From bench to the hemodialysis clinic: protein-bound uremic toxins modulate NF-κB/Nrf2 expression

Uremic toxins produced by gut microbiota (indoxyl sulfate-IS, p-cresyl sulfate-p-CS, and indole-3-acetic acid-IAA) accumulate in hemodialysis (HD) patients and exhibit potent inflammatory effects. However, the impact of these toxins on nuclear E2-related factor 2 (Nrf2) and nuclear factor-kappa B (NF-kappa B) expression in HD patients remains poorly defined. The aim of this study was to evaluate the association between uremic toxins and Nrf2/NF-kappa B expression in vitro (RAW 264.7 macrophage-like cells) and in peripheral blood mononuclear cells from HD patients. Uremic toxins, C-reactive protein (CRP), interleukin-6 (IL-6) and malondialdehyde (MDA) levels were measured in fifteen HD patients and nine healthy individuals. RAW 264.7 macrophage-like cells were incubated with IS, as a prototype of protein-bound uremic toxin. Nrf2 and NF-kappa B expressions were analyzed by RT-qPCR. HD patients presented high levels of inflammatory markers, MDA and uremic toxins. In addition, they presented high NF-kappa B and low Nrf2 expression. Uremic toxins were positively correlated with NF-kappa B expression (IS, rho = 0.58, p < 0.003; p-CS, rho = 0.71, p < 0.001; IAA, rho = 0.62, p < 0.001) and negatively with Nrf2 (IS, rho = - 0.48, p = 0.01; p-CS, rho = - 0.46, p < 0.02). Uremic toxins also exhibited positive correlations with CRP and MDA levels. Multivariate analysis revealed that p-CS is a determinant factor of NF-kappa B expression. In RAW 264.7 culture, NF-kappa B mRNA expression was stimulated by IS, while Nrf2 was downregulated. Thus, uremic toxins may stimulate NF-kappa B mRNA and decrease Nrf2 expression in HD patients and, consequently, trigger inflammation and oxidative stress.