P-cresyl Sulfate Research Articles

P-Cresyl sulfate predicts clinical outcomes in sustained peritoneal dialysis: a 5-year follow-up cohort study and meta-analysis

Background The impact of p-cresyl sulfate (PCS) and indoxyl sulfate (IS) on the prognosis of patients with uremia remains controversial. We performed a prospective study on peritoneal dialysis (PD) to investigate the relationship between PCS or IS levels with clinical outcomes. Methods This prospective cohort study investigated the association of serum PCS and IS with clinical outcomes in patients undertaking PD. We performed a correlations analysis to explore the influencing factors of PCS an IS. Meta-analysis was conducted to objectively evaluate the prognostic effects of PCS and IS on different stages of CKD patients. Results A total of 127 patients were enrolled consecutively and followed with an average period of 51.3 months. Multivariate Cox regression showed that serum total PCS not only contributed to the occurrence of PD failure event (HR: 1.05, 95% CI = 1.02 to 1.07, p < 0.001), but also increased the risk of cardiovascular event (HR: 1.08, 95% CI = 1.04 to 1.13, p < 0.001) and PD-associated peritonitis (HR: 1.04, 95% CI = 1.02 to 1.08, p = 0.001). Dividing the total PCS level by 18.99 mg/L, which was calculated from the best cutoff value of the ROC curve, patients with total PCS higher than 18.99 mg/L had worse prognosis. Meta-analysis confirmed its value in cardiovascular event in PD. Conclusion The serum total PCS concentration was a detrimental factor for higher PD failure event, cardiovascular event, and PD-associated peritonitis. It could be used as an innovative marker in predicting poor clinical outcome in PD.

Proton pump inhibitor-induced risk of chronic kidney disease is associated with increase of indoxyl sulfate synthesis via inhibition of CYP2E1 protein degradation

Proton pump inhibitors (PPIs) are widely used to treat acid-related disorders in the gastrointestinal tract; however, PPI use increases the risk of chronic kidney disease (CKD) through unclear mechanisms. Considering that PPIs disturb the gut microbiome balance, which is involved in the precursor of gut-derived uremic toxin accumulation, and that gut-derived uremic toxins aggravate CKD progression, the aim of this study is to elucidate whether PPIs affect gut-derived uremic toxin metabolism, including indoxyl sulfate (IS), p-cresyl sulfate, and trimethylamine-N-oxide, as a mechanism for causing CKD. The present study showed that 3 week-treatment of PPIs (omeprazole, lansoprazole, and pantoprazole at 30 mg/kg) in mice only increased IS plasma levels among the above three gut-derived uremic toxins. Additionally, lansoprazole increased IS plasma concentrations along with increased exposure dose (7.5–30 mg/kg) and duration (1–3 weeks). However, nephrotoxicity with mild changes in glomerular structure and signs of fibrosis were observed only in groups exposed to a 3-week treatment of PPIs (30 mg/kg). As the concentrations of indole (the precursor of IS from gut metabolism) in the colon were only increased in the pantoprazole-treated group, the mechanism of increased IS exposure remains unclear. Further studies revealed that PPIs (omeprazole and lansoprazole; but not pantoprazole) increased IS production from indole in primary mouse hepatocytes in a concentration-dependent manner. Additionally, the increased protein levels of hepatic CYP2E1 (the key enzyme mediating IS formation) due to suppressed degradation resulted in an increase in IS levels. Although omeprazole and lansoprazole significantly inhibited IS uptake in hOAT1/3 in vitro, 3 weeks of PPI treatment did not reduce IS renal excretion in mice. In conclusion, PPIs induced IS synthesis via increased hepatic CYP2E1 protein level, subsequently leading to increased IS exposure. These findings present a plausible biological mechanism to explain the association of PPI use with the increased risk of CKD.

The New Nordic Renal Diet Induces a Pronounced Reduction of Urine Acid Excretion and Uremic Toxins in Chronic Kidney Disease Patients (Stage 3 and 4)

