Effect of serum derived from non-smokers and smokers and of lipoprotein-depleted serum (LPDS) derived from smokers on intracellular GSH concentration, on intracellular reactive oxygen species (ROS) and on nitric oxide (NO) formation in human umbilical vein endothelial cells (HUVECs).

<p>Confluent HUVECs were incubated without and with 10, 30 and 50% serum derived from non-smokers and smokers and with the corresponding LPDS derived from smokers for 12 hours. Figure shows intracellular GSH concentration (A), intracellular ROS (B) and cumulative basal and bradykinin-stimulated NO production, evaluated by measuring levels of nitrite in the supernatants (C). Data are presented as mean±SD of measurements performed in triplicate in four different occasions; *P<0.01 versus control (no addition of serum derived from the subjects) non-smokers' serum and smokers' LPDS.</p

Effect of increasing concentrations of oxPAPC on Nrf2, GCLC and HO-1 expression in human umbilical vein endothelial cells.

<p>mRNA (A–C) was analysed by quantitative Real-Time PCR. Normalised gene expression levels were given as the ratio between the mean value for the target gene and that for beta-actin in each sample. Results are presented as the mean±SD of measurements performed in triplicate.*P<0.01 versus oxPAPC 0 and 150 µg/mL. Figure (D) shows a representative Western blot analysis of three independent experiments for nuclear Nrf2, GCLC and HO-1.</p

Endothelial function and correlation between flow-mediated vasodilation (FMD) and concentrations of GSH in peripheral blood mononuclear cells (PBMC) of non-smokers and smokers.

<p>FMD and glyceryl trinitrate (GTN)-induced vasodilation in non-smokers and smokers (A); correlation between FMD and intracellular concentrations of GSH in PBMC of non-smokers and smokers (B). Data are presented as mean±SD; FMD and GTN are expressed as maximal percentage change in brachial artery dilation. *P<0.001 versus non-smokers.</p

Effect of increasing concentrations of oxPAPC on intracellular GSH concentration, on intracellular reactive oxygen species (ROS), and on nitric oxide (NO) production in human umbilical vein endothelial cells (HUVECs).

<p>Confluent HUVECs were incubated without and with increasing concentrations (25–150 µg/mL) of oxPAPC for 6 hours. Figure shows intracellular GSH concentration (A), intracellular ROS (B) and cumulative basal and bradykinin-stimulated NO production, evaluated by measuring levels of nitrite in the supernatants (C). Data are presented as mean±SD of measurements performed in triplicate in four different occasions. *P<0.01 versus oxPAPC 0 e 75 µg/mL. # P<0.01 versus basal NO.</p

Effect of serum derived from non-smokers and smokers and of lipoprotein-depleted serum (LPDS) derived from smokers on Nrf2, GCLC and HO-1 expression in human umbilical vein endothelial cells (HUVECs).

<p>Confluent HUVECs were incubated without and with increasing amounts (10, 30 and 50%) of serum derived from smokers (serum S 10, serum S 30, serum S 50), with 50% serum derived from non-smokers (serum N–S 50) and with 50% LPDS derived from smokers (LPDS S 50) for 12 hours. mRNA for Nrf2, GCLC and HO-1 (A–C) was analysed by quantitative Real-Time PCR. Normalised gene expression levels were given as the ratio between the mean value for the target gene and that for beta-actin in each sample. Results are reported as the mean±SD of measurements performed in triplicate. *P<0.01 versus non-smokers and LPDS. (D) shows a representative Western blot analysis of three independent experiments for nuclear Nrf2, for GCLC and HO-1.</p

Effect of small interfering (si) and scrambler (sc) RNA against Nrf2 on oxPAPC-dependent expression of GCLC and HO-1 in human umbilical vein endothelial cells.

<p>mRNA (a) was analyzed by quantitative Real-Time PCR. Normalized gene expression levels were given as the ratio between the mean value for the target gene and that for the beta-actin in each sample. Figure shows a representative Western blot analysis for GCLC and HO-1 and the average quantification obtained by densitometric analysis of all the samples (b). Data on Western blot analysis are expressed as the density ratio of target to control (beta-actin) in arbitrary units ×10. Results are the mean±SD of measurements performed in triplicate in four different occasions. *P<0.01 versus control; †P<0.01 versus scRNA; #P<0.01 versus oxPAPC+scRNA.</p