The Establishment And Application Of Vitro Research Platform In Study Of Bronchial Epithelial Cell Injury By Black Powder Smoke

Objective:1, a smoke-generating device and an apparatus used in smoke treating cell were designed, and the cell research platform of smoke inhalation injury were established basing on the two previous.2, cultured human bronchial epithelial cell injured by civilian black powder smoke were studied through the cell research platform.Methods:1, in the smoke generating device, a wireless electromagnetic heater was used to ignite black powder and produce smoke. The quality of black powder was checked by black powder burn rate. The quality of experimental smoke was indirectly checked through the cell damage.2, the apparatus used in the study of smoke treating cell, was set to a constant temperature 37℃and the humidity was greater than constantly.3, factors which influence smoke treating cell were:cell density, the exposure mode, volume of smoke, testing time, volume of cell culture medium, combustion velocity.4, one test smoke was produced by a dose of 0.5g, 1g,2g,4g black powder burning and test time was set up to 10 minute. The other test smoke was produced by a dose of 2g black powder burning and test time was set up to 5min, 1Omin,15min and 20min. Then status of cells was evaluated to decide their injury by test smoke. 5, intracellular ROS levels were detected by DHE, and were analyzed by fluorescent microscope and IPP software.6, DNA damage in cell nuclei was detected by SCGE, and was analyzed by fluorescent microscope camera and casp software.7, IL-8 secreted by cells was detected through ELISA. Results:1, coefficient variation of different batches of black powder was less than 5%. Coefficient variation of different batches of smoke was less than 10%.2, factors that influence smoking-treating cells:(1) cell density in each hole of 12-well plate was 4×104. (2) the filter were used in the exposure mode. (3) the total smoke volume was greater than 10L. (4) cell culture medium volume was 1ml in each hole of 12-well plate. (5) cell activity was detected after 24h. (6) combustion time was less than 5 seconds.3, cell injury were greater with dose of black powder smoke increased and test time Extended.4, ROS levels were higher with cell injury greater in single cell.5, nuclear DNA breakage was more severe with cell injury greater.6, the amount of IL-8 secreted by cell was increased with cell injury greater in single cell.Conclusion:The experimental devices produced test smoke stably and safely. The apparatus of smoke treating cell fulfilled that cell injury was positively correlated with smoke concentration and exposure time. Bronchial epithelial cell was injured by black powder smoke, and the mechanism was relevant with increased ROS levels, nuclear DNA breakage and the increased secretion of IL-8.