Metabolic acidosis and the uremic toxins, indoxyl sulfate (IS) and p-cresyl sulfate (PCS), are associated with increased risks of kidney disease progression, muscle catabolism, cardiovascular disease, and mortality in patients with chronic kidney disease (CKD). The New Nordic Renal Diet (NNRD) is a plant-focused meal pattern, with reduced phosphorus and protein content compared to an average Danish diet. Due to a higher amount of plant-based products, we hypothesized that NNRD would reduce renal excretion of acids and uremic toxins. Thus, we evaluated the effects of NNRD on metabolic acidosis and uremic toxins in patients with moderate CKD, stages 3-4. This post hoc analysis is based on a randomized controlled crossover trial comparing 1 week of the NNRD to a control 1-week period of an average Danish diet, in patients with CKD stages 3 and 4. Urine pH and urine excretion of bicarbonate, ammonium, titratable acids, IS, and PCS alongside plasma total CO2 (tCO2) were measured at days 1, 4, and 7 in 18 patients. After 7days on NNRD 24-hour urine net acid excretion was decreased by 80% (P<.001), 24-hour urine excretion of ammonium and bicarbonate decreased by 34% (P<.001), and increased by 678% (P<.001), respectively, compared with the control period. Plasma tCO2 was increased by 8% (P=.005). Moreover, 24-hour urine excretion of PCS and IS were reduced by 31% (P=.018) and 29% (P<.001), respectively. One week of NNRD in patients suffering from moderate CKD effectively improved metabolic acidosis with a marked reduction in urine net acid excretion that included a large increase in urine bicarbonate excretion. In addition, NNRD reduced urinary excretion of the uremic toxins PCS and IS. These results encourage further investigations of the long-term effects of NNRD on renal protection in patients with CKD.

Cerebrovascular Disease, Cardiovascular Disease, and Chronic Kidney Disease: Interplays and Influences.

We reviewed reasons for the high cardiovascular risk (CVD) of patients with chronic kidney disease (CKD), and explored alternatives to treatment of traditional risk factors to reduce CVD in CKD. Besides traditional risk factors, patients with CKD are exposed to uremic toxins of two kinds: systemically derived toxins include asymmetric dimethylarginine (ADMA), total homocysteine (tHcy), thiocyanate, tumor necrosis factor alpha, and interleukin 6. Gut-derived uremic toxins (GDUT), products of the intestinal microbiome, include hippuric acid, indoxyl sulfate, p-cresyl sulfate, p-cresyl glucuronide, phenylacetylglutamine, and trimethylamine N-oxide (TMAO). Cyanocobalamin is toxic in patients with CKD. Approaches to reducing plasma levels of these uremic toxins would include diet to reduce GDUT, kidney transplantation, more intensive dialysis, and vitamin therapy to lower tHcy with methylcobalamin rather than cyanocobalamin. The high CVD risk in CKD requires consideration of therapies beyond treatment of traditional risk factors.

Microbiome in Chronic Kidney Disease.

The gut microbiome is a complex collection of microorganisms with discrete characteristics and activities. Its important role is not restricted to food digestion and metabolism, but extends to the evolution, activation and function of the immune system. Several factors, such as mode of birth, diet, medication, ageing and chronic inflammation, can modify the intestinal microbiota. Chronic kidney disease (CKD) seems to have a direct and unique effect, as increased urea levels result in alteration of the gut microbiome, leading to overproduction of its metabolites. Therefore, potentially noxious microbial uremic toxins, which have predominantly renal clearance, including p-cresyl sulfate, indoxyl sulfate and N-oxide of trimethylamine [Trimethylamine-N-Oxide (TMAO)], accumulate in human’s body, and are responsible not only for the clinical implications of CKD, but also for the progression of renal failure itself. Certain changes in gut microbiome are observed in patients with end stage renal disease (ESRD), either when undergoing hemodialysis or after kidney transplantation. The purpose of this review is to summarize the changes of gut microbiome and the protein bound uremic toxins which are observed in CKD and in different kidney replacement strategies. In addition, we attempt to review the connection between microbiome, clinical implications and immune response in CKD.

Altered gut microbiotaand gut-derived p-cresyl sulfate serum levels in peritoneal dialysis patients.

Peritoneal dialysis (PD) is a renal replacement therapy for end-stage renal disease. Gut microbiota-derived uremic solutes, indoxyl sulfate (IS), p-cresyl sulfate (PCS), and trimethylamine-N-oxide (TMAO) accumulate in PD patients. The objective was to explore the gut microbiota and their influence on uremic toxins in PD patients and healthy controls (HC). Fecal samples were collected from PD patients (n = 105) and HC (n = 102). 16S rRNA gene regions were sequenced for gut microbiota analysis. IS, PCS, and TMAO levels were measured using HPLC-MS. PD patients exhibited lower alpha diversity and altered gut microbiota composition compared to HC. At the genus level, PD patients showed increased abundance of opportunistic pathogenic bacteria, and decreased abundance of beneficial bacteria. Three Operational Taxonomic Units discriminated PD patients from HC. Phenylalanine metabolism increased in PD, whereas tryptophan metabolism was unaltered. Low serum PCS did not necessarily mean healthier due to the loss of alpha diversity, increased Proteobacteria and opportunistic pathogenic bacteria. High serum PCS was mainly caused by elevated p-cresol-producing bacteria, enriched amino acid related enzymes, and enhanced sulfur metabolism, rather than declined residual renal function. In patients with different urine volumes, the gut microbiota alpha diversity and composition were unaltered, but serum IS and TMAO were significantly elevated in anuric patients. In conclusion, the gut microbiota abundance, composition, and function were altered in PD patients, which increased the PCS levels. We provided a better understanding of the microbiota-metabolite-kidney axis in PD patients. Targeting certain bacteria could decrease the PCS levels, whereas preserving the residual renal function could reduce the IS and TMAO levels.

