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INTRODUCTION: Trimethylamine (TMA), produced by gut microbiota, is the precursor of trimethylamine-N-oxide (TMAO), a uremic toxin that accumulates in patients with chronic kidney disease (CKD). Elevated TMAO plasma levels are associated with cardiovascular complications and CKD progression. OBJECTIVE: To evaluate the association between gut microbiota composition and TMAO plasma levels in CKD patients undergoing hemodialysis (HD). METHODS: This is a cross-sectional study with 25 patients evaluated (60% female, 53 (18) years, body mass index (BMI) 25.8 (6.75) Kg/m2). They were divided into two groups according to their TMAO plasma levels: normal (≤ 7.4 µM) and high (> 7.4 µM). Uremic toxins such as indoxyl sulfate (IS), p-cresyl sulfate (pCS), and indol acetic acid (IAA) were measured with RP-HPLC, and TMAO plasma levels were quantified using LC-MS/MS. Fecal DNA was extracted with a commercial kit, PCR amplified the V4 region of the 16S rRNA gene, and short-read sequencing was performed on the Illumina platform. Dietary intake, anthropometric measurements, and inflammation markers were also evaluated. Nrf2, NF-κB, IL-1ß, and NLRP3 mRNA expressions were measured from peripheral blood mononuclear cells (PBMC) using quantitative real-time polymerase chain reaction (qPCR). RESULTS: There were significant positive correlations between TMAO and plasma levels of pCS, NLPR3 inflammasome mRNA expression, serum phosphorus levels, and negative correlations with dietary lipid intake. The group with TMAO > 7.4 µM showed an increase in the microbiome abundance of Saccharibacteria (genus incertae sedis), Colidextribacter, Dorea, and Staphylococci genera, and a decrease in abundance in the genera Lachnospira, Lactobacilli, and Victivallis. TMAO plasma level was positively correlated with the abundance of bacteria of the genera Colidextribacter and Helicobacter and was negatively correlated with Sphingomanos, Lachnospira, Streptomyces, and Bacillus genera. CONCLUSION: Saccharibacteria (genus incertae sedis), Colidextribacter, Dorea, and Staphylococci genera showed higher abundance in patients with high TMAO levels. In addition, we observed that elevated plasma TMAO levels are associated with inflammation markers, dietary lipid intake, and serum phosphorus levels in patients undergoing HD.
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Sulforaphane (SFN), found in cruciferous vegetables, is a known activator of NRF2 (master regulator of cellular antioxidant responses). Patients with chronic kidney disease (CKD) present an imbalance in the redox state, presenting reduced expression of NRF2 and increased expression of NF-κB. Therefore, this study aimed to evaluate the effects of SFN on the mRNA expression of NRF2, NF-κB and markers of oxidative stress in patients with CKD. Here, we observed a significant increase in the mRNA expression of NRF2 (p = 0.02) and NQO1 (p = 0.04) in the group that received 400 µg/day of SFN for 1 month. Furthermore, we observed an improvement in the levels of phosphate (p = 0.02), glucose (p = 0.05) and triglycerides (p = 0.02) also in this group. On the other hand, plasma levels of LDL-c (p = 0.04) and total cholesterol (p = 0.03) increased in the placebo group during the study period. In conclusion, 400 µg/day of SFN for one month improves the antioxidant system and serum glucose and phosphate levels in non-dialysis CKD patients.
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Isotiocianatos , NAD(P)H Desidrogenase (Quinona) , Fator 2 Relacionado a NF-E2 , Estresse Oxidativo , RNA Mensageiro , Insuficiência Renal Crônica , Sulfóxidos , Humanos , Isotiocianatos/farmacologia , Isotiocianatos/uso terapêutico , Fator 2 Relacionado a NF-E2/metabolismo , Fator 2 Relacionado a NF-E2/genética , NAD(P)H Desidrogenase (Quinona)/genética , NAD(P)H Desidrogenase (Quinona)/metabolismo , Insuficiência Renal Crônica/metabolismo , Insuficiência Renal Crônica/genética , Insuficiência Renal Crônica/tratamento farmacológico , Insuficiência Renal Crônica/patologia , Masculino , Pessoa de Meia-Idade , Feminino , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Triglicerídeos/sangue , Triglicerídeos/metabolismo , Glicemia/metabolismo , Regulação para Cima/efeitos dos fármacos , Adulto , Idoso , NF-kappa B/metabolismo , NF-kappa B/genéticaRESUMO
Quiescin/sulfhydryl oxidase (QSOX1) is a secreted flavoprotein that modulates cellular proliferation, migration and adhesion, roles attributed to its ability to organize the extracellular matrix. We previously showed that exogenously added QSOX1b induces smooth muscle cells migration in a process that depends on its enzymatic activity and that is mediated by hydrogen peroxide derived from Nox1, a catalytic subunit of NAD(P)H oxidases. Here, we report that exogenous QSOX1b also stimulates the migration of L929 fibroblasts and that this effect is regulated by its endocytosis. The use of endocytosis inhibitors and caveolin 1-knockdown demonstrated that this endocytic pathway is caveola-mediated. QSOX1b colocalized with Nox1 in intracellular vesicles, as detected by confocal fluorescence, suggesting that extracellular QSOX1b is endocytosed with the transmembrane Nox1. These results reveal that endosomal QSOX1b is a novel intracellular redox regulator of cell migration.
