S, MO, USA). The ANOVA gene list was obtained by industrial software Partek genomic Suite. This Minimum Information about a Microarray Experiment-compliant dataset has been deposited in the NCBI Gene Expression Omnibus, GEO Series accession number GSE51834 (http://ncbi.nlm.nih.gov/geo/query/ acc.cgi?token=ezanqyoclzunxcf acc=GSE51834).AhR localized to podocyte nuclei in mouse kidneysIndoxyl sulfate is as an endogenous ligand of AhR, acting as a ligand-activated transcription factor that regulates detoxification, carcinogenesis, and inflammation [16,17]. We localized AhR in typical mouse kidneys by utilizing immunofluorescence (Figure 3a?c). AhR expression was restricted to the glomerulus (Figure 3a) in a subset of podocytes good for WT1 and synaptopodin, exactly where it was largely restricted to the nucleus (Figure 3b and c). Subsequent, we assessed the mRNA expression of Cyp1a1, which can be induced by the activation of AhR following ligand binding. Cyp1a1 mRNA expression in the kidney was considerably elevated by two h right after exposure and peaked at 4 h, having a return to regular by 24 h, and was elevated in glomeruli as well as entire kidneys at 2 h following indoxyl sulfate exposure (Figure 3d and e).Statistical analysesThe results had been expressed as imply 6 SD. Information for two groups had been analyzed utilizing the Student’s t-test. For several comparisons, evaluation was by ANOVA by using a Bonferroni test (to compare all pairs) or Dunnett test (to examine all samples vs. the control samples). Significance was inferred for P,0.05.PLOS 1 | plosone.orgPodocyte Injury by Indoxyl SulfateFigure 1. Indoxyl sulfate induced tubulointerstitial and vascular injuries in mouse kidneys. Serum indoxyl sulfate levels had been measured following single dose exposure (a and b). C57BL/6 mice (800 mg/kg, i.p. provided once). n = 1 (time course, panel a). n 7 (240 min, panel b), mean 6 SD. Histopathology of FVB/N mouse kidneys following chronic exposure to automobile or indoxyl sulfate, administered at 600 mg/kg/d i.p. for 8 w (c ). In contrast to kidneys from vehicle-exposed mice (c), global renal atrophy was observed in 1 of 2 kidneys from an indoxyl sulfate-exposed mouse (d, arrow). Bars = 500 mm. Vehicle-treated mice manifested histologically unremarkable tubules stained with periodic acid Schiff (e).98386-83-5 In stock Inside the macroscopically atrophied kidneys in indoxyl sulfate-exposed mice, prominent tubulointerstitial injury with numerous, prominent protein casts within tubules and comprehensive tubular atrophy (f) and foci of interstitial fibrosis were observed with Masson trichrome staining (g).529476-80-0 Formula Bars = 20 mm.PMID:28322188 doi:ten.1371/journal.pone.0108448.gChronic indoxyl sulfate exposure caused podocyte injury in miceWe characterized podocyte injury in chronically indoxyl sulfateexposed mice following 8 w of exposure to indoxyl sulfate (Figure 4a ). Podocytes manifested prominent but incomplete foot process effacement and podocyte cytoplasmic vacuoles (Figure 4b ) as well as a focal granular/wrinkled pattern of podocin and synaptopodin (Figure 4e). Furthermore, glomerular vimentinpositive signals and AhR-positive cells had been enhanced inside the indoxyl sulfate-exposed glomeruli in comparison with the control (Figure 4e). Quantitative assessment demonstrated a reduce in the synaptopodin-positive region inside the glomeruli of mice showing macroscopic renal atrophy (Figure 4f). Further, a important boost inside the vimentin- or AhR-positive location was observed in indoxyl sulfate-exposed mice (Figure 4f). Immunoblotting demonstrated that podocin a.