柴油废气颗粒的吸入对哮喘大鼠速发相反应的影响

Effects of diesel exhaust particles inhalation on immediate reaction in asthma rats

目的探讨柴油废气颗粒的吸入对哮喘大鼠速发相反应的影响。方法选择Wistar雄性大鼠60只，随机分为6组，每组各10只。A组用生理盐水攻击，B组用卵清蛋白攻击，C、D、E、F组用卵清蛋白攻击以后继续吸入柴油废气颗粒（DEP）各1周、2周、3周和4周。所有抗原攻击结束1周以后，除A组用生理盐水激发30min以外其他组均用卵清蛋白激发30min，之后测定并比较气道阻力，观察支气管肺泡灌洗液中炎症细胞的变化和肺组织病理学的改变，肺组织中白细胞介素5和1干扰素的浓度，血清IgE变化等。结果（1）A、B、C、D、E、F组在卵清蛋白激发后30min时的气道阻力[cmH2O／（ml·s）]分别为（3．56±0．21）、（7．06±0．63）、（6．46±0．38）、（7．47±0．33）、（8．87±0．61）、（11．00±0．69）。A组未发生气道高反应，B、C、D、E、F组的气道阻力均高于A组，E、F组高于B组，差异均有统计学意义。而各组在各时间点内的气道阻力比较同样有统计学意义（F=160．646，148．901，162．204，156．186，P均〈0．01）。DEP吸入的时间和气道阻力之间呈正相关（r=0．948，P〈0．01）。（2）血清IgE浓度在B、C、D、E、F组之间差异无统计学意义（P〉0．05），但均高于A组（F=2．639，P〈0．01）。（3）支气管肺泡灌洗液中的炎症细胞有中性粒细胞、嗜酸性粒细胞和淋巴细胞等，以前者为主。中性粒细胞百分比（％）依次为（4．3±2．0）、（9．7±5．2）、（10．3±5．6）、（13．0±5．2）、（42．6±18．3）、（55．3±6．9），E、F组高于A组和B组，组间差异有统计学意义（F=114．226，P〈0．01）。（4）肺组织病理切片中可以看到A组上皮细胞完整，气道周围未见炎症细胞的浸润，以纤毛柱状上皮细胞为主，仅有少量的杯状细胞，基底膜未见纤维化。随着DEP吸人，逐渐出现上皮细胞的坏死、中断、脱落，杯状细胞增生，气道周围的炎症细胞浸润等改变。（5）肺组织中自细胞介素浓度（pg／mg）在B组（12．8±2．8）和C组（17．1±5．2）、E组（18．6±4．2）间差异有统计学意义（F=4．236，P〈0．01），各组大鼠肺组织中1干扰素的浓度之间比较差异无统计学意义（F=1．185，P〉0．05）。结论DEP的吸入加重了哮喘大鼠的速发相反应。

Objective The role of air pollution on asthma can not be ignored, diesel exhaust particles （DEP） in the air is one of the most important pollutants. This study aimed to investigate the effect and mechanism of DEP inhaled on immediate reaction in the asthma rats. Method Sixty male Wistar rats of ＂Clean＂ grade, 6-7 week-old, with an average weight of （140 ± 20 ） g were used in this study. The rats were randomly divided into 6 groups, 10 in each. Group A was treated with normal saline attack as a negative control, Group B with ovalbumin attack as a positive control. After ovalbumin attack, groups C, D, E, F continued to inhale DEP for 1 week, 2 weeks, 3 weeks and 4 weeks, respectively. The concentration of DEP was 200 ug/ml, the animals were subjected to inhalation of ultrasound nebulized DEP for 30 min per day. One week after all the attacks were concluded, Group A was stimulated with normal saline for 30 min, other groups were stimulated with ovalbumin. Then the airway resistance was determined with multi-channel signal acquisition and processing system and compared. The changes in neutrophils, eosinophils, and other inflammatory cells of BALF and the pathological changes in lung tissue, including epithelial cells loss, the inflammatory cells infiltration around the airway, basement membrane fibrosis, goblet cell hyperplasia etc. were observed. The concentration of IL-5 and γ-interferon in the lung tissues, and the changes of serum IgE etc. were determined. Result Airway resistance values of group A, B, C, D, E, F after ovalbumin excitation for 30minwere （3.56±0.21）, （7.06±0.63）, （6.46±0.38）, （7.47±0.33）, （8.87± 0. 61 ）, （11.00 ± 0.69） cm H2O/（ml · s）. No airway hyperresponsiveness occurred in group A, while Groups B, C, D, E, F had higher airway resistance than group A, group E and F had higher airway resistance than that of group B, the differences were statistically significant. And the airway resistance was different in each group among 0 min, 10 min, 20 min and 30 min （ F = 160. 646, 148. 901, 162. 204, 156. 186, P 〈0. 01 for both）. The time of DEP inhalation and the airway resistance was positively correlated （r =0. 948, P 〈0. 01 ）; IgE concentrations of the serum between groups B, C, D, E, F was not significantly different （P 〉 0. 05）, but higher than that of group A （F = 2. 639, P 〈 0. 01 ）. The infiltrated inflammatory cells included eosinophils and lymphocytes, etc. The percentages of neutrophil （%） were （4. 3 ± 2.0 ）, （ 9.7 ± 5.2）, （10.3±5.6）, （13.0±5.2）, （42.6±18.3）, （55.3±6.9）. The groups E and Fhad higher percentage than Group A and Group B（F = 114. 226, P 〈0. 01 ）. The percentages of eosinophils （ % ） were 0, （11.9 ±3.8） , （15.8 ±6. 3） , （13.0 ±4.9） , （21.1 ±5.6） , （27.1 ±4. 8）. The difference between Groups B, C, D, E, F and Group A was statistically significant. There was significant difference between groups C, D, E, F and group B （ F = 46. 462, P 〈 0.05） ; Lung tissue biopsy in group A showed that the epithelial cells were intact, no inflammatory cells infiltrations were found around the airways, instead, mainly ciliated columnar epithelial cells and only a small number of goblet cells were seen without basement membrane fibrosis. With the inhalation of DEP, the epithelial cells showed gradual necrosis, disruption and loss, goblet ceils showed hyperplasia, and infiltrations with inflammatory cells were seen around the airway. In the lung tissue, concentrations of IL-5 in group B,C, and E were （12.8 ±2.8）, （17.1 ±5.2）, （18.6 ±4. 2） pg/mg, the difference between groups C, E and group B was statistically significant （F = 4. 236, P 〈 0. 01 ）, the difference in γ-interferon concentration among all groups was not statistically significance （ F = 1. 185, P 〉 0. 05）. Conclusion DEP inhalation increased the airway responsiveness of asthma rats in immediate reaction, promoted the lung epithelial cell loss, inflammatory cell infiltration, basement membrane fibrosis and goblet cell hyperplasia.