The Autism Spectrum Disorder-Associated Bacterial Metabolite p-Cresol Derails the Neuroimmune Response of Microglial Cells Partially via Reduction of ADAM17 and ADAM10.

The bacterial metabolite 4-methylphenol (para-cresol or p-cresol) and its derivative p-cresyl sulfate (pCS) are elevated in the urine and feces of children with autism spectrum disorder (ASD). It has been shown that p-cresol administration induces social behavior deficits and repetitive behavior in mice. However, the mechanisms of p-cresol, specifically its metabolite pCS that can reach the brain, in ASD remain to be investigated. The pCS has been shown to inhibit LPS-stimulated inflammatory response. A Disintegrin And Metalloprotease 10 (ADAM10) and A Disintegrin And Metalloprotease 17 (ADAM17) are thought to regulate microglial immune response by cleaving membrane-bound proteins. In the present study, a neuroinflammation model of LPS-activated BV2 microglia has been used to unveil the potential molecular mechanism of pCS in ASD pathogenesis. In microglial cells pCS treatment decreases the expression or maturation of ADAM10 and ADAM17. In addition, pCS treatment attenuates TNF-α and IL-6 releases as well as phagocytosis activity of microglia. In in vitro ADAM10/17 inhibition experiments, either ADAM10 or ADAM17 inhibition reduces constitutive and LPS-activated release of TNF-α, TNFR-1 and IL-6R by microglial cells, while it increases constitutive and LPS-activated microglial phagocytotic activity. The in vivo results further confirm the involvement of ADAM10 and ADAM17 in ASD pathogenesis. In in utero VPA-exposed male mice, elevated concentration in serum of p-cresol-associated metabolites pCS and p-cresyl glucuronide (pCG) is associated with a VPA-induced increased ADAM10 maturation, and a decreased ADAM17 maturation that is related with attenuated levels of soluble TNF-α and TGF-β1 in the mice brain. Overall, the present study demonstrates a partial role of ADAM10 and ADAM17 in the derailed innate immune response of microglial cells associated with pCS-induced ASD pathogenesis.

Lactobacillus acidophilus KBL409 Reduces Kidney Fibrosis via Immune Modulatory Effects in Mice with Chronic Kidney Disease.

Intestinal dysbiosis has been reported to play an important role in the pathogenesis of various diseases, including chronic kidney disease (CKD). Here, to evaluate whether probiotic supplements can have protective effects against kidney injury in an animal model of CKD is aimed. An animal model of CKD is established by feeding C57BL/6 mice a diet containing 0.2% adenine. These model mice are administered Lactobacillus acidophilus KBL409 daily for 4 weeks. Features of adenine-induce CKD (Ade-CKD) mice, such as prominent kidney fibrosis and higher levels of serum creatinine and albuminuria are improved by administration of KBL409. Ade-CKD mice also exhibit a disrupted intestinal barrier and elevate levels of TNF-α, IL-6, and 8-hydroxy-2'-deoxyguanosine. These changes are attenuated by KBL409. Administration of KBL409 significantly reduces macrophage infiltration and promotes a switch to the M2 macrophage phenotype and increasing regulatory T cells. Notably, the NLRP3 inflammasome pathway is activated in the kidneys of Ade-CKD and decreases by KBL409. In primary kidney tubular epithelial cells treated with p-cresyl sulfate, short-chain fatty acids significantly increase M2 macrophage polarization factors and decrease profibrotic markers. These results demonstrate that supplementation with the probiotic KBL409 has beneficial immunomodulating effects and protects against kidney injury.

Protein/Fiber Index Modulates Uremic Toxin Concentrations in Hemodialysis Patients.

