| 摘要: | 近年來慢性呼吸道疾病在全世界的發生率以及嚴重度逐漸增加，成為急待正視的健康問題。慢性呼吸道疾病是由於長期暴露在吸菸煙霧、空氣汙染物、過敏原之下刺激呼吸道造成慢性發炎，進而導致呼吸道上皮重塑的現象，使得氣管壁增厚，增加氣流通過限制，影響病人的呼吸功能。許多報導指出在這些疾病中觀察到促發炎因子，同時也是誘導上皮細胞間質轉化(EMT)的主要細胞激素TGF-β表現量的增加，而EMT也被報導為慢性呼吸道疾病的重要病理機轉之一。近期研究也指出在呼吸道上皮觀察到細胞粒線體功能的受損和葡萄糖代謝的改變，並與呼吸道重塑和慢性發炎有關。然而，TGF-β是否會調控細胞能量代謝改變，並且與EMT相關目前仍不清楚。在本論文，我們發現TGF-β會誘導A549和BEAS-2B細胞進行EMT，包括細胞移動性增加，以及上皮和間質標幟表現量改變。TGF-β也會增加糖解酵素表現、乳酸產生、有氧糖解作用，並引起代謝再程序化。而這些作用與Smad3的磷酸化並結合到Snail、GLUT1和PFKFB3的啟動子區域有關，減弱Smad3表現會顯著降低TGF-β誘導的糖解酵素表現增加和EMT。透過2-deoxy-D-glucose (2DG)和降低PFKFB3表現抑制糖解作用會顯著抑制TGF-β所誘導EMT。而乳酸可以誘導肺部上皮細胞的Smad3磷酸化並結合到Snail的啟動子區域，並且給予2DG可以顯著抑制TGF-β誘導Smad3磷酸化以及轉錄活性增加。此外，TGF-β也可能透過活化AMPK促進TGF-β誘導Smad3磷酸化和Snail表現。總而言之，我們在本論文發現在肺部上皮細胞中，TGF-β透過Smad3誘導EMT時也會增加糖解作用進行，並透過乳酸增加Smad3轉錄活性促進EMT的發生，另一方面，TGF-β也可以誘導AMPK活化而促進EMT，形成TGF-β誘導肺部上皮細胞EMT的惡性循環。透過了解糖解作用參與在EMT以及呼吸道重塑的角色，能量代謝重整可能成為慢性呼吸道疾病具未來發展性的新穎治療標的。
Chronic respiratory diseases (CRDs) have been a major health problem with increasing incidence and severity worldwide. Common risk factors for CRDs include cigarette smoking, prolonged exposure to environmental stressors such as air pollutants. These factors are highly associated with chronic inflammation and respiratory epithelium remodeling which may consequently cause CRDs. Increasing evidence has indicated elevated level of pro-inflammatory cytokine, TGF-β, in these lung diseases. TGF-β is recognized as a prototypical cytokine promoting epithelial-to-mesenchymal transition (EMT), which is the crucial pathogenesis of inflammatory lung diseases. Recent studies have further showed that impaired mitochondrial function and altered glucose metabolism in airway epithelium are associated with lung inflammation and remodeling. However, the effect of TGF-β in cellular energy metabolism in lung epithelial cells and its association with EMT remain elusive. In this study, we showed that TGF-β treatment promoted EMT, supported via increasing cell motility and expression changes in epithelial and mesenchymal markers in lung epithelial cells such as A549 and BEAS-2B cells. Furthermore, TGF-β resulted in increasing glycolytic enzymes expression, lactate production, aerobic glycolysis, known as the Warburg Effect, and metabolic reprogramming. These effects were associated with Smad3 phosphorylation as well as the recruitment of Smad3 to Snail, GLUT1, and PFKFB3 promoter region. Silencing of Smad3 abrogated TGF-β’s enhancing effect on EMT and glycolytic enzymes expression. Inhibition of glycolysis via 2-deoxy-D-glucose (2DG) and knockdown of PFKFB3 significantly suppressed TGF-β-induced EMT. Moreover, lactate is capable of inducing Smad3 phosphorylation and the recruitment of Smad3 to Snail promoter region in lung epithelial cells, whereas these events induced by TGF-β were successfully inhibited via 2DG. In addition, TGF-β induces activation of AMPK, which also enhance Smad3 phosphorylation and Snail expression. In conclusion, we demonstrated that TGF-β/Smad3 signaling pathway induces EMT and increases aerobic glycolysis with lactate production, which in turn promotes Smad3 activity and further exacerbates EMT. Activation of AMPK may also contribute to TGF-β-induced EMT. Establishing the causal role of aerobic glycolysis in TGF-β-induced EMT and subsequent tissue remodeling may shed light on future development of novel CRDs’ treatments strategized via energy reprogramming approach. |
