Kuwano Kazuyoshi, Hagimoto Naoki, Maeyama Takashige, Fujita Masaki, Yoshimi Michihiro, Inoshima Ichiro, Nakashima Nobutaka, Hamada Naoki, Watanabe Kentaro, Hara Nobuyuki
Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Lab Invest. 2002 Dec;82(12):1695-706. doi: 10.1097/01.lab.0000045084.81853.76.
We previously demonstrated that the up-regulation of p53, Fas, and DNA damage are present in lung epithelial cells from patients with idiopathic interstitial pneumonias (IIP). Fas ligation induces apoptosis of lung epithelial cells predominantly through the direct activation of the caspase cascade via caspase-8 activation, whereas the up-regulation of p53 and other cellular stresses can induce mitochondria-mediated apoptosis. In this study, we investigated the incidence of mitochondria-mediated apoptosis of epithelial cells in IIP. We performed TUNEL staining to detect apoptotic cells and western blot analysis and immunohistochemistry to assess the expression and activation of caspases and the cytochrome c release from mitochondria in lung tissues from eight patients with usual interstitial pneumonia, five patients with nonspecific interstitial pneumonia, and eight patients with normal lung parenchyma. The expressions of pro- and cleaved caspase-8, 9, 3, and cytochrome c release from the mitochondria were all significantly increased in the lung tissues of IIP compared with normal lung parenchyma. The positive signals for caspases in epithelial cells were increased in IIP compared with normal lung parenchyma by immunohistochemistry. The results of TUNEL and electron microscopy suggested that apoptotic cells were predominantly epithelial cells. TUNEL-positive cells in % of epithelial cells were significantly increased in IIP compared with normal lung parenchyma, and significantly correlated with cytochrome c release from the mitochondria and with the expression of cleaved caspase-3 in epithelial cells. We conclude that mitochondria-mediated apoptosis may be involved in the pathophysiology of IIP.
我们之前证明,特发性间质性肺炎(IIP)患者的肺上皮细胞中存在p53、Fas上调以及DNA损伤。Fas配体主要通过经由半胱天冬酶-8激活直接激活半胱天冬酶级联反应来诱导肺上皮细胞凋亡,而p53的上调和其他细胞应激可诱导线粒体介导的凋亡。在本研究中,我们调查了IIP中上皮细胞线粒体介导凋亡的发生率。我们对8例普通间质性肺炎患者、5例非特异性间质性肺炎患者以及8例肺实质正常的患者的肺组织进行了TUNEL染色以检测凋亡细胞,并进行了蛋白质印迹分析和免疫组织化学以评估半胱天冬酶的表达和激活以及线粒体细胞色素c的释放。与正常肺实质相比,IIP肺组织中前体和裂解的半胱天冬酶-8、9、3的表达以及线粒体细胞色素c的释放均显著增加。通过免疫组织化学检测,与正常肺实质相比,IIP上皮细胞中半胱天冬酶的阳性信号增加。TUNEL和电子显微镜检查结果表明,凋亡细胞主要是上皮细胞。与正常肺实质相比,IIP中上皮细胞中TUNEL阳性细胞的百分比显著增加,并且与线粒体细胞色素c的释放以及上皮细胞中裂解的半胱天冬酶-3的表达显著相关。我们得出结论,线粒体介导的凋亡可能参与了IIP的病理生理学过程。