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Cavéolas , NADPH Oxidases , Fibroblastos , Endocitose , Proliferação de CélulasRESUMO
BACKGROUND & AIMS: Turmeric (a source of curcumin) is an excellent food to modulate oxidative stress, inflammation, and gut dysbiosis in patients with chronic kidney disease (CKD). However, no studies report the benefits of curcumin in patients undergoing peritoneal dialysis (PD). This study aims to evaluate the effects of curcuminoid supplementation on oxidative stress, inflammatory markers, and uremic toxins originating from gut microbiota in patients with CKD undergoing PD. METHODS: This longitudinal, randomized, single-blind, placebo-controlled trial evaluated 48 patients who were randomized into two groups: Curcumin (three capsules of 500 mg of Curcuma longa extract, with 98.42 % total curcuminoids) or placebo (three capsules of 500 mg of starch) for twelve weeks. In the peripheral blood mononuclear cells (PBMCs), the transcriptional expression levels of Nrf2, HOX-1 and NF-κB were evaluated by quantitative real-time PCR. Oxidative stress was evaluated by malondialdehyde (MDA) and total Thiol (T-SH). TNF-α and IL-6 plasma levels were measured by ELISA. P-cresyl sulphate plasma level, a uremic toxin, was evaluated by high-performance liquid chromatography (HPLC) with fluorescent detection. RESULTS: Twenty-four patients finished the study: 10 in the curcumin group (57.5 ± 11.6 years) and 14 in the placebo group (56.5 ± 10.0 years). The plasma levels of MDA were reduced after 12 weeks in the curcumin group (p = 0.01), while the placebo group remained unchanged. However, regarding the difference between the groups at the endpoint, no change was observed in MDA. Still, there was a trend to reduce the p-CS plasma levels in the curcumin group compared to the placebo group (p = 0.07). Likewise, the concentrations of protein thiols, mRNA expression of Nrf2, HOX-1, NF-κB, and cytokines plasma levels did not show significant changes. CONCLUSION: Curcuminoid supplementation for twelve weeks attenuates lipid peroxidation and might reduce uremic toxin in patients with CKD undergoing PD. This study was registered on Clinicaltrials.gov as NCT04413266.
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Curcumina , Diálise Peritoneal , Insuficiência Renal Crônica , Uremia , Humanos , Curcumina/farmacologia , Curcumina/uso terapêutico , NF-kappa B/metabolismo , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Leucócitos Mononucleares/metabolismo , Método Simples-Cego , Inflamação , Estresse Oxidativo , Insuficiência Renal Crônica/complicações , Insuficiência Renal Crônica/terapia , Diarileptanoides/farmacologia , Diarileptanoides/uso terapêutico , Suplementos Nutricionais , Uremia/tratamento farmacológicoRESUMO
Secreted quiescin/sulfhydryl oxidase (QSOX) is overexpressed in many tumor cell lines, including melanoma, and is usually associated with a pro-invasive phenotype. Our previous work described that B16-F10 cells enter in a quiescent state as a protective mechanism against damage generated by reactive oxygen species (ROS) during melanogenesis stimulation. Our present results show that QSOX activity was two-fold higher in cells with stimulated melanogenesis when compared to control cells. Considering that glutathione (GSH) is one of the main factor responsible for controlling redox homeostasis in cells, this work also aimed to investigate the relationship between QSOX activity, GSH levels and melanogenesis stimulation in B16-F10 murine melanoma cell line. The redox homeostasis was impaired by treating cells with GSH in excess or depleting its intracellular levels through BSO treatment. Interestingly, GSH-depleted cells without stimulation of melanogenesis kept high levels of viability, suggesting a possible adaptive mechanism of survival even under low GSH levels. They also showed lower extracellular activity of QSOX, and higher QSOX intracellular immunostaining, suggesting that this enzyme was less excreted from cells and corroborating with a diminished extracellular QSOX activity. On the other hand, cells under melanogenesis stimulation showed a lower GSH/GSSG ratio (8:1) in comparison with control (non-stimulated) cells (20:1), indicating a pro-oxidative state after stimulation. This was accompanied by decreased cell viability after GSH-depletion, no alterations in QSOX extracellular activity, but higher QSOX nucleic immunostaining. We suggest that melanogenesis stimulation and redox impairment caused by GSH-depletion enhanced the oxidative stress in these cells, contributing to additional alterations of its metabolic adaptive response.
