Sueki Akane, Matsuda Kazuyuki, Iwashita Chinami, Taira Chiaki, Ishimine Nau, Shigeto Shohei, Kawasaki Kenji, Sugano Mitsutoshi, Yamamoto Hiroshi, Honda Takayuki
Department of Laboratory Medicine, Shinshu University Hospital, Matsumoto, Japan; Department of Laboratory Medicine, Shinshu University School of Medicine, Matsumoto, Japan.
Department of Laboratory Medicine, Shinshu University Hospital, Matsumoto, Japan.
Biochem Biophys Res Commun. 2014 Oct 31;453(4):804-9. doi: 10.1016/j.bbrc.2014.10.022. Epub 2014 Oct 14.
Epithelial-mesenchymal transition (EMT) is associated with pulmonary fibrosis, including idiopathic pulmonary fibrosis (IPF). In this study, we investigated EMT of human pulmonary epithelial-derived cells (A549). A549 cells was either cultured by itself or co-cultured with THP-1 macrophages under normoxic (21% O2) and hypoxic (2% O2) conditions. We evaluated the presence of EMT by determining the expression of EMT markers, E-cadherin, vimentin, and fibronectin. To determine the role of TGF-β1 and IL-1β in EMT of the A549 cells, we analyzed the effects of blocking their activity with TGF-β1 inhibitor or IL-1β neutralizing antibody respectively. The A549 cells presented EMT when they were co-cultured with THP-1 macrophages. The EMT of the A549 cells co-cultured with THP-1 macrophages was exacerbated under hypoxia. In addition, the EMT were prevented by the addition of TGF-β1 type I receptor kinase inhibitor. The hypoxic condition increased the mRNA levels of TGF-β1 in A549 cells and THP-1 macrophages and that of IL-1β in THP-1 macrophages when each cells were co-cultured. Anti-IL-1β neutralizing antibody attenuated TGF-β1 secretion in co-culture media under hypoxic conditions. Thus, the IL-1β from THP-1 macrophages up-regulated the TGF-β1 from A549 cells and THP-1 macrophages, and then the TGF-β1 from both cells induced and promoted the EMT of A549 cells when they were co-cultured under hypoxia. Together, these results demonstrate that the interaction between type II pneumocytes and macrophages under hypoxia is necessary for the development of pulmonary fibrosis.
上皮-间质转化（EMT）与肺纤维化相关，包括特发性肺纤维化（IPF）。在本研究中，我们调查了人肺上皮来源细胞（A549）的EMT。A549细胞单独培养或在常氧（21% O₂）和低氧（2% O₂）条件下与THP-1巨噬细胞共培养。我们通过检测EMT标志物E-钙黏蛋白、波形蛋白和纤连蛋白的表达来评估EMT的存在。为了确定转化生长因子-β1（TGF-β1）和白细胞介素-1β（IL-1β）在A549细胞EMT中的作用，我们分别分析了用TGF-β1抑制剂或IL-1β中和抗体阻断其活性的效果。A549细胞与THP-1巨噬细胞共培养时呈现EMT。与THP-1巨噬细胞共培养的A549细胞在低氧条件下EMT加剧。此外，添加TGF-β1 I型受体激酶抑制剂可阻止EMT。当两种细胞共培养时，低氧条件增加了A549细胞和THP-1巨噬细胞中TGF-β1的mRNA水平以及THP-1巨噬细胞中IL-1β的mRNA水平。抗IL-1β中和抗体在低氧条件下减弱了共培养基中TGF-β1的分泌。因此，THP-1巨噬细胞产生的IL-1β上调了A549细胞和THP-1巨噬细胞产生的TGF-β1，然后当它们在低氧条件下共培养时，两种细胞产生的TGF-β1诱导并促进了A549细胞的EMT。总之，这些结果表明低氧条件下II型肺上皮细胞与巨噬细胞之间的相互作用对于肺纤维化的发生发展是必要的。
