| Background:In recent years, particulate matter has become the most important pollutant in most cities of China, and it is the main factor that affects the urban air quality. Studies have shown that long-term exposure to airborne particulate matter is a risk factor for respiratory disease occurred in the population. The settlement of the particles in the air is related to the particle size, the smaller the particle size, the more slowly the sedimentation rate is, the easier it is to be absorbed into the body. Because of the small particle size, PM2.5 is easy to be inhaled in the body deposition in the alveolar,directly affect the ventilation of lung ventilation function, make the body to a state of hypoxia. And because of its large surface area, so that bacteria, viruses, organic matter and heavy metals and other toxic substances adsorbed on the surface of the particles into the depth of the lung, further increase the damage to the lungs. In haze weather,people pay more attention to the protection of the respiratory system, in fact, the environmental pollution factors on the skin damage can not be ignored. Skin diseases like dermatitis, eczema, acne, psoriasis, pigmented diseases and skin cancer or other skin aging problems are related to environmental pollution. Most of the previous studies focused on effects of PM2.5 on the respiratory system, but to study the skin injury little. Therefore, our study was to clear the effect and mechanism of different concentrations of PM2.5 on Ha Ca T cells.Objective:To study the effects of different concentrations of PM2.5 on the injury of Ha Ca T cells and the damage effect of NF-κB inhibitor PDTC on Ha Ca T cells induced by PM2.5.Methods:In this study, we collected the PM2.5 of particulate matter, and formulated it into different concentrations, and then stimulated Ha Ca T cells with different concentrations of PM2.5. Cell viability was measured by MTT method, and determine the concentration of PM2.5 and reaction time. Give a certain concentration of PDTC intervention, the cells were divided into four groups: normal group, PM2.5group,PDTC group and PDTC+PM2.5 group. Apoptosis was detected with DAPI staining and Western blot method. MDA content in Ha Ca T cells was measured by TBA assay.Use SOD, GSH-px and CAT assay kit to detect the cell antioxidant defense status.The levels of IL-6 and INF-α in cell supernatant were determined by ELISA. Protein expressions of Bax, Bcl-2, TLR-2, TLR-4 and NF-κB p65 in Ha Ca T cells was detected by Western blot.Results:1. Effect of PM2.5 on cell viability of Ha Ca T cells: Cell viability decreased with increasing concentrations of PM2.5 in certain range, the cell viability of 200、400、800μg/ml groups are significantly lower than the normal group(P<0.05), There was no significant difference in the cell viability of Ha Ca T cells cultured for 24 h and 48 h at the same concentration of PM2.5.2. Effect of PM2.5 on apoptosis of Ha Ca T cells: By DAPI staining analyzing,normal group of cells to produce weaker blue light, the nuclei are round, the edge is clear and dyeing uniformity. PM2.5 group of cells apoptosis, resulting in a strong blue light, the edge of the nucleus is irregular; In PDTC group, the cell staining results were nearly no different from the normal group. There were a small part of the cells to produce a strong blue light in PM2.5+PDTC group.3. Effects of PM2.5 on the content of MDA in Ha Ca T cells :Compared with normal group, MDA contents in PM2.5 group and PM2.5+PDTC group increased significantly(P<0.05). Compared with PM2.5 group, MDA contents in PDTC group and PM2.5+PDTC group decreased significantly(P<0.05). Compared with PDTC group, MDA content in PM2.5+PDTC group increased significantly(P<0.05).4. Effects of PM2.5 on the levels of ROS in Ha Ca T cells: Compared with normal group, the levels of ROS in PM2.5 group and PM2.5+PDTC group increased significantly(P<0.05). Compared with PM2.5 group, the levels of ROS in PDTC group and PM2.5+PDTC group decreased significantly(P<0.05).5. Effects of PM2.5 on the enzyme activities of SOD, GSH-Px and CAT in Ha Ca T cells: Compared with normal group, enzyme activities of SOD, GSH-Px and CAT in PM2.5 group and PM2.5+PDTC group decreased significantly(P<0.05).Compared with PM2.5 group, enzyme activities of SOD, GSH-Px and CAT in PDTC group and PM2.5+PDTC group increased significantly(P<0.05). Compared with PDTC group, enzyme activities of SOD, GSH-Px and CAT in PM2.5+PDTC groupdecreased significantly(P<0.05).6. Effect of PM2.5 on the levels of IL-6 and TNF-α in supernatant of Ha Ca T cells: IL-6 and TNF-α in cell supernatant of PM2.5 group is significantly higher than that in normal group(P<0.05). IL-6 and TNF-α in cell supernatant of PDTC group and PM2.5+PDTC group is significantly lower than that in PM2.5 group.7. Effect of PM2.5 on the expression of Bax and Bcl-2 in Ha Ca T cells:Compared with normal group, Bax/Bcl-2 rate in PM2.5 group and PDTC group increased significantly(P<0.05). There was no significant difference of Bax/Bcl-2rate between PM2.5+PDTC group and normal group. Compared with PM2.5 group,Bax/Bcl-2 rate in PM2.5+PDTC group decreased significantly(P<0.05).8. Effect of PM2.5 on the expression of TLR-2, TLR-4 and NF-κB p65 in Ha Ca T cells: Compared with normal group, TLR-2 expression levels in PM2.5 group and PM2.5+PDTC group increased significantly(P<0.05). Compared with PM2.5 group,TLR-2 expression levels in PDTC group and PM2.5+PDTC group decreased significantly(P<0.05). Compared with PDTC group TLR-2 expression level in PM2.5+PDTC group increased significantly(P<0.05).Compared with normal group, TLR-4 expression levels in PM2.5 group and PM2.5+PDTC group increased significantly(P<0.05). TLR-4 expression level in PDTC group decreased significantly(P<0.05). Compared with PM2.5 group, TLR-4expression levels in PDTC group and PM2.5+PDTC group decreased significantly(P<0.05). Compared with PDTC group TLR-4 expression level in PM2.5+PDTC group increased significantly(P<0.05).Compared with normal group, NF-κB p65 expression levels in PM2.5 group and PM2.5+PDTC group increased significantly(P<0.05). NF-κB p65 expression level in PDTC group decreased significantly(P<0.05). Compared with PM2.5 group, NF-κB p65 expression levels in PDTC group and PM2.5+PDTC group decreased significantly(P<0.05). Compared with PDTC group NF-κB p65 expression level in PM2.5+PDTC group increased significantly(P<0.05).Conclusion:1. PM2.5 makes Ha Ca T cell viability decreased and causes apoptosis.2. PM2.5 can cause oxidative damage to Ha Ca T cells, and PDTC can alleviate the damage.3. PM2.5 causes inflammatory cytokines IL-6 and TNF-α levels increased of Ha Ca T cells in supernatants, resulting in inflammatory injury, while PDTC can reducethe effects of this damage.4. PM2.5 can make NF- kappa B pathway activated through the signal transduction pathways of TLR-2 and TLR-4.5. NF-κB inhibitor PDTC can reduce the damage of PM2.5 on Ha Ca T cells in a certain extent. |
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