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Melanoma , Oxirredutases atuantes sobre Doadores de Grupo Enxofre , Animais , Camundongos , Glutationa/metabolismo , Melanoma/metabolismo , Oxirredução , Oxirredutases/metabolismo , Espécies Reativas de Oxigênio , Oxirredutases atuantes sobre Doadores de Grupo Enxofre/metabolismoRESUMO
BACKGROUND: Patients with chronic kidney disease (CKD) have reduced expression of erythroid nuclear factor-related factor 2 (NRF2) and increased nuclear factor κB (NF-κB). "Food as medicine" has been proposed as an adjuvant therapeutic alternative in modulating these factors. No studies have investigated the effects of sulforaphane (SFN) in cruciferous vegetables on the expression of these genes in patients with CKD. OBJECTIVE: The study aimed to evaluate the effects of SFN on the expression of NRF2 and NF-κB in patients on hemodialysis (HD). DESIGN AND METHODS: A randomized, double-blind, crossover study was performed on 30 patients on regular HD. Fourteen patients were randomly allocated to the intervention group (1 sachet/day of 2.5 g containing 1% SFN extract with 0.5% myrosinase) and 16 patients to the placebo group (1 sachet/day of 2.5 g containing corn starch colored with chlorophyll) for 2 months. After a washout period of 2 months, the groups were switched. NRF2 and NF-κB mRNA expression was evaluated by real-time quantitative polymerase chain reaction, and tumor necrosis factor alpha and interleukin-6 levels were quantified by enzyme-linked immunosorbent assay. Malondialdehyde was evaluated as a marker of lipid peroxidation. RESULTS: Twenty-five patients (17 women, 55 [interquartile range = 19] years and 55 [interquartile range = 74] months on HD) completed the study. There was no significant difference concerning the expression of mRNA NRF2 (P = .915) and mRNA NF-κB (P = .806) after supplementation with SFN. There was no difference in pro-inflammatory and oxidative stress biomarkers. CONCLUSION: 150 µmol of SFN for 2 months had no antioxidant and anti-inflammatory effect in patients with CKD undergoing HD.
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Isotiocianatos , NF-kappa B , Insuficiência Renal Crônica , Sulfóxidos , Humanos , Feminino , NF-kappa B/genética , NF-kappa B/metabolismo , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Estudos Cross-Over , Estresse Oxidativo , Diálise Renal/efeitos adversos , Insuficiência Renal Crônica/tratamento farmacológico , Insuficiência Renal Crônica/etiologia , RNA Mensageiro/metabolismo , RNA Mensageiro/farmacologia , Suplementos NutricionaisRESUMO
The precise mechanisms underlying the cardiovascular complications due to acute kidney injury (AKI) and the retention of uremic toxins like p-cresyl sulfate (PCS) remain incompletely understood. The objective of this study was to evaluate the renocardiac effects of PCS administration in animals subjected to AKI induced by ischemia and reperfusion (IR) injury. C57BL6 mice were subjected to distinct protocols: (i) administration with PCS (20, 40, or 60 mg/L/day) for 15 days and (ii) AKI due to unilateral IR injury associated with PCS administration for 15 days. The 20 mg/L dose of PCS led to a decrease in renal mass, an increase in the gene expression of Cystatin C and kidney injury molecule 1 (KIM-1), and a decrease in the α-actin in the heart. During AKI, PCS increased the renal injury biomarkers compared to control; however, it did not exacerbate these markers. Furthermore, PCS did not enhance the cardiac hypertrophy observed after 15 days of IR. An increase, but not potentialized, in the cardiac levels of interleukin (IL)-1ß and IL-6 in the IR group treated with PCS, as well as in the injured kidney, was also noticed. In short, PCS administration did not intensify kidney injury, inflammation, and cardiac outcomes after AKI.
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Injúria Renal Aguda , Traumatismo por Reperfusão , Animais , Camundongos , Sulfatos , Camundongos Endogâmicos C57BL , Rim , Isquemia/complicações , Traumatismo por Reperfusão/complicaçõesRESUMO
Oxidative stress (OS) is essential in uremia-associated comorbidities, including renal anemia. Complications experienced by hemodialysis (HD) patients, such as hypoxemia and uremic toxins accumulation, induce OS and premature death of red blood cells (RBC). We aimed to characterize reactive oxygen species (ROS) production and antioxidant pathways in HD-RBC and RBC from healthy controls (CON-RBC) and evaluate the role of uremia and hypoxia in these pathways. ROS production, xanthine oxidase (XO) and superoxide dismutase (SOD) activities, glutathione (GSH), and heme oxygenase-1 (HO-1) levels were measured using flow cytometry or spectrophotometry in CON-RBC and HD-RBC (pre- and post-HD), at baseline and after 24 h incubation with uremic serum (S-HD) and/or under hypoxic conditions (5% O2 ). Ketoprofen was used to inhibit RBC uremic toxins uptake. HD-RBC showed higher ROS levels and lower XO activity than CON-RBC, particularly post-HD. GSH levels were lower, while SOD activity and HO-1 levels of HD-RBC were higher than control. Hypoxia per se triggered ROS production in CON-RBC and HD-RBC. S-HD, on top of hypoxia, increased ROS levels. Inhibition of uremic toxins uptake attenuated ROS of CON and HD-RBC under hypoxia and uremia. CON-RBC in uremia and hypoxia showed lower GSH levels than cells in normoxia and non-uremic conditions. Redox mechanisms of HD-RBC are altered and prone to oxidation. Uremic toxins and hypoxia play a role in unbalancing these systems. Hypoxia and uremia participate in the pathogenesis of OS in HD-RBC and might induce RBC death and thus compound anemia.
