Effect Of Traffic Related Fine Particulate On Apoptosis Of CEM-6T

PartⅠInducing effects of traffic related fine particulate on apoptosis of CEM-6TObjective: To discuss the inducing effects of apoptosis of CEM-6T caused by traffic related fineparticulate.Methods: The proliferation function of CEM-6T exposed to 24h and 48h traffic related fineparticulate was determined by MTT assay. Apoptosis of early and necrosis of CEM-6Tlabelled with FITC-AnnexinV/PI were detected by flow cytometry. The morphologicalchange of apoptotic cells was investigated by AO/EB fluorescent staining under fluorescentmicroscope. The level of expression of caspase-3 mRNA was detected by RT-PCR.Results: The SI of CEM-6T decreased markedly in both 160(excluding 48h), 320, 800μg/mLPM2.5 after 24h and 48h exposure compared to control group, and the differences werestatistically significant(P<0.05). Through AO/BE double fluorescence staining, we foundthat the level of late apoptosis and necrosis increased with the dose increased. Determiningby flow cytometry, after 24h exposure, the early apoptosis rate of CEM-6T in320μg/mLPM2.5 was higher than the control group and the difference of the late apoptosisand necrosis had no statistical significance. After 48h exposure, the early apoptosis rates in20, 80, 320μg/mL PM2.5 were higher than the control group, and the late apoptosis andnecrosis in 320μg/mL PM2.5 was higher than the control group, further more thesedifferences were statistically significant(P<0.05). The levels of expression of caspase-3mRNA were increased in both 20, 80, 320μg/mL PM2.5 after 24h exposure and 48h, and thedifferences were statistically significant(P<0.05).Conclusions: The proliferation inhibition and induction of apoptosis of CEM-6T could becaused by traffic related fine particles.PartⅡThe mechanism of the apoptosis of CEM-6T treated with traffic related fineparticulateObjective: To explore the mechanism of inducing effects of the apoptosis of CEM-6T treatedwith traffic related fine particles.Methods: The measurement of the mitochondrial transmembrane potential of CEM-6T wasdetected by Rh-123 single staining. The expressions of Caspase-8, Caspase-9, Cyt-c, Fas-L,TNF-αprotein were measured by Western Blot.Results: The mitochondrial transmembrane potential of CEM-6T decreased in 80(excluding48h) , 160, 320μg/mL PM2.5 groups after 24h and 48h exposure compared to control group(P<0.05). Gray-scale ratio of Cyt-c protein was increased in 20(excluding 48h), 80,320μg/mL PM2.5 compared to control group after 24h and 48h exposure. the differenceswere statistically significant(P<0.05). Gray-scale ratios of Caspase-9 protein in 20, 80,320μg/mL PM2.5 were higher than control group after 24h and 48h exposure, and thedifferences were statistically significant(P<0.05). Gray-scale ratios of Caspase-8 proteinwere increased in 20,80,320μg/mL PM2.5 compared to control group after 24h and 48hexposure, and the differences were statistically significant(P<0.05). Gray-scale ratios ofFas-L protein were increased in 20(excluding 48h), 80, 320μg/mL PM2.5 compared tocontrol group after 24h and 48h exposure, and the differences were statisticallysignificant(P<0.05). Gray-scale ratio of TNF-αprotein was increased in 20(excluding 48h),80, 320μg/mL PM2.5 compared to control group after 24h and 48h exposure, and thedifference was statistically significant(P<0.05).Conclusions: Traffic related fine particulate can induce apoptosis of CEM-6T both viamitochondrial pathway and membrane death receptor pathway.