The uremic solute indoxyl sulfate induces oxidative stress in endothelial cells.
BACKGROUND
Endothelial dysfunction and oxidative stress are matters of concern in patients with chronic renal failure (CRF). Uremic solutes retained in these patients could be involved in these processes. Notably, the protein-bound uremic solute indoxyl sulfate induces endothelial dysfunction in vitro, and has shown pro-oxidant effects.


OBJECTIVE
To demonstrate that indoxyl sulfate is a potential mediator of oxidative stress in endothelial cells in vitro.


METHODS
Indoxyl sulfate-induced oxidative stress in human umbilical vein endothelial cells (HUVEC) was studied by measuring reactive oxygen specie (ROS) production by cytofluorimetry, by analyzing the involvement of the pro-oxidative enzymes NAD(P)H oxidase, xanthine oxidase, and NO synthase, and by measuring the levels of the non-enzymatic antioxidant glutathione.


RESULTS
We showed that indoxyl sulfate induced a significant production of ROS in HUVEC, with or without human serum albumin. We then investigated the role of pro-oxidative enzymes and measured the levels of the antioxidant glutathione. The NAD(P)H oxidase inhibitors, DPI, and apocynin, inhibited ROS production, whereas inhibitors of xanthine oxidase, NO synthase, and mitochondrial ROS had no effect. Interestingly, indoxyl sulfate strongly decreased the levels of glutathione, one of the most active antioxidant systems of the cell. In addition, the ROS production mediated by indoxyl sulfate was inhibited by the antioxidants vitamin C, vitamin E, and NAC.


CONCLUSIONS
The uremic solute indoxyl sulfate enhances ROS production, increases NAD(P)H oxidase activity, and decreases glutathione levels in endothelial cells. Thus, indoxyl sulfate induces oxidative stress by modifying the balance between pro- and antioxidant mechanisms in endothelial cells.
Dou
L
L
UFR Pharmacie, Université Aix-Marseille, Marseille, France. laetitia.dou@pharmacie.univ-mrs.fr
Jourde-Chiche
N
N
Faure
V
V
Cerini
C
C
Berland
Y
Y
Dignat-George
F
F
Brunet
P
P
D000975:Antioxidants / Q000494:pharmacology; D001483:Base Sequence; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism*; D006801:Humans; D007200:Indican / Q000378:metabolism / Q000494:pharmacology*; D019255:NADPH Oxidases / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000235:genetics; D052250:Nitric Oxide Synthase Type III / Q000235:genetics; D018384:Oxidative Stress / Q000187:drug effects*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D014511:Uremia / Q000378:metabolism*; D014969:Xanthine Oxidase / Q000378:metabolism
D000975:Antioxidants; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; C496319:NOS2 protein, human; C496314:NOS3 protein, human; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; D014969:Xanthine Oxidase; D019255:NADPH Oxidases; D007200:Indican