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Anemia , Uremia , Humanos , Eritrócitos/metabolismo , Uremia/metabolismo , Diálise Renal , Estresse Oxidativo , Glutationa/metabolismo , Hipóxia/metabolismo , Anemia/metabolismoRESUMO
BACKGROUND: Although high-volume online hemodiafiltration has been associated with higher clearance and lower pre-dialysis concentration of middle molecular weight toxins compared to hemodialysis, its effect on protein-bound uremic toxins has shown inconclusive results. In this study, we investigated whether hemodiafiltration impacts pre-dialysis plasma levels of the toxins indoxyl sulfate, p-cresyl sulfate, and indole-3-acetic acid compared to high-flux hemodialysis. METHODS: This is a post-hoc analysis of the multicenter, randomized controlled trial HDFit (ClinicalTrials.gov: NCT02787161). Uremic toxins were determined by high performance liquid chromatography at baseline, 3, and 6 months. Mean differences in monthly changes of pre-dialysis uremic toxin concentrations between hemodiafiltration and high-flux hemodialysis were analyzed using linear mixed-effect models. RESULTS: One hundred ninety-three patients (mean age 53 years old, 71% males) were analyzed. There were no differences between groups regarding clinical and biochemical characteristics at baseline or duration of dialysis session and blood flows throughout the follow-up. Mean differences in rates of change (µM/month, [confidence interval CI]) in high-flux hemodialysis vs. hemodiafiltration were 2.4 [0.3 to 4.56], 3.94 [- 1.54 to 9.41] and 0.06 [- 0.6 to 0.5] for indoxyl sulfate, p-cresyl sulfate and indole-3-acetic acid, respectively. In the exploratory analysis, these differences in high-flux hemodialysis vs. hemodiafiltration subgroup with convective volume > 27.5 L were 2.86 [0.43 to 5.28], 7.43 [0.7 to 14.16] and - 0.19 [- 0.88 to 0.50]. CONCLUSION: These exploratory findings suggest that hemodiafiltration is more effective in reducing indoxyl sulfate as compared to standard high-flux hemodialysis, and also that this effect was extended to p-cresyl sulfate in patients achieving higher convective volumes.
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Hemodiafiltração , Indicã , Diálise , Feminino , Hemodiafiltração/métodos , Humanos , Masculino , Pessoa de Meia-Idade , Diálise Renal/efeitos adversos , Diálise Renal/métodos , SulfatosRESUMO
OBJECTIVES: Uremic toxins such as indoxyl sulfate (IS), p-cresyl sulfate (pCS), and indole-3-acetic acid (IAA) produced by the gut microbiota are recognized as risk factors for many comorbidities, including cardiovascular diseases. Patients with chronic kidney disease (CKD) have an accumulation of these toxins, and nutritional strategies have been proposed to mitigate gut dysbiosis and, consequently, reduce these toxins. This study aimed to evaluate the effects of resveratrol supplementation on the plasma levels of IS, pCS, and IAA in nondialyzed patients with CKD. METHODS: In this placebo-controlled crossover study, twenty nondialyzed patients were randomly divided into two groups: they received either one capsule/day containing 500 mg of trans-resveratrol (63 ± 7.5 years, glomerular filtration rate [GFR]: 34 ± 14 mL/min, body mass index: 26.8 ± 5.6 kg/m2) or a placebo containing 500 mg wheat flour (62 ± 8.4 years, GFR: 34 ± 13 mL/min, body mass index: 28.6 ± 4.4 kg/m2) during 4 weeks. After 8 weeks of washout (no supplementation), another 4 weeks of supplementation with crossover was initiated. IS, IAA, and pCS plasma levels were quantified by the reverse phase high-efficiency liquid chromatography method with fluorescent detection. The mRNA expression of nuclear factor erythroid 2-related factor 2 and nuclear factor kappa B in peripheral blood mononuclear cells was evaluated by polymerase chain reaction. C-reactive protein plasma levels were also evaluated. RESULTS: As expected, the uremic toxin levels were negatively correlated with the GFR, but no effect of trans-resveratrol supplementation was found on levels of IS, IAA, and pCS. There was a positive correlation between IS and nuclear factor erythroid 2-related factor 2 (r = 0.24, P = .03) and also between IS and C-reactive protein (r = 0.21, P = .05). CONCLUSION: Supplementation with trans-resveratrol did not reduce the plasma levels of IS, pCS, and IAA in nondialyzed patients with CKD. The interactions among uremic toxins and anti-inflammatory and proinflammatory pathways deserve more studies.