Background: Indoxyl sulfate (IS) and p-cresyl sulfate (PCS), two uremic toxins (UTs), are associated with increased mortality in patients with chronic kidney disease (CKD). These toxins are produced by the microbiota from the diet and excreted by the kidney. The purpose of this study was to analyze the effect of diet on IS and PCS concentration in hemodialysis (HD) patients. Methods: We performed a prospective monocentric study using a seven-day diet record and determination of serum IS and PCS levels in HD patients. We tested the association between toxin concentrations and nutritional data. Results: A total of 58/75 patients (77%) completed the diet record. Mean caloric intake was 22 ± 9.2 kcal/kg/day. The protein/fiber index was 4.9 ± 1.8. No correlation between IS or PCS concentration and protein/fiber index was highlighted. In the 18 anuric patients (31%) in whom residual renal function could not affect toxin concentrations, IS and PCS concentrations were negatively correlated with fiber intake and positively correlated with the protein/fiber index. In a multivariate analysis, IS serum concentration was positively associated with the protein/fiber index (p = 0.03). Conclusions: A low protein/fiber index is associated with low concentrations of uremic toxins in anuric HD patients. Diets with an increased fiber intake must be tested to determine whether they reduce PCS and IS serum concentrations.

The Impact of Synbiotic Treatment on the Levels of Gut-Derived Uremic Toxins, Inflammation, and Gut Microbiome of Chronic Kidney Disease Patients—A Randomized Trial

Altering dysbiotic gut flora through synbiotic supplementation has recently been recognized as a potential treatment strategy to reduce the levels of gut-derived uremic toxins and decrease inflammation. Assessing its efficacy and safety has been the main goal of our randomized, double-blind, placebo-controlled study. A total of 34 nondialyzed chronic kidney disease patients, aged ≥18years, with an estimated glomerular filtration rate between 15 and 45mL/minute, were randomized either to an intervention group (n=17), receiving synbiotic (Lactobacillus acidophilus, Lactobacillus casei, and Bifidobacterium lactis, 32 billion colony forming units per day plus 3.2g of inulin), or control group (n=17), receiving placebo during 12weeks. The impact of treatment on the dynamic of serum levels of gut-derived uremic toxins, total serum indoxyl sulfate, p-cresyl sulfate, and trimethylamine N-oxide, was defined as the primary outcome of the study. Secondary outcomes included changes in the stool microbiome, serum interleukin-6 levels, high-sensitivity C-reactive protein, estimated glomerular filtration rate, albuminuria, diet, gastrointestinal symptom dynamics, and safety. Serum levels of uremic toxins were determined using ultraperformance liquid chromatography. The stool microbiome analysis was performed using the 16S ribosomal ribonucleic acid gene sequencing approach. Synbiotic treatment significantly modified gut microbiome with Bifidobacteria, Lactobacillus, and Subdoligranulum genera enrichment and consequently reduced serum level of indoxyl sulfate (ΔIS -21.5% vs. 5.3%, P<.001), improved estimated glomerular filtration rate (ΔeGFR 12% vs. 8%, P=.029), and decreased level of high-sensitivity C-reactive protein (-39.5 vs. -8.5%, P<.001) in treated patients. Two patients of the intervention arm complained of increased flatulence. No other safety issues were noted. Synbiotics could be available, safe, and an effective therapeutic strategy we could use in daily practice in order to decrease levels of uremic toxins and microinflammation in chronic kidney disease patients.

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.

Contribution of Hypoalbuminemia and Anemia to the Prognostic Value of Plasma p-Cresyl Sulfate and p-Cresyl Glucuronide for Cardiovascular Outcome in Chronic Kidney Disease.

Free plasma concentrations of protein-bound uremic toxins (PBUTs) may be influenced by serum albumin and hemoglobin. The potential association of serum albumin and hemoglobin with free levels of p-cresyl sulfate (pCS) and p-cresyl glucuronide (pCG) and their predictive value for cardiovascular morbidity and mortality were explored. A total of 523 non-dialysis chronic kidney disease (CKD) stages G1–G5 patients were prospectively followed for the occurrence of fatal or non-fatal cardiovascular events over a 5.5-year period. A negative correlation was found between albumin and between hemoglobin, and both total and free pCS and pCG. In multiple linear regression, PBUTs were negatively associated with eGFR (estimated glomerular filtration rate) and hemoglobin but not albumin. In multivariate Cox regression analysis, albumin was a predictor of outcome, independent of pCS and pCG, without interactions between albumin and pCS or pCG. The relation of low hemoglobin with adverse outcome was lost when albumin was entered into the model. Lower concentrations of pCS and pCG are associated with higher serum albumin and hemoglobin. This may indicate that there are two pathways in the blood that potentially contribute to attenuating the vasculotoxic effects of these PBUTs. The association of PBUTs with cardiovascular risk is not explained by albumin levels, which remains a strong and independent predictor for adverse outcome.

