H oxidase appears to be responsible for ROS generation A 120 Wild-type gp91 phox KO in MPMVEC exposed to UFPs and/or CSE. 3.3. Co-exposure to UFPs and CSE resulted in increased 100 80 60 40 20 phosphorylation of MAPKs To investigate whether UFPs and/or CSE-induced ROS genera- tion could activate endothelial cells, the effects of UFPs and/or CSE on p38 and ERK1/2 mitogen-activated protein kinase (MAPK) activation in MPMVEC FK-506 were studied by Western blot. Our results showed that in MPMVEC from wild-type mice, the phosphorylation 0 0 10 20 50 100 200 of p38 and ERK1/2 MAPKs increased signiantly after 20 and 50 l g/ml of UFPs and 2.5% of CSE exposure for one hour ( Fig. 3 A UFPs (/ml) and B). This effect was enhanced when cells were co-exposed to both UFPs and CSE ( Fig. 3 A and B).

However, no such effects were B 120 Wild-type gp91 phox KO observed when MPMVEC from gp91 phox mice were exposed to UFPs and/or CSE ( Fig. 3 C and D). These results suggest that ROS generation in MPMVEC exposed to UFPs and/or CSE is involved in 100 80 60 the activation of MAPKs. 3.4. Co-exposure to UFPs and CSE led to increased Egr-1 expression 40 20 Exposure of MPMVEC from wild-type mice to 20 and 50 l g/ml of UFPs AZD2171 and 2.5% of CSE caused increased Egr-1 expression at both 0 0 1 2.5 5 CSE (%) 7.5 10 15 20 350 300 Wild-type gp91 phox KO   C 120 100 Wild-type phox 250 200 80 60 40 150 100 50 20 0 UFPs (/ml) 0 0 0 50 50 50 0 UFPs (/ml) CSE (%) 0 0 20 0 50 0 0 2.5 20 2.5 50 2.5 CSE (%) 0 2.5 5 0 2.5 5 Fig. 2.

Effects of UFPs, CSE and UFPs with CSE on ROS generation in MPMVEC from wild-type or gp91 phox KO mice. 1 10 4 cells were seeded into each well of 96-well Fig. 1.  purchase VX-950 Cytotoxicity of UFPs (A), CSE (B) and UFPs with CSE (C) on MPMVEC from wild-type or gp91 phox KO mice. 3 10 3 cells were seeded into each well of 96-well plates. After overnight culture, cells were treated with UFPs, CSE or UFPs with CSE, respectively. Cytotoxicity was determined with MTS assay kit (Promega) after 24 h treatment. Cells without UFPs or CSE treatment were used as controls. Data are shown as mean SD of three experiments with six replicates in each experiment.  Signiant difference as compared with the control, p < 0.05. plates. After overnight culture, cells were pretreated with 5 l M H 2 -DCFDA for 2 h prior to exposure to UFPs, CSE or UFPs with CSE for 4 h. Data are shown as mean SD of three experiments with six replicates in each experiment.

DCF rescence in cells with 5 l M H 2 -DCFDA pre-treatment but without UFPs and CSE  order VX-950 treatment was used as control. Signiant difference as compared with the control, p < 0.05;  Signiant difference as compared with the UFPs alone or CSE alone treated group, p < 0.05. Cell Viability (% of control) Cell Viability (% of control) (% of control) Cell Viability (% of control) DCF Fluorescence gp91 KO Page 4  Y. Mo et al. / Toxicology in Vitro 26 (2012) 29503 299 A UFPs (/ml) CSE (%) 0 0 0 2.5 20 0 50 0 20 2.5 50 2.5 C UFPs (/ml) CSE (%) 0 0 0 2.5 50 0 50 2.5 Phospho-p38 Total-p38 Phospho-ERK1/2 Total-ERK1/2 Phospho-p38 Total-p38 Phospho-ERK1/2 Total-ERK1/2 B 600 500 400 300 P-p38   D 200 150 100 P-p38 200 50 100 0 0 600 P-ERK-1   200 P-ERK-1 500 400 P-ERK-2   150 P-ERK-2 300 100 200 50 100 0 0 UFPs (/ml) CSE (%) 0 0 0 2.5 20 0 50 0 20 2.5 50 2.5 UFPs (/ml) CSE (%) 0 0 0 2.5 50 0 50 2.5 Fig. 3.

Increased phosphorylation of p38 and ERK1/2 in MPMVEC from wild-type mice exposed to UFPs, CSE and UFPs with CSE (A and B). In MPMVEC from gp91 phox in a 27% increase in ruxolitinib AUC, which is perhaps not clinically relevant in determining the starting dose chronic of ruxolitinib because the magnitude of the AUC change is within the intersubject variability (% coefficient of variation [CV]) observed in this or other clinical studies conducted with ruxolitinib. Mini- mal changes in ruxolitinib C max or t 1/2 were observed with erythromycin coadministration. Rifampin is a dual inducer of CY