Resistant starch supplementation effects on plasma indole 3-acetic acid and aryl hydrocarbon receptor mRNA expression in hemodialysis patients: Randomized, double blind and controlled clinical trial / Efeitos da suplementação de amido resistente no ácido indol-3-acético plasmático e na expressão do mRNA do receptor aril-hidrocarboneto em pacientes em hemodiálise: ensaio clínico randomizado, duplo-cego e controlado

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.

Resistant starch supplementation effects on plasma indole 3-acetic acid and aryl hydrocarbon receptor mRNA expression in hemodialysis patients: Randomized, double blind and controlled clinical trial.

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.

Aryl Hydrocarbon Receptor and Uremic Toxins from the Gut Microbiota in Chronic Kidney Disease Patients: Is There a Relationship between Them?

Recent studies have suggested that uremic toxins such as indoxyl sulfate (IS) and indole-3-acetic acid (IAA) from the metabolism of the gut microbiota may be involved in the inflammatory signaling pathway in chronic kidney disease (CKD) patients through the activation of the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor. The objective of this study was to investigate the possible relationship between uremic toxins (IS and IAA) and AhR protein expression in CKD patients. A cross-sectional observational study involving 17 hemodialysis (HD) [11 men, 55.5 ± 11.7 years of age, 54.0 (25.5-136.0) months of HD, body mass index (BMI) of 25.8 ± 3.8 kg/m2] and 15 non-dialysis-dependent (NDD) CKD (8 men, 54.1 ± 18.2 years of age, glomerular filtration rate of 34.8 ± 21.0 mL/min/1.73 m2, BMI of 27.4 ± 5.0 kg/m2) patients was conducted. IS and IAA levels were measured by reversed-phase high-performance liquid chromatography, and the protein expression levels of AhR and nuclear factor κ B (NF-κB) were evaluated by a Western blot assay. There was no difference in the expression of either AhR or NF-κB in the patients, and as expected, uremic toxin levels were higher in HD patients than in NDD patients. In the overall analysis, AhR protein expression was positively associated with IAA plasma levels ( r = 0.4; p = 0.03) and NF-κB protein expression ( r = 0.62; p = 0.001). Although the role of AhR in inflammation and CVD in CKD patients is far from being completely understood, the association between IAA and AhR observed in this study suggests a possible role for uremic toxins in the cell signaling pathway involved in inflammation in CKD patients.

Is there a relationship between tryptophan dietary intake and plasma levels of indoxyl sulfate in chronic kidney disease patients on hemodialysis?

INTRODUCTION: Gut microbiota is involved in generation of uremic toxins in chronic kidney disease (CKD) patients on hemodialysis (HD), like indoxyl sulfate (IS) that is originated from tryptophan amino acid fermentation. OBJECTIVE: To evaluate the tryptophan intake by chronic renal failure patients on HD and its possible relationship with IS plasma levels. METHODS: Participated of the study 46 patients with CKD on HD regular program (56.5% men; 52.7 ± 10.3 years; 63 (32.2-118.2) months on HD; BMI 25.6 ± 4.9 kg/m2). The tryptophan intake was evaluated by a 24-hours dietary recall (R-24h) performed on 3 different days. Routine biochemical tests and anthropometric measurements were evaluated. IS plasma levels were determined by High Performance Liquid Chromatography (HPLC) with fluorescent detection and the interleukin-6 (IL-6) plasma levels by immunoenzymatic method (ELISA, Enzyme Linked Immunosorbent Assay). RESULTS: The average of tryptophan intake was according to recommendation, but IS plasma levels (35.0 ± 11.9 mg/L) were elevated, however according to the EUTox values for uremic individuals. There was no correlation between the tryptophan intake and IS plasma levels. However, there was positive correlation between protein intake and tryptophan and variables used to evaluate lean body mass, and moreover, IS levels were positively associated with IL-6 (r = 0.6: p = 0.01). CONCLUSION: The present study suggests that tryptophan dietary intake may not be a determinant factor to IS levels. However, it suggests that gut microbiota may play an important role in systemic inflammation in patients with CKD.

