen species (ROS) production in human umbilical vein endothelial cells (HUVEC) were measured with a ROS probe. Serum levels of 11 of the solutes (all except 4-ethylphenyl sulfate) were significantly increased in HD patients compared to healthy subjects. All 12 solutes showed changes in their protein-binding ratios. In particular, indoxyl sulfate, p-cresyl sulfate, CMPF, and 4-ethylphenyl sulfate showed high protein-binding ratios (>95 %) and low reduction rates by HD (<35 %). Indoxyl sulfate at its mean and maximum pre-HD serum concentrations-even with 4 % albumin-stimulated ROS production in HUVEC most intensely, followed by CMPF. In conclusion, the serum levels of 11 protein-bound uremic toxins were increased in HD patients. Indoxyl sulfate, p-cresyl sulfate, and CMPF could not be removed efficiently by HD due to their high protein-binding ratios. Indoxyl sulfate most intensely induced endothelial ROS production, followed by CMPF.

The uremic solute indoxyl sulfate acts as an antioxidant against superoxide anion radicals under normal-physiological conditions

The effect of the uremic solute indoxyl sulfate (IS) on scavenging superoxide anion radicals (O2-) generated from both the xanthine/xanthine oxidase (X/XO) system and activated neutrophils was investigated by electron paramagnetic resonance spectroscopy, combined with 2-ethoxycarbonyl-2-methyl-3,4-dihydro-2H-pyrrole-1-oxide (EMPO). The findings show that the presence of normal-physiological serum concentrations of IS (0.1–10μM) resulted in decreased formation of EMPO-superoxide adduct without affecting XO activity. Furthermore, IS showed scavenging activity against cell-derived O2- generated from activated neutrophils. In addition, IS also eliminated hydroxyl radicals. These findings suggest that IS acts as a novel endogenous antioxidant under normal-physiological conditions.

Oral charcoal adsorbent (AST-120) prevents progression of cardiac damage in chronic kidney disease through suppression of oxidative stress

Chronic kidney disease (CKD) is an important risk factor for cardiovascular disease (CVD). Increased oxidative stress plays a role in the pathogenesis of CVD in CKD patients. The oral charcoal adsorbent AST-120 attenuates the progression of CKD possibly by removing uraemic toxins such as indoxyl sulfate (IS), and reduces oxidative stress. We investigated the relationship between oxidative stress and cardiac damage in CKD and its prevention by AST-120. Male Lewis rats were administered adriamycin at 8 weeks of age, and the right kidney was removed at 12 weeks of age. From 14 weeks of age, the rats were treated daily with AST-120 (n = 8) or were untreated (control group, n = 8). At 34 weeks of age, the rats were killed and urinary and blood biochemical tests as well as cardiac histological analyses were performed. At 14 weeks of age, there were no significant differences in blood pressure, renal function (creatinine clearance: 1.54 +/- 0.28 mL/min versus 1.60 +/- 0.22 mL/min), oxidative stress markers or other biochemical data between the control and AST-120 groups. At 34 weeks, despite similar blood pressure and renal function (creatinine clearance: 0.78 +/- 0.46 mL/min versus 0.75 +/- 0.54 mL/min), serum concentrations of IS and urinary excretion of 8-hydroxydeoxyguanosine (8-OHdG), acrolein and IS were significantly lower in the AST-120 group than in the control group. Heart volume, left ventricular volume and cardiac fibrosis were significantly smaller in the experimental AST-120 group than in the control group. Immunohistological analysis revealed that the numbers of 8-OHdG- and acrolein-positive cardiomyocytes and the degrees of myocardial and perivascular fibrosis were ameliorated by AST-120 administration. The myocardial fibrosis score was significantly associated with the 8-OHdG- (r = 0.848, P < 0.001) and acrolein-positive (r = 0.812, P < 0.001) cell scores. The perivascular fibrosis score was also significantly associated with the 8-OHdG- (r = 0.906, P < 0.0001) and acrolein-positive (r = 0.789, P < 0.001) cell scores. Oxidative stress is suggested to play a key role in the development of cardiac hypertrophy and fibrosis in CKD. AST-120 may suppress oxidative stress and reduce cardiac damage in CKD.