Dysregulation of CLOCK gene expression in hyperoxia-induced lung injury
Dysregulation of CLOCK gene expression in hyperoxia-induced lung injury
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DOI：
10.1152/ajpcell.00064.2013
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发表时间：
2014-06-01
影响因子：
5.5
通讯作者：
Kolliputi, Narasaiah
中科院分区：
文献类型：
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作者：
Lagishetty, Venu;Parthasarathy, Prasanna Tamarapu;Kolliputi, Narasaiah
高氧急性肺损伤（HALI）是以炎症和上皮细胞死亡为特征的疾病。CLOCK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，CLOCK基因在高氧肺损伤中的关系尚未研究。这项研究将确定HALI是否改变CLOCK基因表达。为了测试这一点，将野生型和NALP 3（-/-）小鼠暴露于室内空气或高氧24、48或72小时。此外，小鼠暴露于不同浓度的高氧（50，75，或100%O-2）或室内空气72小时。肺CLOCK基因的mRNA和蛋白质水平，分别基于定量PCR和Western印迹分析，以及它们的靶基因在暴露于高氧的小鼠与对照组相比显著升高。与对照组相比，CLOCK基因的改变与高氧小鼠支气管肺泡灌洗液中炎性标志物的增加有关。对暴露于高氧的小鼠肺的组织学检查显示，与对照组相比，炎症和肺泡充血增加。我们的研究结果表明，与对照组相比，暴露于高氧的小鼠肺中的CLOCK基因表达顺序增加。此外，数据表明随着氧浓度的增加，CLOCK基因表达呈剂量依赖性增加。为了验证与CLOCK基因相关的表达变化是否确实与炎症相关，引入NALP 3（-/-）来分析炎症中的功能丧失。蛋白质印迹分析显示，与野生型对照组相比，NALP 3（-/-）小鼠的CLOCK基因显著下调。总之，我们的研究结果表明，高氧介导的肺部炎症与CLOCK基因表达的改变有关。
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% O-2) 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 hyperoxiamediated lung inflammation is associated with alterations in CLOCK gene expression.