Is there a relationship between tryptophan dietary intake and plasma levels of indoxyl sulfate in chronic kidney disease patients on hemodialysis? / Há relação entre ingestão alimentar de triptofano e níveis plasmáticos de indoxil sulfato em pacientes renais crônicos em hemodiálise?

Abstract Introduction: Gut microbiota is involved in generation of uremic toxins in chronic kidney disease (CKD) patients on hemodialysis (HD), like indoxyl sulfate (IS) that is originated from tryptophan amino acid fermentation. Objective: To evaluate the tryptophan intake by chronic renal failure patients on HD and its possible relationship with IS plasma levels. Methods: Participated of the study 46 patients with CKD on HD regular program (56.5% men; 52.7 ± 10.3 years; 63 (32.2-118.2) months on HD; BMI 25.6 ± 4.9 kg/m2). The tryptophan intake was evaluated by a 24-hours dietary recall (R-24h) performed on 3 different days. Routine biochemical tests and anthropometric measurements were evaluated. IS plasma levels were determined by High Performance Liquid Chromatography (HPLC) with fluorescent detection and the interleukin-6 (IL-6) plasma levels by immunoenzymatic method (ELISA, Enzyme Linked Immunosorbent Assay). Results: The average of tryptophan intake was according to recommendation, but IS plasma levels (35.0 ± 11.9 mg/L) were elevated, however according to the EUTox values for uremic individuals. There was no correlation between the tryptophan intake and IS plasma levels. However, there was positive correlation between protein intake and tryptophan and variables used to evaluate lean body mass, and moreover, IS levels were positively associated with IL-6 (r = 0.6: p = 0.01). Conclusion: The present study suggests that tryptophan dietary intake may not be a determinant factor to IS levels. However, it suggests that gut microbiota may play an important role in systemic inflammation in patients with CKD.

Resumo Introdução: A microbiota intestinal está envolvida na geração de toxinas urêmicas presentes nos pacientes com doença renal crônica (DRC) em hemodiálise (HD) como indoxil sulfato (IS), formado a partir da fermentação do aminoácido triptofano. Objetivo: Avaliar a ingestão de triptofano alimentar pelos pacientes renais crônicos em HD e sua possível relação com os níveis plasmáticos de IS. Métodos: Participaram do estudo 46 pacientes com DRC em programa regular de HD (56,5% homens; 52,7 ± 10,3 anos; 63 (32,2-118,2) meses em HD; IMC 25,6 ± 4,9kg/m2. A ingestão de triptofano foi avaliada por meio do recordatório alimentar de 24 (R-24h) realizado em três diferentes dias. Exames bioquímicos de rotina, bem como a avaliação antropométrica foram avaliados. Os níveis plasmáticos de IS foram determinados por cromatografia líquida de alto desempenho (HPLC) com detecção fluorescente e as concentrações plasmáticas de interleucina-6 (IL-6) pelo método imunoenzimático (ELISA, Enzyme Linked Immunosorbent Assay). Resultados: A ingestão média de triptofano estava dentro do recomendado, já os níveis plasmáticos de IS (35,0 ± 11,9mg/L) estavam elevados, porém de acordo com os valores da EUTox para indivíduos urêmicos. Não houve correlação entre a ingestão de triptofano e os níveis plasmáticos de IS. Contudo, houve correlação positiva entre ingestão de proteína e triptofano e variáveis que avaliam massa magra e, além disso, os níveis IS foram positivamente associados com os de IL-6 (r = 0,6: p = 0,01). Conclusão: O presente estudo sugere que a ingestão alimentar de triptofano pode não ser um fator determinante dos níveis de IS. No entanto, sugere que o intestino pode ter importante papel na inflamação sistêmica presente nos pacientes com DRC.