不同粒径的大气细颗粒物对A549细胞的毒性研究

研究了不同粒径的大气细颗粒物对人肺癌上皮细胞A549的生物学效应,探讨了大气中细颗粒物粒径与细胞毒性的关系。采集杭州市大气中不同粒径的细颗粒物,制备成不同粒径的颗粒物总悬浮液,将A549细胞暴露于该制备液24 h后,测定细胞活力(MTT法)和培养液上清液中的LDH含量;并选择最佳浓度为后续暴露剂量,测定ROS生成量,评价不同粒径大气细颗粒物对细胞损伤效应。此外,还采用RTPCR测定凋亡基因P50、BAX、BCL-2表达情况。不同粒径的细颗粒物均显著抑制A549细胞活力,LDH显著增多,并存在剂量-效应关系;最佳染毒质量浓度为10μg/m L;ROS的生成量和各个凋亡基因的mRNA表达量均高于对照组。大气中不同粒径的细颗粒物均能对A549细胞产生毒性,并且随粒径增加,颗粒物对细胞的毒性呈降低趋势。

On the toxicity of the fine particles with different sizes to A549 cells

The paper is entitled to engage in an investigation and exploration of the relationship between the atmospheric fine particulates and their cytotoxicity by taking different sizes of fine particulates collected in Hangzhou with DLPI sample of 13 ranks.For the above said purpose,we have collected sample particulates known as PM0.030-0.108,PM0.108-0.170,PM0.170-0.260,PM0.260-0.400,PM0.400-0.650,PM0.650-1.000,PM1.000-1.600 and PM1.600-2.500 and organized them into a series of different content concentrations of the corresponding suspension particles.To examine and determine the cytotoxicity of the said fine particulate matters of different sizes,we have diluted them through DMEM with 1％ FBS of 1 000-fold,so as to ensure their concentrations within a range of 1μg/mL,5 μg/mL,10 μg/mL,50 μg/mL and 100 μg/mL,respectively,and then let them exposed to for 24 h.And,the following step is to determine the cell viability with the MTT assay and the LDH release of the culture supernatant with ELISA.Furthermore,we have chosen the optimum concentrations of cytotoxicity from MTF assay for further in-depth studies.And,afterwards,the effects of cell damage can be evaluated and determined under the optimum concentration at ROS level,whereas the expression of apoptotic genes (P53,BAX and BCL-2) can be determined by using RT-PCR assay.As a result,we have found that the viability of A549 cells has been subjected to be significantly inhibited by the different sizes of the fine particles while LDH tends to increase significantly (p ＜ 0.01) with the increase of the particle sizes.On the other hand,when the mass concentration of the particulates remains at about 10 μg/mL,the cells tend to show great toxicity.Moreover,when the mass concentration of the particulates is getting bigger than 10 μg/mL,the cell viability may turn to be almost the same with that in the case of 10 μg/mL.However,when we have chosen the particulates of 10 μg/mL to conduct the rest experiments,it has been found that the particulates of 10 μg/mL should be attributed to and selected as the optimum concentration.The results of our study have proven that the amount of ROS generation and apoptosis gene at mRNA expression level can be much greater than those of the control group.In addition,their generation and mRNA expression can be expected to be affected by the particle size.Or,more exactly speaking,the smaller the particle size,the greater the degree of the oxidative damage,and,in turn,the stronger the expression viability of the cell apoptosis related genes.Therefore,different sizes of fine particles tend to reveal their different cytotoxicity to A549 cells,with such a kind of toxicity dropping down with the increase of the particle size.