Damage of uremic myocardium by p-cresyl sulfate and the ameliorative effect of Klotho by regulating SIRT6 ubiquitination

Uremic cardiomyopathy (UCM) is a common complication in patients with chronic kidney disease (CKD) and an important risk factor for death. P-Cresyl sulfate (PCS) is a damaging factor in UCM, and Klotho is a protective factor. However, the molecular mechanisms of Klotho and PCS in UCM and the relationship between PCS and Klotho are unclear. In vitro, Klotho treatment inhibited PCS-induced cardiomyocyte hypertrophy and apoptosis by blocking mTOR phosphorylation and inhibiting DNA double-strand breaks (DSBs), respectively. Moreover, PCS increased SIRT6 protein ubiquitination and downregulated SIRT6 protein expression, while Klotho inhibited SIRT6 protein ubiquitination and upregulated SIRT6 protein expression. In a mouse model of 5/6 nephrectomy (5/6Nx)-induced UCM, the expression of Klotho in the kidney and serum was decreased, and the expression of SIRT6 protein in myocardial tissues was lower. PCS further reduced Klotho and SIRT6 expression, aggravated heart structure and function abnormalities, and increased myocardial cell apoptosis in UCM mice. Administration of Klotho protein inhibited the downregulation of SIRT6 protein expression and improved cardiac structure and function. Furthermore, serum PCS level was associated with the left ventricular mass (LVM) and left ventricular mass index (LVMI) in hemodialysis patients. In conclusion, the uremic toxin PCS injures cardiomyocytes via mTOR phosphorylation and DSBs, and Klotho antagonizes the damaging effects of PCS. Moreover, the SIRT6 protein plays an important role in UCM, and Klotho suppresses SIRT6 ubiquitination induced by PCS, further improves cardiac structure and function in UCM and exerts protective effects.

STRUCTURAL AND FUNCTIONAL INTESTINAL BARRIER ABNORMALITIES AND CHRONIC KIDNEY DISEASE. LITERATURE REVIEW. PART II

The last few decades have been marked by significant progress in the investigation of the intestinal microbial-tissue complex and its role in the pathogenesis of a wide range of diseases. The presence of intestinal barrier dysfunction has also been confirmed in various nephropathies. Patients with chronic kidney disease (CKD) are characterized by specific alterations of the qualitative and quantitative composition of the gut microbiota. These changes contribute to an increase in the fermentation of food proteins into uremic toxins, such as p-cresyl sulfate, indoxyl sulfate and trimethylaminoxide, disorders of immune tolerance mechanisms of the mucous membrane, disorganization of intestinal epithelium tight junctions, which inevitably leads to an increase of transepithelial permeability. Translocation of bacteria and microbial metabolism products from the intestinal lumen into the systemic circulation is responsible for systemic inflammation, which is currently considered as one of the leading causes of the CKD progression and related complications. However, the exact mechanisms of gut-kidney interaction remain poorly understood. The second part of the review gives a detailed description of the structural and functional disorders of the intestinal muco-epithelial barrier identified in various nephropathies. The mechanisms of uremia-induced intestinal epithelial disruption are discussed, as well as the current therapeutic strategies that may attenuate consequences of intestinal barrier dysfunction in patients with CKD.

Effect of Fatty Acids and Uremic Toxins on the Binding of Nateglinide, an Insulin Secretagogue, to Site II on Human Serum Albumin.

Nateglinide (NAT) is used to treat diabetes, stimulating pancreatic islet β-cells with residual insulin secretory capacity to increase insulin secretion. NAT has been reported to bind to human serum albumin (HSA), but the detail is still unclear. In the current study, we investigated the location and the affinity for the binding of NAT to HSA. Quantitative analysis data from the ultrafiltration experiment indicated that NAT binds strongly to a primary site on HSA with a high affinity. The presence of diazepam (DZP) or ibuprofen (IB), the specific site II ligands of HSA, decreased the binding constants of NAT respectively, without the significant changes in the number of binding sites. Whereas warfarin (WF), a site I specific ligand, did not affect the binding of NAT. Fluorescent replacement experiment showed that NAT replaced dansylsarcosine (DNSS), a site II probe of HSA, but not WF. An increasing level of myristate and uremic toxins, indoxyl sulphate (IS), indoxyl acetate (IA) and p-cresyl sulphate (PCS), during renal disease significantly increased the concentration of unbound NAT. These findings suggest that NAT specifically binds to site II of HSA and the binding capacity and pharmacokinetics of NAT change in renal diseases.