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Microbioma Gastrointestinal , Insuficiência Renal Crônica , Humanos , Resveratrol , Toxinas Urêmicas , Proteína C-Reativa , Leucócitos Mononucleares/metabolismo , Estudos Cross-Over , Farinha , Triticum , IndicãRESUMO
BACKGROUND & AIMS: Patients with Chronic Kidney Disease (CKD) have an imbalance in the gut microbiota that can lead to increase levels of lipopolysaccharides (LPS) and uremic toxins such as indoxyl sulfate (IS), p-cresyl sulfate (p-CS), and indole-3 acetic acid (IAA). Among the therapeutic options for modulating gut microbiota are the bioactive compounds such as polyphenols present in cranberry, fruit with potential antioxidant and anti-inflammatory effects. This clinical trial focuses on evaluating the effects of supplementation with a dry extract of cranberry on plasma levels of LPS and uremic toxins in non-dialysis CKD patients. METHODS: It was a randomized, double-blind, placebo-controlled study. Patients were randomized into two groups: the cranberry group received 500 mg of dry cranberry extract (2 times daily), and the placebo group received 500 mg of corn starch (2 times daily) for two months. LPS plasma levels were evaluated by enzyme-linked immunosorbent assay (ELISA) and uremic toxins (IS, p-CS, and IAA) by high-performance liquid chromatography-fluorescence detection. Anthropometric measurements and food intake using the 24-h food recall technique were also evaluated before and after the intervention. RESULTS: Twenty-five participants completed two months of supplementation: 12 patients in the cranberry group (8 women, 56.7 ± 7.5 years, estimated glomerular filtration rate (eGFR) of 39.2 ± 21.9 mL/min); 13 patients in the placebo group (9 women, 58.8 ± 5.1 years, eGFR of 39.7 ± 12.9 mL/min). As expected, there was a negative association between glomerular filtration rate and p-CS and IS plasma levels at the baseline. No change was observed in the uremic toxins and LPS levels. CONCLUSION: Cranberry dry extract supplementation for two months did not reduce the LPS and uremic toxins plasma levels produced by the gut microbiota in non-dialysis CKD patients.
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Microbioma Gastrointestinal , Insuficiência Renal Crônica , Vaccinium macrocarpon , Suplementos Nutricionais , Feminino , Frutas , Humanos , Projetos Piloto , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêuticoRESUMO
BACKGROUND: Anemia is one of the most frequent complications in patients with chronic kidney disease (CKD). Despite being multifactorial, the relative or absolute deficiency of erythropoietin production is the leading cause. Recent studies have shown that uremic toxins produced by the gut microbiota also may play a role in the genesis of anemia in these patients. OBJECTIVE: To evaluate the possible association between uremic toxins plasma levels and anemia in patients with CKD on hemodialysis (HD). METHODS: This cross-sectional study evaluated one hundred fifty-four patients (53.2% men, 51.2 ± 11.2 years, hemoglobin (Hb) levels of 11.2 ± 1.6 g/dL). Biochemical variables such as urea, creatinine, hemoglobin, hematocrit, were measured according to standard methods and uremic toxins such as indoxyl sulfate (IS), indole-3-acetic acid (IAA), p-cresyl sulfate (p-CS) plasma levels were measured by reverse-phase high-performance liquid chromatography (RP-HPLC). RESULTS: The levels of uremic toxins such as IS, IAA, p-CS were increased in all patients. However, no correlation was found between uremic toxins plasma levels and anemia parameters. Only patients with Hb < 11 g/dL presented a negative correlation between hematocrit and IAA plasma levels. CONCLUSION: There is no strong evidence that uremic toxins produced by the gut microbiota may be associated with anemia in patients with CKD on HD.
