Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida.
Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida
Am J Physiol Cell Physiol. 2014 Jun 1;306(11):C999-C1007. doi: 10.1152/ajpcell.00064.2013. Epub 2014 Apr 2.
Hyperoxic acute lung injury (HALI) is characterized by inflammation and epithelial cell death. CLOCK genes are master regulators of circadian rhythm also implicated in inflammation and lung diseases. However, the relationship of CLOCK genes in hyperoxia-induced lung injury has not been studied. This study will determine if HALI alters CLOCK gene expression. To test this, wild-type and NALP3(-/-) mice were exposed to room air or hyperoxia for 24, 48, or 72 h. In addition, mice were exposed to different concentrations of hyperoxia (50, 75, or 100% O2) or room air for 72 h. The mRNA and protein levels of lung CLOCK genes, based on quantitative PCR and Western blot analysis, respectively, and their target genes are significantly elevated in mice exposed to hyperoxia compared with controls. Alterations in CLOCK genes are associated with increased inflammatory markers in bronchoalveolar lavage fluid of hyperoxic mice compared with controls. Histological examination of mice lungs exposed to hyperoxia show increased inflammation and alveolar congestion compared with controls. Our results indicate sequential increase in CLOCK gene expression in lungs of mice exposed to hyperoxia compared with controls. Additionally, data suggest a dose-dependent increase in CLOCK gene expression with increased oxygen concentrations. To validate if the expression changes related to CLOCK genes are indeed associated with inflammation, NALP3(-/-) was introduced to analyze loss of function in inflammation. Western blot analysis showed significant CLOCK gene downregulation in NALP3(-/-) mice compared with wild-type controls. Together, our results demonstrate that hyperoxia-mediated lung inflammation is associated with alterations in CLOCK gene expression.
高氧性急性肺损伤（HALI）的特征是炎症和上皮细胞死亡。时钟基因是昼夜节律的主要调节因子，也与炎症和肺部疾病有关。然而，时钟基因在高氧诱导的肺损伤中的关系尚未得到研究。本研究将确定 HALI 是否改变时钟基因的表达。为了验证这一点，野生型和 NALP3（-/-）小鼠分别暴露于空气或高氧中 24、48 或 72 小时。此外，小鼠还暴露于不同浓度的高氧（50、75 或 100% O2）或空气 72 小时。基于定量 PCR 和 Western blot 分析，分别检测肺时钟基因的 mRNA 和蛋白水平，以及它们的靶基因，结果显示，与对照组相比，暴露于高氧中的小鼠的表达水平显著升高。与对照组相比，高氧小鼠的支气管肺泡灌洗液中时钟基因的改变与炎症标志物的增加有关。高氧暴露小鼠的肺部组织学检查显示，与对照组相比，炎症和肺泡充血增加。我们的结果表明，与对照组相比，暴露于高氧中的小鼠的肺时钟基因表达呈序贯增加。此外，数据表明，随着氧气浓度的增加，时钟基因的表达呈剂量依赖性增加。为了验证与时钟基因表达变化相关的炎症是否确实与炎症有关，我们引入了 NALP3（-/-）来分析炎症中功能丧失的情况。Western blot 分析显示，与野生型对照组相比，NALP3（-/-）小鼠的时钟基因表达显著下调。总之，我们的结果表明，高氧介导的肺炎症与时钟基因表达的改变有关。