Translational Medicine in Uremic Vascular Calcification: Scavenging ROS Attenuates p-Cresyl Sulfate-Activated Caspase-1, NLRP3 Inflammasome and Eicosanoid Inflammation in Human Arterial Smooth Muscle Cells.

We formerly proved that uremic vascular calcification (UVC) correlates tightly with oxidative elastic lamina (EL) injury and two cell fates (apoptosis and osteocytic conversion) in smooth muscle cells (SMC) of chronic kidney disease (CKD) patients and eliminating p-cresyl sulfate (PCS)-activated intracellular ROS ameliorates the MAPK signaling pathway in a human arterial SMC (HASMC) model. Nonetheless, whether ROS scavenger attenuates PCS-triggered inflammasome activation and eicosanoid inflammation in the UVC process remains unknown. Patients with lower extremity amputation were categorized into CKD and normal control group according to renal function. We used immunohistochemistry stain to analyze UVC in arterial specimens, including oxidative injury (8-hydroxy-2′-deoxyguanosine (8-OHdG) and internal EL disruption), cytosolic phospholipase A2 (cPLA2), cyclooxygenase 2 (COX2), interleukin-1 beta (IL-1β), caspase-1 and NLRP3. To simulate the patho-mechanism of human UVC, the therapeutic effects of ROS scavenger on PCS-triggered inflammatory pathways was explored in a HASMC model. We found CKD patients had higher circulating levels of PCS and an increase in medial arterial calcification than the control group. In CKD arteries, the severity of UVC corresponded with expressions of oxidative EL disruption and 8-OHdG. Furthermore, coupling expressions of cPLA2 and COX2 were accentuated in CKD arteries, indicative of eicosanoid inflammation. Notably, tissue expressions of IL-1β, caspase-1 and NLRP3 were enhanced in parallel with UVC severity, indicative of inflammasome activation. From bedside to bench, ROS scavenger attenuates PCS-activated expressions of cPLA2/COX2, pro-caspase-1 and NLRP3 in the HASMC model. UVC as an inevitable outcome is predictive of death in CKD patients. Nonetheless, UVC remain pharmacoresistant despite the evolution of treatment for mineral-parathyroid hormone-vitamin D axis. Beyond the mineral dysregulation, the stimulation of pro-oxidant PCS alone results in eicosanoid inflammation and inflammasome activation. Concerning the key role of Caspase-1 in pyroptosis, cell fates of HASMC in uremic milieu are not limited to apoptosis and osteogenesis. In view of this, reducing ROS and PCS may act as a therapeutic strategy for UVC-related cardiovascular events in CKD patients.

MO660: Medium Chain Fatty Acids used as Binding Competitors of Albumin Improve the Dialytic Clearance of Protein-Bound Uraemic Toxins

