AMPK减轻香烟烟雾诱发的肺细胞衰老和肺气肿及其分子机制

Cigarette smoke is a major risk factor for the development of chronic obstructive pulmonary disease (COPD), and it accounts for at least 75% of COPD deaths. No current drug therapy available reduces the relentless progression of COPD, which is due to lack of understanding of the mechanisms underlying the destructive processes in lungs of this disease. We and others have recently shown that COPD develops as a result of accelerated premature lung aging due to cellular senescence. The AMP-activated protein kinase (AMPK) has emerged as an important integrator of signals that control energy balance and lipid metabolism. Recent studies have shown that AMPK improves cellular stress resistance as well as protects against premature aging and cellular senescence. However, there are no studies regarding the regulation of AMPK in cigarette smoke-induced lung cellular senescence and subsequent emphysema/airspace enlargement. Our preliminary data have shown for the first time that the phosphorylation of AMPKα 1(Thr172), which reflects AMPK activity, was decreased in lung epithelium of COPD patients and in mice with pulmonary emphysema caused by cigarette smoke. Treatment with AMPK activator (AICAR) significantly reduced cigarette smoke extract-induced increase in senescence-associated β-galactosidase activity and pro-inflammatory cytokine IL-6 release in normal human small airway epithelial cells. Therefore, we hypothesize that AMPK protects against cigarette smoke-induced lung cellular senescence and inflammatory response, thereby prevents the progression of COPD/emphysema. To test this hypothesis, we will first determine the effects of AMPK on senescence and inflammatory mediator release in vitro in human small airway epithelial cells using AMPK activators, inhibitors, gene knockdown and overexpression. Next, the effects of AMPK on lung cellular senescence and injurious response will be investigated in vivo in mouse model of cigarette smoke-induced emphysema. Finally, the downstream signaling of AMPK in reducing lung cellular senescence and emphysema will be determined using the pharmacological and genetic approaches. The proposed study will provide the insights into novel molecular events that result in tobacco smoke-induced lung premature aging and emphysema. Importantly, the outcome of this study has high translational potential; such as knowledge will provide the basis for novel approaches to halt the progression of COPD.

慢性阻塞性肺病 (COPD)是一种以香烟烟雾吸入为主要病因的小气道疾病。目前其治疗药物均不能改善肺功能的降低。最近研究提示，肺上皮细胞特别是小气道上皮细胞经历衰老程序，从而损伤肺重新上皮化，以致启动或促进COPD的发生和发展。腺苷酸活化蛋白激酶 (AMPK) 是一类重要的蛋白激酶,除了参与能量和脂质代谢外，AMPK还能调节细胞衰老。我们前期数据首次表明，COPD患者和香烟烟雾引起肺气肿小鼠的肺上皮细胞中AMPK活性降低。另外，小分子AMPK激活剂抑制香烟提取物引起的小气道上皮细胞衰老和致炎因子IL-6产生。因此，我们推测AMPK活性降低参与了COPD的病理机制；激活AMPK信号能减轻COPD的进展。据此，本项目将从肺上皮细胞入手，利用小鼠COPD模型、采取分子遗传学和药理学手段，多层次地深入研究AMPK抗肺细胞衰老和COPD作用，为其防治COPD的临床应用提供重要的实验数据。

慢性阻塞性肺病 (COPD)是一种以香烟烟雾吸入为主要病因的小气道疾病。目前其治疗药物均不能改善肺功能的降低。最近研究提示，肺上皮细胞特别是小气道上皮细胞经历衰老程序，从而损伤肺重新上皮化，以致启动或促进COPD的发生和发展。腺苷酸活化蛋白激酶(AMPK) 是一类重要的蛋白激酶,除了参与能量和脂质代谢外，AMPK还能调节细胞衰老。我们前期数据首次表明，COPD患者和香烟烟雾引起肺气肿小鼠的肺上皮细胞中AMPK活性降低。另外，小分子AMPK激活剂抑制香烟提取物引起的小气道上皮细胞衰老和致炎因子IL-6产生。因此，我们推测AMPK活性降低参与了COPD的病理机制；激活AMPK信号能减轻COPD的进展。据此，我们研究了以下两方面的内容： 1）在细胞水平研究AMPK 是否抑制香烟提取物 (Cigarette smoke extract, CSE)诱发的正常人和COPD 患者小气道上皮细胞衰老和炎症介质产生。2) 在整体动物模型上研究AMPK是否减轻小鼠肺细胞衰老和肺气肿。结果显示: 1) AMPK 激活剂AICAR (1mM)显著降低CSE诱导的小气道上皮细胞 (SAEC)和支气管上皮细胞 (BEAS-2B)炎症介质　(IL-6和IL-8) 的释放和衰老基因 (p16，p21和 p66shc) mRNA的表达，而AMPK 抑制剂Compound C (5 µM)进一步加重上述的影响。2）基因抑制AMPKα1/α2显著增加BEAS-2B细胞衰老基因 (p16，p21和 p66shc) mRNA的表达。3）AMPK激活剂二甲双胍（50 和 250 mg/kg）预防性给药显著抑制，而Compound C (4 和20 mg/kg)加重弹性蛋白酶诱发的小鼠肺炎症反应，细胞衰老和肺泡扩大。4）AMPK激活剂二甲双胍（50 mg/kg）治疗性给药显著抑制弹性蛋白酶诱发的小鼠肺炎症反应，细胞衰老和肺泡扩大。结果表明，AMPK具有抑制肺炎症反应，抗肺细胞衰老和COPD作用，这为AMPK激活剂防治COPD的临床应用提供重要的实验数据。