小剂量茶碱对大鼠气道上皮细胞NF-κB活性的影响

目的 :观察小剂量茶碱是否对体外培养的哮喘模型大鼠气道上皮细胞中NF κB的活性存在影响。以进一步阐明茶碱类药物抗炎作用和免疫调节作用的机制。方法 :雄性Wistar大鼠随机分为 2组 ,对照组 (5只 ) ,哮喘模型组 (15只 ) ;模型组以卵清蛋白致敏和激发 ,建立哮喘模型。体外培养哮喘模型组和对照组大鼠的气道上皮细胞。分别观察加入脂多糖 (LPS)刺激前后NF κB活性的变化 ;在哮喘模型组大鼠的气道上皮细胞中加入不同浓度的氨茶碱与LPS(1μg·ml- 1 )共同培养 4h后 ,以及加入同一浓度氨茶碱和LPS(1μg·ml- 1 )共同培养不同时间后NF κB活性的变化。NF κB活性采用免疫组织化学方法检测 ,以细胞核染色作为阳性标准 ,以细胞核阳性的百分比表示NF κB的活性。结果 :经LPS刺激前 ,对照组和哮喘组大鼠气道上皮细胞胞浆NF κB均有一定程度的表达。细胞核表达的细胞 ,哮喘组为 (6 .4 %± 1.2 ) % ,而对照组为 (4 .7%± 1.7) % ,二者相比较 ,其差异具有统计学意义 (P <0 .0 5 )。经 1μg·ml- 1 脂多糖刺激 2h后 ,两组细胞胞核NF κB表达均明显上升 ,哮喘组为 (82 .5± 5 .2 ) % ,对照组为(79.3± 3.4 ) % ,两者相比 ,无统计学差异 (P >0 .0 5 )。在哮喘组大鼠上皮细胞中加入 1μg·ml- 1 脂多糖及各种浓度氨茶碱

Effects of Low-dose Theophylline on Activity of NF-κB in Airway Epithelial Cells of Asthmatic Rats

Objective: To observe whether low dose theophylline has effects on activity of NF κB in airway epithelial cells (AEC) of asthmatic rats. Methods: 20 male Wistar rats were randomly assigned to control group (n=5) and asthma group (n=15). Asthma group were sensitized and challenged by OVA to establish an animal model of asthma. Airway epithelial cells of two groups were cultured in vitro,and activity of NF κB was observed pre and post administration of LPS. Airway epithelial cells of asthma group were added in 0 μg·ml -1 ,5 μg·ml -1 ,10 μg·ml -1 ,15 μg·ml -1 amionphylline respectively,cultured with 1 μg·ml -1 LPS for 4 hours or added in 10 μg·ml -1 amionphylline,cultured with 1 μg·ml -1 LPS for 2,4 and 24 hours respectively. The activity of NF κB was observed respectively. Immunohistochemical stain of NF κB P65 protein was used to evaluate the activity of NF κB. Results: Before treatment with LPS,the level of nuclear NF κB protein expression showed a significant difference between asthma and normal group (6.4%±1.2% versus 4.7 %±1.7%; P<0.05),but there was no significant difference between these two groups after the treatment with LPS (82.5%±5.2% versus 79.3%±3.4%,P>0.05). The level of nuclear NF κB protein expression rate was 84.3%±3.2%,82.8%±2.5%,78.4%±6.5% and 75.6%±4.5% after the AEC of asthma group were cultured with 0 μg·ml -1 ,5 μg·ml -1 ,10 μg·ml -1 ,and 15 μg·ml -1 amionphylline and 1 μg·ml -1 LPS,significant differences(P<0.05) were seen among these groups except for 0 μg·ml -1 and 5 μg·ml -1 group. The level of NF κB protein expression rate was 79.7 %±4.8%,77.5%±5.2% and 78.8%±4.6%,and after cultured for 2 h,4 h and 24 h with 10 μg·ml -1 aminophylline and 1 μg·ml -1 LPS,no significant difference was observed among them. Conclusion: Baseline NF κB activity is significantly higher in airway epthelial cells of athmatic rats. Theophylline could inhibit activity of NF κB in airway epithelial cells of athmatic rats in vitro. This inhibition depends on concentration of theophylline, and shows no significant relation with time.