咖啡因经Nrf2通路调节支气管肺发育不良新生大鼠肺氧化应激损伤

Caffeine regulates lung oxidative stress injury through Nrf2 pathway in neonatal rats with bronchopulmonary dysplasia

目的探讨咖啡因对高氧诱导的新生SD大鼠支气管肺发育不良(BPD)氧化应激损伤的保护作用及相关机制。方法幼鼠分为空气对照组(N组)、咖啡因空气对照组(NC组)、模型组(H组)、咖啡因干预组(HC组)。采用高氧诱导法建立BPD模型。各组于第3、7、14、21天分别取6只大鼠收集血液和肺组织标本。各组按4个日龄分为4个亚组,4组的21天亚组测量体质量。各组右肺上叶测定湿干重比值(W/D);右肺下叶石蜡包埋后切片进行HE染色观察形态学改变并行辐射状肺泡计数(RAC);检测血、肺组织丙二醛(MDA)、超氧化物歧化酶(SOD)含量评估幼鼠机体氧化和抗氧化稳态失衡情况;RT-qPCR检测肺组织转录因子NF-E2相关因子2(Nrf2)mRNA相对表达量。结果①H组第3天后渐出现活动度下降,第14天后体质量增长呈上升趋势;②H组W/D值第14天达高峰,HC组变化趋势与H组相近;③H组肺组织结构不规则,RAC值第7天达高峰后显著下降,HC组和H组比较差异有统计学意义(P<0.01);④H组MDA值第7天时呈上升趋势,第14天后渐下降,SOD活性第7天时明显降低,H组和HC组于第14、21天比较差异有统计学意义(P<0.05);⑤H组Nrf2 mRNA在第7天时表达显著增强,第14天时趋于平稳,H组和HC组于第3、7、14天比较差异有统计学意义(P<0.05);⑥Nrf2 mRNA相对表达量与MDA呈正相关,与SOD呈负相关。结论咖啡因可经Nrf2通路,调节氧化应激反应,减轻高氧诱导的新生大鼠BPD肺氧化应激损伤。

Objective To investigate the protective effect of caffeine on oxidative stress injury induced by hyperoxia in neonatal SD rats with bronchopulmonary dysplasia(BPD)and its related mechanism.Methods Neonatal SD rats were randomly divided into air control group(N group),caffeine air control group(NC group),model group(H group)and caffeine intervention group(HC group).The high oxygen induction method was used to establish the BPD model.Blood and lung tissue of six samples were collected from each group on day 3,7,14 and 21.Each group was divided into four subgroups according to four days of age.The 21-day panel of four groups measured their body weights.The upper lobe of the right lung was used to measure wet-dry weight ratio(W/D)of lung tissue in each group;the lower lobe of the right lung was sliced after paraffin embedding and stained with hematoxylin-eosin(HE)to observe morphological changes and calculate radial alveolar count(RAC)values;the levels of malondialdehyde(MDA)and superoxide dismutase(SOD)in blood and lung tissues were determined to evaluate the imbalance of oxidative and antioxidant homeostasis in neonatal rats;real-time fluorescence quantitative reverse transcription polymerase chain reaction(RT-qPCR)was used to detect the relative expression level of nuclear factor-erythroid 2-related factor 2(Nrf2)mRNA in lung tissues.Results①H group showed a gradual decline in activity after 3 days and an increasing trend in body weight after 14 days.②W/D value of H group reached its peak at day 14,and the trend of HC group was similar to that of H group.③The lung tissue structure of H group was irregular,RAC value of which decreased significantly after 7 days of peak,and the difference between H group and HC group was statistically significant(P<0.01).④The MDA value of H group increased on day 7 and gradually decreased on day 14,while the SOD activity decreased obviously on day 7,and there were significant differences in MDA and SOD activity between H group and HC group at 14 days and 21 days(P<0.05).⑤The expression of Nrf2 mRNA in H group was significantly enhanced at day 7 and stabilized at day 14,and there were statistically significant differences between H and HC groups at day 3,day 7 and day 14(P<0.05).⑥The relative expression level of Nrf2 mRNA was positively correlated with MDA and negatively correlated with SOD.Conclusion Caffeine can regulate oxidative stress response through Nrf2 pathway and alleviate lung oxidative stress injury induced by hyperoxia in neonatal rats with BPD.