Abstract BACKGROUND AND AIMS Protein-bound uraemic toxins (PBUTs) remain a concerning burden in patients with end-stage renal disease (ESRD) since their removal in haemodialysis (HD) is limited by their strong binding to plasma proteins. By increasing their circulating free fraction during HD, "binding competitors" of albumin could be used to increase their dialytic clearance. Medium-chain fatty acids (MCFAs, 4–10 carbon length) are potent candidates for that purpose. The aim of this work was to evaluate the displacing capacities of MCFAs. METHOD Sodium salts of butanoic acid (C4), hexanoic acid (C6), octanoic acid (C8) and decanoic acid (C10) were purchased from Sigma-Aldrich. The interaction between MCFAs and serum albumin was explored using two fluorescent probes: warfarin (specific to Sudlow's site I) and dansylsarcosine (specific to Sudlow's site II). Indoxyl-sulfate (IS, final concentration 212 µM) and p-cresyl sulfate (p-CS, final concentration 250 µM) were added to a 600 µM bovine serum albumin-phosphate buffered saline solution. The free fractions of IS and p-CS were assayed with or without MCFAs (1–2 mM) using ultrafiltration devices. To mimic the removal of PBUTs during an HD session, batches of 2L of fresh bovine blood were loaded with IS and p-CS (final concentration 200 µM). A 2-h closed-loop HD session was performed using a Fresenius 5008 CorDiax HD generator (Fresenius, Germany). A solution of 224 mM of sodium octanoate (C8) was perfused using the electric syringe pump at the rate of 150 µL/min; a solution of saline 0.9% was used as a control. A total of 1 mL of blood was sampled every 15 min through the arterial sampling port and concentrations of IS and pCS were assayed by HPLC coupled with fluorescence detection. The haemolytic effect of MCFAs was evaluated in vitro by assaying the free haemoglobin concentration. RESULTS Among the short-chain fatty acids tested, octanoic (C8) and decanoic (C10), acids were more prone to displace dansylsarcosine from Sudlow's site II of albumin (which is the main binding site of IS and p-CS). In vitro, the incubation with 2 mM of sodium octanoate or decanoate increased the free fraction of PBUTs from 12 to 53% for p-CS (4.4 folds change, P &amp;lt; .05) and from 11 to 45% for IS (4.1 folds change, P &amp;lt; .05). The per-dialytic infusion of sodium octanoate significantly increased the fractional removal of p-CS (from 38 to 88%, P &amp;lt; .001) and IS (from 36 to 91%, P &amp;lt; .001) (Figure 1). No significant haemolysis was observed for the concentration of MCFAs ˂ 2 mmol/L. CONCLUSION MCFAs and especially C8 and C10 are serious candidates to displace the binding of PBUTs such as p-CS and IS. The per-dialytic administration of MCFA significantly increased the removal of PBUTs and could constitute a new strategy to get rid of these compounds and prevent their accumulation in end-stage kidney disease patients. Due to their safeness (toxicity and metabolism profiles in vivo), MCFAs could be better tolerated than other chemical compounds that have already been tested clinically such as ibuprofen. Further in vivo studies are, however needed to carefully evaluate this potentially new therapeutic option.

MO277: Effect of P-Cresyl Sulfate in Cardiorenal Axis in Mice

Abstract BACKGROUND AND AIMS Uremic toxins (UT) are extremely relevant for the understanding of pathologies of the cardiovascular system once kidney and heart have a close relationship and share different functions. UT are divided into three groups according to physicochemical characteristics: small water-soluble compounds, medium compounds, and protein-bound uremic toxins (PBUT). The PBUT are cytotoxic molecules that have shown deleterious effects in various tissues, including the cardiovascular system. P-Cresyl (PCS), a toxin derived from tyrosine, is one of most important UT once it has a strong affinity for serum proteins, which makes their removal by dialysis difficult. Although some deleterious effects of PCS are known, systematic studies that address the cellular and molecular mechanisms of action of PCS, in the kidney and heart, are still scarce [1, 2]. Moreover, many works in the literature diverge as to the experimental model and the dose of PCS to be used. In this sense, the present study aimed define the dose of PCS capable of leading to renal impairment in mice and also evaluate the effect of PCS alone (without renal injury installed) in kidney and heart. METHODS C57BL/6J male mice 6–8 weeks old (22–28 g) were obtained from the Animal Facility of Federal University of ABC (SP, Brazil). All procedures and protocols were approved by the Ethics Committee on Animal Use of the Federal University of ABC (no. 6 548 291 020). A dose response curve was performed, and the mice were divided into four groups and treated for 21 days, i.p.: Vehicle (saline), PCS 60 mg/L, PCS 40 mg/L and 20 mg/L. Doses were based on uremic maximum concentration available on EUTox-European Uremic Toxins Work Group [3]. After treatment, kidneys, heart, blood, tibia were collected to morphometric and biochemical analysis. Gene expression was evaluated using real-time PCR. RESULTS The EUTox suggested that ∼40 mg/L is the maximum uremic dose for humans. However, the dose–response curve showed that only PCS 20 mg/L was able to increase the mRNA levels of renal injury markers such as Cystatin (3x), KIM-1 (6x) and Vimentin (12x) in relation to control animals (P &amp;lt; .05). Besides, it was possible to observe a decrease on mRNA gene expression of IL-1β levels in kidney (40%, P &amp;lt; .05) when compared with control group. No alterations were observed in MCP-1 levels. In relation to cardiac changes, heart weight/tibia length ratio, showed a cardiac atrophy about 13% after PCS treatment. The OAT 1 and 3 expression did not alter after different doses of PCS treatment. CONCLUSION The results indicated that lowest dose of PCS was able to promote renal injury and cardiac remodeling, despite the changes in OAT1 and OAT3 in kidney and heart. In addition, the establishment of critical dose in mice model is very important since the use of knockout mice models could help to understand the role of UT in organ–organ crosstalk. Finally, this study indicates that the PCS has an impact in kidney/heart axis independent of previous renal injury. The molecular mechanisms involved in cardiac atrophy and inflammatory status are being investigated by our group.

