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Unveiling the links between cysteine and outcomes In peritoneal dialysis
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RESUMO: O aumento global da prevalência de doença renal crónica (DRC) tem conduzido a um número crescente de doentes a atingir o estádio 5 - doença renal terminal e, por isso, a necessitar de terapêutica de substituição renal. Entre as opções disponíveis, a diálise peritoneal (DP) oferece vantagens clínicas e de qualidade de vida, mas a sua eficácia a longo prazo é limitada pela disfunção da membrana peritoneal (PM), um processo intimamente ligado à homeostase redox local e sistémica.
Hipotetizámos que os aminotióis, incluindo cisteína (Cys), γ-glutamilcisteína (GlyCys), homocisteína (HCys), cisteinilglicina (CysGly), glutationa (GSH) e N-acetilcisteína (NAC) urinária, designados em conjunto por “tioloma”, podem fornecer novas pistas sobre a integridade da MP e o equilíbrio redox, servindo como potenciais biomarcadores do estado da membrana e dos resultados em DP.
Este estudo apresenta uma caracterização inovadora e multi-compartimental do tioloma numa coorte real de doentes em DP, em plasma, urina, efluente peritoneal noturno (PDE) e efluentes recolhidos às 2 h e 4 h do teste de equilíbrio peritoneal (PET), um teste clínico padronizado utilizado para avaliar as características de transporte da PM. O nosso objetivo foi identificar ligações entre o tioloma associado à cisteína e os desfechos em doentes em diálise peritoneal, avaliar o seu potencial como biomarcadores da função da membrana peritoneal e dos resultados clínicos da DP, e explorar se a sua integração em modelos clínicos existentes poderá melhorar a estratificação dos doentes e ajudar a identificar fármacos que contribuam para manter a eficácia da DP. Análises multivariadas (PCA, PLS-DA), univariadas e de regressão linear revelaram que as concentrações de aminotióis são influenciadas por fatores clínicos. A CysGly destacou-se como marcador sensível de stress oxidativo e turnover redox extracelular, enquanto a GSH refletiu preferencialmente alterações locais no equilíbrio redox peritoneal. Destaca-se que os doentes tratados com o fármaco X apresentaram níveis significativamente mais elevados de GSH no PDE noturno, efluentes PET e urina, independentemente da presença de diabetes, com níveis plasmáticos inalterados, sugerindo maior disponibilidade e turnover de GSH peritoneal. A cinética de transporte dos aminotióis durante o PET diferiu dos solutos clássicos, com a GSH a apresentar maior variabilidade interindividual e, em alguns doentes, com rácios dialisado/plasma (D/P) superiores a um entre as 2 e 4 horas, consistentes com produção intraperitoneal in situ. Estes trajetos D/P específicos de cada soluto reforçam a utilidade do perfil multi-soluto 11 no PET para complementar a classificação tradicional do tipo de transportador e caracterizar melhor a função da PM. Para explorar os mecanismos subjacentes, foi usado um modelo celular de mesotélio humano (MET-5A), exposto a fluidos brancos do PET e moduladores redox. A citotoxicidade dependeu do momento de aplicação das 4 h de estímulo, com exposição precoce a permitir recuperação parcial, enquanto exposição tardia resultou em perda sustentada de viabilidade. A quantificação extracelular de tióis revelou diminuição de Cys e aumento de GSH no meio condicionado, consistente com uma resposta adaptativa de tamponamento redox ao stress oxidativo induzido pelo PET.
Em suma, este estudo demonstra que o perfil tiolómico abrangente fornece novas informações sobre a biologia redox da MP e identifica os aminotióis como potenciais biomarcadores da saúde da membrana e dos resultados em DP. Estes resultados servem como pilares para estudos translacionais futuros, visando personalizar a gestão da DP através da estratificação redox dos doentes e do desenvolvimento de biomarcadores de alerta precoce.
