## 3 Results
### Chemical and Biological Reactivities: CM Railroad Sites  
The collection sites were located at different distances (0.03 to 1.3 mi from the railyards (Table 1) identified as CM1 to CM4 in increasing distance. The PM2.5 prooxidant and VOC electrophile content decreased as a function of distance from the railroad emission source (Figure 2) with the exception of CM2 which was adjacent to two railyards. Prooxidant content was mostly associated with PM2.5 samples while electrophile content was mostly associated with the VOC samples. VOC prooxidant and PM2.5 electrophile contents remained close to background levels for all samples (Figure 6). The biological reactivities paralleled the profiles of the regional samples. Further, PM2.5 TNF-\(\alpha\) and VOC HO-1 expression showed a 'distance' pattern, with CM2 being the most potent and CM4 (most distant) being least potent; PM2.5 HO-1 expression remained close to background levels for all samples (Figure 7). Figure 5: Relative potencies of VOC and PM 2.5 air samples from CM, LB (**a**) and SB (**b**) to increase HO-1 expression in RAW 264.7 macrophages. Cell extracts were measured following exposure to the indicated concentrations of either VOC or its corresponding PM for 16 h. The lines represent the best line fit following linear regression analysis of the results (n = 3 for each data point) with potencies indicated by the slopes (values of the regression analysis are detailed in supplemental data—Table S5). Figure 6: Chemical properties of PM and VOC air samples collected near to the CM railroad area (CM1—closest, CM4 most distant). For each PM and VOC local sample, the nmol DTT consumed (prooxidant content (**a**)) and NEM equivalents (electrophile content (**b**)) were measured. Error bars indicate 95% CI for DTT (n = 3) results and SEM (n = 3) for the GAPDH results; analysis methods as described in Figure 4, (values of the CIs are detailed in supplemental data—Tables S2 and S3). SB VOC Pretreatment Effects on SB PM2.5 2.5 Expression of TNF-\(\alpha\) and HO-1 in RAW 264.7 Macrophages  
The anti-inflammatory properties of the VOC suggested that VOC pretreatment may affect the attenuation of the pro-inflammatory effects of a subsequent PM2.5 challenge. We selected the most potent samples--SB VOC for the pretreatment and SB PM2.5 for the challenge. A sequential exposure experiment was performed in which cells were first exposed to the SB VOC for 24 h. The media was then replaced with fresh media containing control filter extracts or suspensions of the SB PM2.5 and the exposure continued for 16 h. Macrophages and media were analyzed for HO-1 and TNF-\(\alpha\) content, respectively (Figure 8). The reason for the two-stage exposure was the limitation in the volume of sample solutions that could be added to the plate cells while maintaining the incubation conditions (see Methods for details). The results show a 54% decrease in the TNF-\(\alpha\) response to the PM2.5 challenge following pretreatment to SB VOCs at 1 m\({}^{3}\)/mL, n = 4; \(p^{\star\star\star}\) < 0.005, together with a significant 200% increase in HO-1 expression (Blank XAD/SB vs. SB VOC/SB 2.5; (n = 4; p < 0.001). Activation of the antioxidant Response Element (ARE) in RAW 264.7 Macrophages by Air Samples from CM, LB and SB  
In prior studies, we measured an HO-1 response in VOC samples collected at Riverside CA (a sample site nearby SB). Those samples also showed an electrophilic effect on the ARE [23]. Figure 8: SB VOC pretreatment effects on SB PM 2.5 expression of TNF-\(\alpha\) (**a**) and HO-1 (**b**) in RAW 264.7 macrophages. Cells were preincubated with VOC (1 m\({}^{3}\)/mL) for 24 h followed by incubation with PM 2.5 (1 m\({}^{3}\)/mL) for 16 h; TNF-\(\alpha\)—XAD Blk/PM 2.5 vs. VOC/PM 2.5 vs. VOC/PM 2.5, n = 4; \(p^{\star\star}\) < 0.005; HO-1—XAD Blk/PM 2.5 vs. VOC/PM 2.5, n = 4; \(p<0.001\). Figure 7: TNF-\(\alpha\) expression (**a**) and HO-1 expression (**b,c**) in RAW 264.7 macrophages using air samples collected nearby CM railroads (CM1—closest, CM4 most distant).