Melatonin regulates the lipopolysaccharide-induced iNOS expression in endothelial cells.

<p>iNOS expression in cultures obtained from naïve (black bar), vehicle injected (white bars), lipopolysaccharide injected (LPS, 0.5 mg/kg; grey bars) or LPS + melatonin injected (MEL 3.0 mg/kg; grey diagonal striped bars) animals was determined by confocal immunofluorescence. The injections were performed 2 or 6 hours before sacrifice (6 hours after lights off –24h00). Data are expressed as mean ± SEM, n = 4–8 per group; *significantly different (P<0.05) versus 2 hours vehicle group (white bars).</p

Melatonin regulates the lipopolysaccharide-induced iNOS expression in endothelial cells.

<p>iNOS expression in cultures obtained from naïve (black bar), vehicle injected (white bars), lipopolysaccharide injected (LPS, 0.5 mg/kg; grey bars) or LPS + melatonin injected (MEL 3.0 mg/kg; grey diagonal striped bars) animals was determined by confocal immunofluorescence. The injections were performed 2 or 6 hours before sacrifice (6 hours after lights off –24h00). Data are expressed as mean ± SEM, n = 4–8 per group; *significantly different (P<0.05) versus 2 hours vehicle group (white bars).</p

Plasma melatonin concentration is inversely correlated with the expression of adhesion molecules and iNOS in endothelial cells.

<p>The plasma concentration of melatonin (pg/mL) in each rat was plotted against the expression of PECAM-1 (A), ICAM-1 (B) and iNOS (C) shown as the percentage of the fluorescence in the naïve group seeded in the same chamber slide. The values of the slopes and the Pearson's coefficient are shown in the text.</p

Standard curve of leukocyte adhesion assay.

<p>Representative image of experimental myeloperoxidase assay (A). The number of polymorphonuclear cells (PMN) was determined by comparison with standard curve (B). Confluent endothelial cell culture adhered (D) or not (C) with leukocytes. Arrow indicates leukocytes adhered to endothelial cells. Scale bar  = 50 µm.</p

Melatonin regulates the lipopolysaccharide-induced iNOS, PECAM-1 and ICAM-1 expression in endothelial cells.

<p>Representative images of immunofluorescence assays of iNOS (A–C), PECAM-1 (D–F) and ICAM-1 (G–I). Columns show cells obtained from vehicle (A, D and G), lipopolysaccharide (LPS) (B, E and H) and LPS + melatonin (MEL) (C, F and I) injected animals. Scale bar  = 50 µm.</p

Melatonin regulates leukocyte adhesion to cultured endothelial cells.

<p>Leukocyte adhesion on endothelial cells cultures obtained from naïve (black bar), vehicle injected (white bars), lipopolysaccharide injected (LPS, 0.5 mg/kg; grey bars) or LPS + melatonin injected (MEL 3.0 mg/kg; grey diagonal striped bars) animals was determined by leukocyte-endothelial adhesion assay. The injections were performed 2 or 6 hours before sacrifice (6 hours after lights off –24 h 00). Data are expressed as mean ± SEM, n = 4–8 per group of at least two different experiments; * significantly different (P<0.05) versus 2 hours vehicle group (white bars).</p

Experimental protocols.

<p>Wistar rats were divided into four groups: naïve, vehicle, lipopolysaccharide (LPS, 0.5 mg/kg) and LPS + melatonin (MEL 3 mg/kg). The intravenous (caudal vein) injections of vehicle, LPS or LPS + melatonin were performed two (A) or six (B) hours before sacrifice (6 hours after lights off –24h00). Grey portion represents the dark phase of the day.</p

Melatonin regulates the lipopolysaccharide-induced PECAM-1 and ICAM-1 expression in endothelial cells.

<p>Expression of PECAM-1 (A) and ICAM-1 (B) in cultures obtained from naïve (black bar), vehicle injected (white bars), lipopolysaccharide injected (LPS, 0.5 mg/kg; grey bars) or LPS + melatonin injected (MEL 3.0 mg/kg; grey diagonal striped bars) animals were determined by confocal immunofluorescence. The injections were performed 2 or 6 hours before sacrifice (6 hours after lights off –24h00). Data are expressed as mean ± SEM, n = 4–8 per group; * significantly different (P<0.05) versus 2 hours vehicle group (white bars).</p