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Anemia , Microbioma Gastrointestinal , Insuficiência Renal Crônica , Uremia , Anemia/complicações , Estudos Transversais , Feminino , Humanos , Indicã , Masculino , Diálise Renal/efeitos adversos , Diálise Renal/métodos , Insuficiência Renal Crônica/complicações , Insuficiência Renal Crônica/terapia , Uremia/complicações , Uremia/terapia , Toxinas UrêmicasRESUMO
PURPOSE: Regular physical exercise may result in many benefits to patients with chronic kidney disease (CKD) on hemodialysis (HD), including gut microbiota modulation and solute removal. The study aimed to evaluate the effects of two programs of intradialytic exercises on uremic toxins plasma levels in HD patients. METHODS: In experiment 1, twenty HD patients [12 men, 44.1 ± 8.9 years, BMI of 23.4 ± 2.4 kg/m2] were randomized into two groups: Aerobic exercise group (AEG, n = 11) that performed aerobic exercise on an adapted exercise bike three times a week for three months (36 sessions) and Control group (CG, n = 9). In experiment 2, twenty-six HD patients [19 men, 47.6 ± 11.0 years, BMI of 25.9 ± 3.6 kg/m2] were randomized into Resistance exercise group (REG, n = 14) that performed a resistance exercise program (using elastic bands and ankle cuffs with both lower limbs) monitored three times a week, during six months (72 sessions) and CG (n = 12). P-cresyl sulfate (p-CS), indoxyl sulfate (IS), and indol-3-acetic acid (IAA) plasma levels were determined by high-performance liquid chromatography (HPLC) with fluorescent detection. RESULTS: The uremic toxins plasma levels did not reduce in both exercise programs, aerobic exercise (IS: 32.7 ± 14.0 vs 33.0 ± 15.4 mg/L, p = 0.86; p-CS: 59.9 ± 39.3 vs 60.0 ± 41.2 mg/L, p = 0.99; IAA: 2233 [1488-2848] vs 2227 [1275-2824] µg/L, p = 0.72) and resistance exercise (IS: 28.3 ± 11.3 vs 29.1 ± 9.7 mg/L, p = 0.77; p-CS: 31.4 ± 21.3 vs 34.2 ± 19.8 mg/L, p = 0.63; IAA: 1628 [1330-3530] vs 2000 [971-3085] µg/L, p = 0.35) in HD patients. CONCLUSION: According to our findings, physical exercise does not appear to alter the levels of uremic toxins produced by the gut microbiota in HD patients.
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Exercício Físico , Microbioma Gastrointestinal , Diálise Renal , Insuficiência Renal Crônica/sangue , Insuficiência Renal Crônica/terapia , Toxinas Urêmicas/sangue , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
BACKGROUND/AIMS: Chronic kidney disease is frequently accompanied by anemia, hypoxemia, and hypoxia. It has become clear that the impaired erythropoietin production and altered iron homeostasis are not the sole causes of renal anemia. Eryptosis is a process of red blood cells (RBC) death, like apoptosis of nucleated cells, characterized by Ca2+ influx and phosphatidylserine (PS) exposure to the outer RBC membrane leaflet. Eryptosis can be induced by uremic toxins and occurs before senescence, thus shortening RBC lifespan and aggravating renal anemia. We aimed to assess eryptosis and intracellular oxygen levels of RBC from hemodialysis patients (HD-RBC) and their response to hypoxia, uremia, and uremic toxins uptake inhibition. METHODS: Using flow cytometry, RBC from healthy individuals (CON-RBC) and HD-RBC were subjected to PS (Annexin-V), intracellular Ca2+ (Fluo-3/AM) and intracellular oxygen (Hypoxia Green) measurements, at baseline and after incubation with uremic serum and/or hypoxia (5% O2), with or without ketoprofen. Baseline levels of uremic toxins were quantified in serum and cytosol by high performance liquid chromatography. RESULTS: Here, we show that HD-RBC have less intracellular oxygen and that it is further decreased post-HD. Also, incubation in 5% O2 and uremia triggered eryptosis in vitro by exposing PS. Hypoxia itself increased the PS exposure in HD-RBC and CON-RBC, and the addition of uremic serum aggravated it. Furthermore, inhibition of the organic anion transporter 2 with ketoprofen reverted eryptosis and restored the levels of intracellular oxygen. Cytosolic levels of the uremic toxins pCS and IAA were decreased after dialysis. CONCLUSION: These findings suggest the participation of uremic toxins and hypoxia in the process of eryptosis and intracellular oxygenation.
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Eriptose , Eritrócitos/metabolismo , Oxigênio/sangue , Insuficiência Renal Crônica/sangue , Uremia/sangue , Adolescente , Adulto , Idoso , Anexina A5/sangue , Cálcio/sangue , Hipóxia Celular , Eritrócitos/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Insuficiência Renal Crônica/patologia , Uremia/patologiaRESUMO
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.
