Indoxyl Sulfate Induces Leukocyte-Endothelial Interactions through Up-regulation of E-selectin

Abstract

Cardiovascular disease is a major cause of death in chronic kidney disease (CKD) 2 (1). Atherosclerosis is highly prevalent in patients with severe renal failure and advances more rapidly in individuals with renal dysfunction compared with the general population (2). Reduced kidney function is associated with the risk of cardiovascular events, even when the dysfunction is mild (3).Leukocyte-endothelial interactions play an important role in the development of atherosclerosis (4). Cell adhesion molecules belonging to the immunoglobulin superfamily, such as ICAM-1 (intercellular cell adhesion molecule-1) and VCAM-1 (vascular cell adhesion molecule-1), together with members of the selectin family, including E-selectin, are upregulated to mediate monocyte/macrophage infiltration into atherosclerotic lesions (4, 5).Indoxyl sulfate is a uremic toxin synthesized in the liver from indole, a metabolite of tryptophan produced by the intestinal flora (6). In CKD patients, the serum levels of indoxyl sulfate are increased significantly compared with those in healthy individuals (7), and a number of studies have indicated that indoxyl sulfate accelerates glomerular sclerosis, whereas its accumulation promotes renal failure (8 -10). Other studies also showed that indoxyl sulfate induces endothelial dysfunction by releasing endothelial microparticles (11) and producing reactive oxygen species (ROS) (12). However, its effect on endothelial inflammatory processes such as leukocyte recruitment to vascular endothelium has not been reported.We report for the first time that indoxyl sulfate enhances monocyte adhesion to vascular endothelium through up-regulation of E-selectin and augmentation of oxidative stress in both in vitro and in vivo models. The underlying mechanisms seem to involve activation of JNK and NF-B. Our findings reveal a previously unrecognized molecular link between uremic toxins and cardiovascular diseases. EXPERIMENTAL PROCEDURESReagents-Indoxyl sulfate, N-acetylcysteine, probenecid, RPMI 1640 medium, and Dulbecco's PBS were obtained from Sigma. The JNK phosphorylation inhibitor SP600125, the p38 MAPK phosphorylation inhibitor SB203580, the ERK1/2 inhibitor U0126, and the IB phosphorylation inhibitor BAY11-7082 were purchased from Calbiochem. Recombinant human TNF-␣ was obtained from R&D Systems (Minneapolis, MN). A monoclonal antibody against E-selectin (clone 7A9) was obtained from American Type Culture Collection (Manassas, VA) (13). Antibodies against ICAM-1, VCAM-1, the NF-B p65 subunit, and the phospho-NF-B p65 subunit and a monoclonal blocking antibody against mouse E-selectin (clone UZ4) were obtained from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA). Anti-ERK, anti-phospho-ERK, anti-p38 MAPK, anti-phospho-p38 MAPK, anti-JNK, and anti-phos-* This work was supported in part by Ministry of Education, Science, and Technology Grant-in-aid for Scientific Research 10178102 and special coordination funds, a grant-in-aid from the Ministry of Culture of Japan, a grant from the Ministry of Health, Labor, and We...

Protein-Bound Uremic Toxins Stimulate Crosstalk between Leukocytes and Vessel Wall

“…These in vivo data strengthen previous in vitro findings showing that IS influences expression of endothelial adhesion molecules, such as intercellular adhesion molecule 1 32 and E-selectin. 33 In addition, Ito et al 33 showed IS-induced leukocyte adhesion to the femoral artery after administering IS in the drinking water of nephrectomized mice for 10 days, which was significantly reduced by anti-E-selectin antibody treatment. The firm arrest of leukocytes, which was seen in our study after exposure to IS, has already been observed for another tryptophan metabolite, kynurenic acid, in an in vitro model.…”

Aryl Hydrocarbon Receptor Mediates Indoxyl Sulfate-Induced Cellular Senescence in Human Umbilical Vein Endothelial Cells

“…Yeager et al reported that AhR plays a role in Nrf2 activation in the liver in mice administered TCDD 60) , and Tsuji et al demonstrated that the antiinflammatory effects of ketoconazole, a widely used imidazole antifungal agent, via Nrf2 activation require AhR signaling in normal human epidermal keratinowhich accumulates as renal dysfunction progresses, enhances oxidative stress in the setting of CKD. Furthermore, the mechanism by which IS enhances oxidative stress has been reported to involve the transportation of IS into the cell via OAT1 and 3, where it subsequently generates superoxide by activating NADPH oxidase in endothelial, renal tubular and glomerular mesangial cells [34][35][36][37][38] . Although IS is known to induce endothelial senescence due to oxidative stress [49][50][51] , no previous reports have investigated whether Sirt1 is involved in the pathogenesis of ISinduced endothelial senescence.…”

“…13,33) . It has been reported that the mechanism by which IS induces oxidative stress involves the transport of IS into the cell via organic anion transporters (OAT1, OAT3), where it then activates nicotinamide adenine dinucleotide phosphate (NADPH) oxidase in endothelial, renal tubular and glomerular mesangial cells [34][35][36][37][38] . Although it is unknown how IS activates NADPH oxidase, we recently found that aryl hydrocarbon receptor (AhR), which is expressed in human umbilical vein endothelial cells (HUVECs), is involved in the upregulation of NADPH oxidase-4 induced by IS 39) .…”

Neurocritical Care for Patients with Kidney Dysfunction

“…Both toxins induced production of reactive oxygen species and decreased cell viability; phosphate additionally caused eNOS uncoupling [101]. Indoxyl sulfate can induce reactive oxygen species production in various cell types other than vascular endothelial cells, including VSMC, renal tubular cells, monocytes, and macrophages [102][103][104][105]. These findings remain to be validated using in vivo studies of BBB integrity.…”