1. Volume 85, Issue 1, Pages 72-81 (January 2014)
TNF-mediated damage to glomerular endothelium is an important determinant of acute kidney injury in sepsis 
Chang Xu, Anthony Chang, Bradley K. Hack, Michael T. Eadon, Seth L. Alper, Patrick N. Cunningham 
Kidney International 
Volume 85, Issue 1, Pages (January 2014)
DOI: /ki
Copyright © 2014 International Society of Nephrology Terms and Conditions

2. Figure 1
Endotoxemia induces tumor necrosis factor receptor 1 (TNFR1)-dependent acute kidney injury (AKI) and increases urine concentration of albumin, in association with decreased fenestral density and increased fenestral diameter of glomerular capillary endothelium. Effect of lipopolysaccharide (LPS; 10mg/kg for 24h) on (a) plasma blood urea nitrogen concentration, (b) urine albumin-to-creatinine ratio, (c) weight loss, (d) fenestral density, and (e) fenestral diameter in wild-type (WT) and Tnfr1−/− mice. Control, WT treated with normal saline; WT LPS, WT treated with LPS; TNFR1−/− LPS, Tnfr1−/− mice treated with LPS. Values are mean±s.e.m. for 3–5 animals (a–c), 60–70 capillary loops (d), and 33–67 fenestrae (e). *P<0.05 vs. other groups; **P<0.01 vs. other groups; ##P<0.01 vs. WT mice injected with LPS.
Kidney International  , 72-81DOI: ( /ki )
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3. Figure 2
Transmission electron microscopic ultrastructure of glomerular endothelial cells in glomeruli of 8-week-old WT control mice (a, d) or WT mice injected with LPS (10mg/kg for 24h; b, e) or Tnfr1−/− mice injected with LPS (10mg/kg for 24h; c, f). (a–c) Low-magnification electron micrographs. Control, WT treated with normal saline; WT LPS, WT treated with LPS; TNFR1−/− LPS, Tnfr1−/− mice treated with LPS. Scale bar, 2μm. (d, e) High-magnification electron micrographs, with en face view of the fenestrae. Scale bar, 0.5μm. Arrows indicate endothelial fenestrae. Arrowheads in (b) indicate endothelial cell detachment. CL, capillary lumen; F, fenestrate; LPS, lipopolysaccharide; P, podocyte foot processes; RBC, red blood cell; WT, wild-type.
Kidney International  , 72-81DOI: ( /ki )
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4. Figure 3
Intravenous injection of tumor necrosis factor-α (TNF-α) recapitulates a form of acute kidney injury similar to that produced by lipopolysaccharide (LPS) injection, with blood urea nitrogen (BUN) elevation (a), pathological renal injury (b), and renal influx of neutrophils (c). Vacuolization and disruption of tubules is seen, with areas of leukocyte infiltration (arrow). Low-dose intravenous TNF-α did not elevate BUN. BUN elevation required TNF-α levels similar to those achieved after LPS injection (3–10ng/ml). i.v., intravenous.
Kidney International  , 72-81DOI: ( /ki )
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5. Figure 4
Transmission electron microscopic ultrastructure of glomerular endothelial cells in glomeruli of wild-type mice 24h after intravenous administration of saline (control, a, c) or 2.5μg of tumor necrosis factor-α (TNF-α; b, d). (a, b) Loss of endothelial fenestrae in the glomerular capillaries in TNF-treated mice compared with controls. (c, d) High-magnification electron micrographs, with en face view of the fenestrae, showing enlarged glomerular endothelial fenestrae in TNF-treated mice. Scale bar, 1μm. Arrows indicate endothelial fenestrae.
Kidney International  , 72-81DOI: ( /ki )
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6. Figure 5
Effect of lipopolysaccharide (LPS) on vascular endothelial growth factor (VEGF) expression in the kidney. (a) Time-dependent effect of LPS (10mg/kg, 6, 24, and 48h) on renal VEGF mRNA expression, as normalized to 18S mRNA expression. (b) Effect of LPS (10mg/kg for 24h) on VEGF protein expression, as normalized to the expression of β-actin protein. Top inset: representative VEGF immunoblot of kidney lysate. Values are means±s.e.m. for 4–6 animals. **P<0.01 vs. control.
Kidney International  , 72-81DOI: ( /ki )
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7. Figure 6
Effect of lipopolysaccharide (LPS) on vascular endothelial growth factor receptor 2 (VEGFR2) expression in the kidney. (a, b) Indirect immunofluorescence photomicrographs of frozen kidney cortex sections from wild-type (WT) control mice (a) and WT mice treated 24h with 10/ mg/kg LPS (b), incubated with anti-VEGFR2 antibody. LPS did not change VEGFR2 expression in glomeruli. (c) Time-dependent effect of LPS (10mg/kg, 6, 24, and 48h) on renal VEGFR2 mRNA expression, as normalized to 18S mRNA expression. Values are means±s.e.m. for 4–6 animals. Analysis of variance shows no significant difference between the LPS and control groups. Scale bar 20μm.
Kidney International  , 72-81DOI: ( /ki )
Copyright © 2014 International Society of Nephrology Terms and Conditions

8. Figure 7
Effect of lipopolysaccharide (LPS) on glomerular endothelial surface layer (ESL). (a–l) Glomerular ESL is shown by Alexa Fluor 594-WGA staining and co-staining for the endothelial junctional proteins vascular endothelial (VE)-cadherin and CD31 in wild-type (WT) mice treated for 24h with 10mg/kg LPS (d–f) and their matched controls (a–c), or in mice treated 24h with 2.5μg tumor necrosis factor (TNF) intravenously (j–l) and their matched controls (g–i). Glomerular ESL is also shown by staining proteoglycans (PGs) containing heparan sulfate (HS) glycosaminoglycan (GAG) chains in WT control mice (m) and in WT mice 24h after treatment with 10mg/kg LPS (n). (o) Fluorescence intensity of glomerular wheat germ agglutinin (WGA) staining. Data (normalized to staining intensity of control mice) are expressed as mean±s.e.m. **P<0.01 vs. WT control. Scale bar 20μm.
Kidney International  , 72-81DOI: ( /ki )
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9. Figure 8
Effect of lipopolysaccharide (LPS) on glomerular heparanase levels and expression. Indirect immunofluorescence photomicrographs of frozen kidney sections from wild-type (WT) control mice (a–c) and from WT mice treated for 24h with LPS (10mg/kg) (d–f), incubated with antibodies against heparanase-1 (red; a, d) and against nephrin (green; b, e). (g) Top inset: representative immunoblot of heparanase-1 protein expression in kidney. Bar graph quantifying kidney heparanase-1 expression from immunoblot densitometry. Data normalized to ‘control’ band are expressed as mean±s.e.m. **P<0.01 vs. WT control. Scale bar 20μm.
Kidney International  , 72-81DOI: ( /ki )
Copyright © 2014 International Society of Nephrology Terms and Conditions