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Intestinos/química , Musa , Adulto , Idoso , Estudos Cross-Over , Método Duplo-Cego , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Diálise Peritoneal , Diálise Renal , Insuficiência Renal Crônica , Toxinas BiológicasRESUMO
BACKGROUND: Gut dysbiosis is common in patients with chronic kidney disease (CKD) and is closely related to inflammatory processes. Some nutritional strategies, such as bioactive compounds present in curcumin, have been proposed as an option to modulate the gut microbiota and decrease the production of uremic toxins such as indoxyl sulfate (IS), p-cresyl sulfate (pCS) and indole-3 acetic acid (IAA). OBJECTIVE: To evaluate the effects of curcumin supplementation on uremic toxins plasma levels produced by gut microbiota in patients with CKD on hemodialysis (HD). METHODS: Randomized, double-blind trial in 28 patients [53.6 ± 13.4 years, fourteen men, BMI 26.7 ± 3.7 kg/m2, dialysis vintage 37.5 (12-193) months]. Fourteen patients were randomly allocated to the curcumin group and received 100 mL of orange juice with 12 g carrot and 2.5 g of turmeric and 14 patients to the control group who received the same juice but without turmeric three times per week after HD sessions for three months. IS, pCS, IAA plasma levels were measured by reverse-phase high-performance liquid chromatography RESULTS: After three months of supplementation, the curcumin group showed a significant decrease in pCS plasma levels [from 32.4 (22.1-45.9) to 25.2 (17.9-37.9) mg/L, p = 0.009], which did not occur in the control group. No statistical difference was observed in IS and IAA levels in both groups. CONCLUSION: The oral supplementation of curcumin for three months seems to reduce p-CS plasma levels in HD patients, suggesting a gut microbiota modulation.
Assuntos
Cresóis/sangue , Curcumina/uso terapêutico , Suplementos Nutricionais , Microbioma Gastrointestinal , Indicã/sangue , Ácidos Indolacéticos/sangue , Diálise Renal , Ésteres do Ácido Sulfúrico/sangue , Toxinas Biológicas/sangue , Uremia/sangue , Adulto , Idoso , Método Duplo-Cego , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Projetos PilotoRESUMO
A Disintegrin And Metalloprotease 23 (ADAM23) is a member of the ADAMs family of transmembrane proteins, mostly expressed in nervous system, and involved in traffic and stabilization of Kv1-potassium channels, synaptic transmission, neurite outgrowth, neuronal morphology and cell adhesion. Also, ADAM23 has been linked to human pathological conditions, such as epilepsy, cancer metastasis and cardiomyopathy. ADAM23 functionality depends on the molecule presence at the cell surface and along the secretory pathway, as expected for a cell surface receptor. Because endocytosis is an important functional regulatory mechanism of plasma membrane receptors and no information is available about the traffic or turnover of non-catalytic ADAMs, we investigated ADAM23 internalization, recycling and half-life properties. Here, we show that ADAM23 undergoes constitutive internalization from the plasma membrane, a process that depends on lipid raft integrity, and is redistributed to intracellular vesicles, especially early and recycling endosomes. Furthermore, we observed that ADAM23 is recycled from intracellular compartments back to the plasma membrane and thus has longer half-life and higher cell surface stability compared with other ADAMs. Our findings suggest that regulation of ADAM23 endocytosis/stability could be exploited therapeutically in diseases in which ADAM23 is directly involved, such as epilepsy, cancer progression and cardiac hypertrophy.
Assuntos
Proteínas ADAM/metabolismo , Endocitose , Membrana Celular/metabolismo , Células Cultivadas , Endossomos/metabolismo , Meia-Vida , Humanos , Microdomínios da Membrana/metabolismoRESUMO
ABSTRACT Background: Patients with chronic kidney disease (CKD) present an imbalance of the gut microbiota composition, leading to increased production of uremic toxins like p-cresyl sulfate (PCS), product from bacterial fermentation of the amino acids tyrosine (Tyr) and phenylalanine (Phe) from the diet. Thus, diet may be a determinant in the uremic toxins levels produced by the gut microbiota. The aim of this study was to evaluate the possible relationship between Tyr and Phe intake and PCS plasma levels in non-dialysis CKD patients. Methods: Twenty-seven non-dialysis CKD patients (stages 3 and 4) without previous nutritional intervention were evaluated. The dietary intake was evaluated using a 24-hour recall, 3-day food record and protein intake was also estimated by Protein Nitrogen Appearance (PNA). The plasma levels of PCS were measured using reverse phase high performance liquid chromatography. Results: The evaluated patients (GRF, 34.8 ± 12.4 mL/min, 54.2 ± 14.3 years, BMI, 29.3 ± 6.1 kg/m2) presented mean protein intake of 1.1 ± 0.5 g/kg/day), Tyr of 4.5 ± 2.4 g/day and Phe of 4.6 ± 2.5 g/day. PCS plasma levels (20.4 ± 15.5 mg/L) were elevated and positively associated with both, Tyr (r = 0.58, p = 0.002) and Phe intake (r = 0.53, p = 0.005), even after adjustments for eGFR and age. Conclusion: This study suggests that the diet is an important modulator of the uremic toxins plasma levels produced by the gut microbiota, in non-dialysis CKD patients.
