Wang Jing, Liu Wei, Peng Yang, Li Wenbin, Cheng Tingting, Gao Chunfang, Mo Luxia, Zhao Zhen, Chang Liwen
Department of Pediatrics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Cell Physiol Biochem. 2014;33(4):953-66. doi: 10.1159/000358667. Epub 2014 Mar 31.
To explore the potential mechanism of the synergetic effect of α-lipoic acid with keratinocyte growth factor (KGF) on protecting alveolar epithelial type II cells (ATIICs) from hyperoxia-induced injury.
Primary culture of ATIICs from the Sprague-Dawley rat fetuses was examined under room air and 95% of O2. Various KGF concentrations (0 to 100 ng/mL) and 0.5 mM of α-lipoic were added into the cell culture. Levels of intracellular reactive oxygen species, necrosis, and proliferation of ATIICs were measured using flow cytometry, ELISA, and MTT assays, respectively. RT-PCR was performed to detect KGFR mRNA expression. Western blot was employed to detect the expression of KGFR, phospho-p53, HDAC1, and acetylated H3 and H4.
KGF promoted the proliferation and inhibited the apoptosis of ATIICs in room air or under temporary exposure to hyperoxia. However, the resistance of ATIICs to KGF was observed after prolonged exposure. Further investigation demonstrated that down-regulation of KGF receptor via activation of p53 and recruitment of HDAC1 induced by oxidative stress contributed to KGF resistance. This resistance could be attenuated by α-lipoic acid, a powerful antioxidant.
Application of KGF combined with α-lipoic acid could inhibit KGF resistance to provide maximum protection to ATIICs from hyperoxic injury.
探讨α-硫辛酸与角质形成细胞生长因子（KGF）协同保护Ⅱ型肺泡上皮细胞（ATIICs）免受高氧诱导损伤的潜在机制。
对来自Sprague-Dawley大鼠胎儿的ATIICs进行原代培养，并在常氧和95%氧气条件下进行检测。将不同浓度的KGF（0至100 ng/mL）和0.5 mM的α-硫辛酸加入细胞培养物中。分别使用流式细胞术、酶联免疫吸附测定（ELISA）和MTT法检测ATIICs的细胞内活性氧水平、坏死情况和增殖情况。进行逆转录聚合酶链反应（RT-PCR）以检测KGFR mRNA表达。采用蛋白质免疫印迹法检测KGFR、磷酸化p53、组蛋白去乙酰化酶1（HDAC1）以及乙酰化H3和H4的表达。
在常氧或短暂暴露于高氧条件下，KGF可促进ATIICs的增殖并抑制其凋亡。然而，在长时间暴露后观察到ATIICs对KGF产生了抗性。进一步研究表明，氧化应激诱导的p53激活和HDAC1募集导致KGF受体下调，从而导致KGF抗性。一种强大的抗氧化剂α-硫辛酸可减弱这种抗性。
联合应用KGF和α-硫辛酸可抑制KGF抗性，为ATIICs提供最大程度的保护，使其免受高氧损伤。
