The Involvement Of Heme Oxygenase In PM_2.5-toxicity In Human Umbilical Vein Endothelial Cells

Objective To investigate the involvement of heme oxygenase (HO-1) in the toxicity induced by PM2.5in human umbilical vein endothelial cells (HUVECs) by observing the effects of ZnPP, an inhibitor of HO-1, and Tiron a scavenger of ROS on the toxicity.Methods and materials Revive the frozen cells and choose cells in logarithmic phase for following experiment. The experiment groups are as follows:(1) control group.(2)PM2.5treatment groups:the cells were cultured with PM2.5(200、400or800μg/ml) for24h.(3) PM2.5+Trion group:the cells were pre-treated by Trion (10μM) for1h before the co-culture with PM2.5(400μg/ml) for24h.(4) PM2.5+ZnPP group:the cells were pre-treated by ZnPP (10μM) for1h before the co-culture with PM2.5(400μg/ml) for24h. MTT assay was used to detect cell viability. Reverse transcription polymerase chain reaction (RT-PCR) and indirect immunofluorescence assay were used for the analysis of mRNA and protein. Fluorescence labeling probe method was used to measure intracellular reactive oxygen species (ROS) level and flow cytometry was used for cell apoptosis.Results1.Compared with the control group, the cell viability of PM2.5-treated group decreased with its concentration increasing. when the concentration of PM2.5is200,400,800μg/ml, the cell survival rate was (67.02±5.12)%、(54.52±5.74)%、(44.81±7.80)%. When the concentration of PM2.5is400μg/ml, in PM2.5, PM2.5+Tiron and PM2.5+ZnPP groups, the cell viability were (54.52±5.74)%、(78.46±7.74)%、(40.40±4.58)%respectively (P<0.05)2. When the concentration of PM2.5is400μg/ml, the optical density ratio of HO-1mRNA in control, PM2.5, PM2.5+Tiron, PM2.5+ZnPP groups respectively were0.83±0.04,1.24±0.04,0.60±0.21,0.68±0.08(P<0.05)3. When the concentration of PM2.5is400μg/ml, the mean fluorescence density of HO-1protein in control, PM2.5, PM2.5+Tiron, PM2.5+ZnPP groups respectively were26.16±4.31,83.10±4.80,60.90±1.14,45.41±1.44(P<0.01) 4. When the concentration of PM2.5is400μg/ml, the intracellular ROS level in control, PM2.5, PM2.5+Tiron, PM2.5+ZnPP group respectively were4.00±0.95,12.88±2.26,9.29±1.06,29.33±0.51(P<0.05)5. When the concentration of PM2.5is400μg/ml, the cell apoptosis rate in control, PM2.5, PM2.5+Tiron, PM2.5+ZnPP groups respectively were (3.12±3.49)%,(13.30±5.22)%,(6.05±1.25)%,(22.13±3.09)%(P<0.05)Conclusion PM2.5can produce oxidative stress injury in HUVECs, heme oxygenase may involve the anti-oxidative effects, the activation of HO-1may be related with the expression of ROS;HO-1may play an important role in response of HUVECs to PM2.5.