RESUMO Introdução: Pacientes com doença renal crônica (DRC) apresentam desequilíbrio na composição da microbiota intestinal, gerando toxinas urêmicas, como o p-cresil sulfato (PCS), pela fermentação bacteriana dos aminoácidos tirosina (Tyr) e fenilalanina (Phe) da dieta. Assim, a dieta pode ser determinante nos níveis de toxinas urêmicas produzidos pela microbiota intestinal. O objetivo deste estudo foi avaliar a possível relação entre a ingestão de Tyr e Phe e os níveis plasmáticos de PCS em pacientes com DRC não dialisados. Métodos: Foram avaliados 27 pacientes com DRC em tratamento conservador (estágios 3 e 4), sem intervenção nutricional prévia. A ingestão alimentar foi avaliada pelo recordatório alimentar de 24h (R-24h) de 3 dias, e a ingestão proteica também foi verificada através do Protein Nitrogen Appearance (PNA). Os níveis plasmáticos de PCS foram determinados por cromatografia líquida de fase reversa. Resultados: Os pacientes avaliados (TFG, 34,8 ± 12,4 mL/min, 54,2 ± 14,3 anos, IMC 29,3 ± 6,1 kg/m2) apresentaram ingestão média de proteína de 1,1 ± 0,5 g/kg/dia, Tyr de 4,5 ± 2,4 g/dia e Phe de 4,6 ± 2,5 g/dia. Os níveis plasmáticos de PCS (20,4 ± 15,5 mg/L) foram elevados e positivamente associados à ingestão de Tyr (r = 0,58, p = 0,002) e Phe (r = 0,53, p = 0,005), mesmo após ajustes pela TFG e idade. Conclusão: Este estudo sugere que a dieta é um importante modulador dos níveis plasmáticos de toxinas urêmicas produzidas pela microbiota intestinal em pacientes com DRC não dialisados.
Assuntos
Humanos , Fenilalanina , Tirosina , Dieta , Insuficiência Renal Crônica , Indicã , Sulfatos , Ésteres do Ácido Sulfúrico , Cresóis , Ingestão de AlimentosRESUMO
ABSTRACT Introduction: Gut microbiota imbalance is linked to high uremic toxins production such as indole-3-acetic acid (IAA) in chronic kidney disease patients. This toxin can activate the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor involved with inflammation. Strategies to restore gut microbiota balance can be associated with reduced production of IAA and its deleterious effects. This study aimed to evaluate prebiotic resistant starch (RS) supplementation effects on IAA plasma levels and AhR mRNA expression in CKD patients on hemodialysis (HD). Methods: This randomized, double-blind and placebo-controlled clinical trial evaluated forty-two stable HD patients allocated in RS (n=22) or placebo (n=20) groups. Patients received, alternately, cookies and sachets containing 16 g/day of RS (Hi-Maize 260®) or manioc flour for four weeks. Fasting pre-dialysis blood samples were collected and IAA plasma levels measured by high performance liquid chromatography. Peripheral blood mononuclear cells were isolated and processed for AhR and nuclear factor kappa B (NF-κB) mRNA expression analyzes by quantitative real-time PCR. Anthropometric and biochemical parameters, as well as food intake were also evaluated. Results: Thirty-one patients completed the study, 15 in the RS group and 16 in the placebo group. Although there was no significant alteration in IAA plasma levels, neither in AhR mRNA expression and NF-κB mRNA expression after RS supplementation, a positive correlation (r=0.48; p=0.03) was observed between IAA plasma levels and AhR expression at baseline. Conclusion: Even though prebiotic RS supplementation did not influence IAA levels or AhR expression, their positive association reinforces a possible interaction between them.
RESUMO Introdução: O desequilíbrio da microbiota intestinal associa-se à alta produção de toxinas urêmicas tais como ácido indol-3-acético (AIA), em renais crônicos. Essa toxina ativa o receptor aril hidrocarboneto (AhR) - fator de transcrição ativado por ligante, na inflamação. Restaurar o equilíbrio da microbiota intestinal associa-se à produção reduzida de AIA e efeitos deletérios. Avaliamos os efeitos da suplementação de amido resistente prebiótico (AR) sobre AIA sérico e expressão de AhR mRNA em renais crônicos em HD. Métodos: Estudo clínico randomizado, duplo-cego, controlado por placebo, com 42 pacientes em HD, nos grupos AR (n = 22) ou placebo (n = 20). Os pacientes receberam, alternadamente, biscoitos e sachês com 16 g/dia de AR ou polvilho - 4 semanas. Coletamos amostras de sangue em jejum pré-diálise e medimos níveis séricos de AIA por cromatografia líquida de alta eficiência. Isolamos e processamos as células mononucleares do sangue periférico para avaliar expressão AhR mRNA e NF-κB por PCR quantitativo em tempo real. Avaliamos parâmetros antropométricos, bioquímicos e ingestão alimentar. Resultados: 31 pacientes, 15 AR e 16 no placebo. Apesar de não apresentarem alteração significativa nos níveis de AIA, nas expressões de AhR ou NF-κB mRNA pós- suplementação com AR, foi verificada uma correlação positiva (r = 0,48; p = 0,03) entre AIA sérico e expressão de AhR na linha basal. Conclusão: Embora a suplementação com o prebiótico de AR não tenha influenciado os níveis de AIA ou a expressão de AhR, sua associação positiva reforça possível interação entre eles.