MO488: Serum P-CRESYL Sulfate Level is an Independent Marker of Peripheral Arterial Stiffness as Assessed Using Brachial-Ankle Pulse Wave Velocity in Patients With Non-Dialysis Chronic Kidney Disease Stage 3–5

Abstract BACKGROUND AND AIMS p-cresyl sulfate (PCS) is associated with inflammation and its contribution to cardiovascular damage in patients with chronic kidney disease (CKD). The brachial-ankle pulse wave velocity (baPWV) is a simple marker to assess the peripheral artery stiffness (PAS) and is a useful marker to predict future cardiovascular events. The aim of this study was to determine the relationship between serum PCS levels and PAS measured by baPWV values in patients with CKD stage 3–5. METHOD Fasting blood samples and baseline characteristics were obtained from 160 CKD stage 3–5 patients. Automatic pulse wave analyzer (VaSera VS-1000) to measure baPWV values, and both sides of baPWV value that was &amp;gt;18.0 m/s were determined as PAS. Serum total PCS levels were determined by high-performance liquid chromatography–mass spectrometry. RESULTS In the study cohort, 54 patients (33.8%) were in the PAS group. When compared with those in the control group, the PAS group had a high prevalence of hypertension (P = .021), older age (P &amp;lt; .001), lower height (P = .033) and body weight (P = .012), higher systolic blood pressure (P &amp;lt; .001), diastolic blood pressure (P = .008), serum calcium–phosphorus product levels (P = .030) and PCS levels (P = .008). The multivariable logistic regression analysis revealed that serum levels of PCS {odds ratio (OR): 1.098, [95% confidence interval (95% CI): 1.029–1.171]; P = .005}, advanced age (OR: 1.105, 95% CI: 1.055–1.159; P &amp;lt; .001) and diastolic blood pressure (OR: 1.058, 95% CI: 1.002–1.118; P = .043) were independently associated with PAS in patients with CKD stage 3–5. CONCLUSION Higher serum PCS levels, diastolic blood pressure and advanced age were associated with PAS in patients with CKD stage 3–5.

Gut Microbiota Mediates the Protective Effects of Traditional Chinese Medicine Formula Qiong-Yu-Gao against Cisplatin-Induced Acute Kidney Injury.

ABSTRACTOur previous study found that Qiong-Yu-Gao (QYG), a traditional Chinese medicine formula derived from Rehmanniae Radix, Poria, and Ginseng Radix, has protective effects against cisplatin-induced acute kidney injury (AKI), but the underlying mechanisms remain unknown. In the present study, the potential role of gut microbiota in the nephroprotective effects of QYG was investigated. We found that QYG treatment significantly attenuated cisplatin-induced AKI and gut dysbiosis, altered the levels of bacterial metabolites, with short-chain fatty acids (SCFAs) such as acetic acid and butyric acid increasing and uremic toxins such as indoxyl sulfate and p-cresyl sulfate reducing, and suppressed histone deacetylase expression and activity. Spearman’s correlation analysis found that QYG-enriched fecal bacterial genera Akkermansia, Faecalibaculum, Bifidobacterium, and Lachnospiraceae_NK4A136_group were correlated with the altered metabolites, and these metabolites were also correlated with the biomarkers of AKI, as well as the indicators of fibrosis and inflammation. The essential role of gut microbiota was further verified by both the diminished protective effects with antibiotics-induced gut microbiota depletion and the transferable renal protection with fecal microbiota transplantation. All these results suggested that gut microbiota mediates the nephroprotective effects of QYG against cisplatin-induced AKI, potentially via increasing the production of SCFAs, thus suppressing histone deacetylase expression and activity, and reducing the accumulation of uremic toxins, thereby alleviating fibrosis, inflammation, and apoptosis in renal tissue.IMPORTANCE Cisplatin-induced acute kidney injury is the main limiting factor restricting cisplatin’s clinical application. Accumulating evidence indicated the important role of gut microbiota in pathogenesis of acute kidney injury. In the present study, we have demonstrated that gut microbiota mediates the protective effects of traditional Chinese medicine formula Qiong-Yu-Gao against cisplatin-induced acute kidney injury. The outputs of this study would provide scientific basis for future clinical applications of QYG as prebiotics to treat cisplatin-induced acute kidney injury, and gut microbiota may be a promising therapeutic target for chemotherapy-induced nephrotoxicity.