ABSTRACT: The global rise in chronic kidney disease (CKD) prevalence has led to an increasing number of patients progressing to stage 5 - end-stage kidney disease (ESKD), and thus requiring renal replacement therapy. Among available modalities, peritoneal dialysis (PD) offers significant clinical and quality-of-life advantages, yet its long-term efficacy is limited by peritoneal membrane (PM) dysfunction, a process closely linked to local and systemic redox homeostasis. We hypothesized that aminothiols, such as cysteine (Cys), γ-glutamylcysteine (GlyCys), homocysteine (HCys), cysteinylglycine (CysGly), glutathione (GSH), and urinary N-acetylcysteine (NAC), collectively termed the “thiolome”, may provide novel insights into PM integrity and redox balance, serving as candidate biomarkers for PM status and PD outcomes.This study presents an innovative, multi-compartmental characterization of the thiolome in a real-world PD cohort across plasma, urine, overnight peritoneal effluent (PDE), and 2-h and 4-h effluents of the peritoneal equilibration test (PET), a standardized clinical test used to assess PM transport characteristics and to adapt the PD prescription. Our goal was to unveil links between the Cys-related thiolome and outcomes in PD patients, to evaluate their potential as biomarkers of PM function and PD outcomes, and to explore whether their integration into existing clinical frameworks may enhance patient stratification and help identify drugs that contribute to maintaining PD effectiveness. Multivariate (PCA, PLS-DA), univariate, and regression analyses revealed that aminothiol concentrations are shaped by patient-related factors. CysGly emerged as a sensitive marker of oxidative stress and extracellular redox turnover, while GSH appeared to preferentially reflect local changes in peritoneal redox balance. Notably, patients treated with drug X exhibited significantly higher GSH levels in overnight PDE, PET effluents, and urine, independent of diabetes status, while plasma levels remained unchanged, consistent with enhanced peritoneal GSH availability and turnover. Aminothiol transport kinetics during PET differed from classical solutes, with GSH displaying the greatest interindividual variability and, in some patients, dialysate-to-plasma (D/P) ratios higher than one between 2- and 4-h, consistent with in situ intraperitoneal generation. These solute-specific D/P trajectories support the utility of multi-solute PET profiling to complement standard transporter classification and to better characterize PM function. To uncover underlying mechanisms, a human mesothelial cell model (MET-5A) was used, and a preliminary study was conducted, exposing cells to PET blank fluids and redox modulators. Cytotoxicity was influenced by the timing of the 4-hour stimulus, with early exposure allowing partial recovery, while late exposure resulted in sustained loss of viability. Extracellular thiol quantification revealed decreased Cys and increased GSH in conditioned media, consistent with an adaptive redox-buffering response to PET-induced oxidative stress. In summary, this study demonstrates that comprehensive thiolomic profiling provides novel insights into PM redox biology and identifies aminothiols as candidate biomarkers for membrane health and PD outcomes. These findings lay the groundwork for future translational studies aiming to personalize PD management through redox-based patient stratification and to develop early-risk biomarkers.
ABSTRACT: The global rise in chronic kidney disease (CKD) prevalence has led to an increasing number of patients progressing to stage 5 - end-stage kidney disease (ESKD), and thus requiring renal replacement therapy. Among available modalities, peritoneal dialysis (PD) offers significant clinical and quality-of-life advantages, yet its long-term efficacy is limited by peritoneal membrane (PM) dysfunction, a process closely linked to local and systemic redox homeostasis. We hypothesized that aminothiols, such as cysteine (Cys), γ-glutamylcysteine (GlyCys), homocysteine (HCys), cysteinylglycine (CysGly), glutathione (GSH), and urinary N-acetylcysteine (NAC), collectively termed the “thiolome”, may provide novel insights into PM integrity and redox balance, serving as candidate biomarkers for PM status and PD outcomes.This study presents an innovative, multi-compartmental characterization of the thiolome in a real-world PD cohort across plasma, urine, overnight peritoneal effluent (PDE), and 2-h and 4-h effluents of the peritoneal equilibration test (PET), a standardized clinical test used to assess PM transport characteristics and to adapt the PD prescription. Our goal was to unveil links between the Cys-related thiolome and outcomes in PD patients, to evaluate their potential as biomarkers of PM function and PD outcomes, and to explore whether their integration into existing clinical frameworks may enhance patient stratification and help identify drugs that contribute to maintaining PD effectiveness. Multivariate (PCA, PLS-DA), univariate, and regression analyses revealed that aminothiol concentrations are shaped by patient-related factors. CysGly emerged as a sensitive marker of oxidative stress and extracellular redox turnover, while GSH appeared to preferentially reflect local changes in peritoneal redox balance. Notably, patients treated with drug X exhibited significantly higher GSH levels in overnight PDE, PET effluents, and urine, independent of diabetes status, while plasma levels remained unchanged, consistent with enhanced peritoneal GSH availability and turnover. Aminothiol transport kinetics during PET differed from classical solutes, with GSH displaying the greatest interindividual variability and, in some patients, dialysate-to-plasma (D/P) ratios higher than one between 2- and 4-h, consistent with in situ intraperitoneal generation. These solute-specific D/P trajectories support the utility of multi-solute PET profiling to complement standard transporter classification and to better characterize PM function. To uncover underlying mechanisms, a human mesothelial cell model (MET-5A) was used, and a preliminary study was conducted, exposing cells to PET blank fluids and redox modulators. Cytotoxicity was influenced by the timing of the 4-hour stimulus, with early exposure allowing partial recovery, while late exposure resulted in sustained loss of viability. Extracellular thiol quantification revealed decreased Cys and increased GSH in conditioned media, consistent with an adaptive redox-buffering response to PET-induced oxidative stress. In summary, this study demonstrates that comprehensive thiolomic profiling provides novel insights into PM redox biology and identifies aminothiols as candidate biomarkers for membrane health and PD outcomes. These findings lay the groundwork for future translational studies aiming to personalize PD management through redox-based patient stratification and to develop early-risk biomarkers.
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Chronic kidney disease Peritoneal dialysis