Cigarette smoking is the chief etiological factor for chronic obstructive pulmonary disease (COPD). Oxidative stress induced by cigarette smoke (CS) causes protein degradation, DNA damage, and cell death, thereby resulting in acute lung injury (ALI). In this regard, autophagy plays a critical role in regulating inflammatory responses by maintaining protein and organelle homeostasis and cellular viability. Expression of autophagy-related proteins (ARPs) is regulated by the fork head box class O (FOXO) transcription factors. In the current study, we examined the role of FOXO family proteins—FOXO1 and FOXO3a—in regulating CS extract (CSE)-induced autophagy. Using human lung adenocarcinoma cells with type II alveolar epithelial characteristics (A549), we observed CSE-mediated downregulation of FOXO3a. In contrast, there was a pronounced increase in the expression of FOXO1 at both the transcriptional and translational levels in the CSE-challenged cells compared with controls. Interestingly, knockdown of FOXO3a heightened the CSE-mediated increase in expression of cytokines/chemokines (IL-6, IL-8, and MCP-1), ARPs, and the FOXO1 transcription factor. Moreover, FOXO1 knockdown rescued CSE-mediated upregulation of ARPs in A549 cells. In addition, using the ROS inhibitor N-acetyl-L-cysteine (NAC), we observed abrogated mRNA expression of several ARPs and production of inflammatory cytokines/chemokines (IL-6, IL-8, MCP-1, and CCL-5) in the CSE-challenged cells suggesting an important role of ROS in regulating CSE-induced autophagy. Chromatin immunoprecipitation of FOXO1 and FOXO3a demonstrated increased binding of the former to promoter regions of autophagy genes- BECLIN1, ATG5, ATG12, ATG16, and LC3 in CSE challenged cells. These findings suggest the role of FOXO1 in regulating the expression of these genes during CSE exposure. Overall, our findings provide evidence for FOXO3a-dependent FOXO1-mediated regulation of autophagy in the CSE-challenged cells.

吸烟是慢性阻塞性肺疾病（COPD）的主要病因。香烟烟雾 (CS) 引起的氧化应激会导致蛋白质降解、DNA 损伤和细胞死亡，从而导致急性肺损伤 (ALI)。在这方面，自噬通过维持蛋白质和细胞器的稳态以及细胞活力在调节炎症反应中起关键作用。自噬相关蛋白 (ARPs) 的表达受叉头盒 O 类 (FOXO) 转录因子的调节。在目前的研究中，我们检查了 FOXO 家族蛋白——FOXO1 和 FOXO3a——在调节 CS 提取物 (CSE) 诱导的自噬中的作用。使用具有 II 型肺泡上皮特征的人肺腺癌细胞 (A549)，我们观察到 CSE 介导的 FOXO3a 下调。相比之下，与对照相比，CSE 攻击细胞中 FOXO1 的表达在转录和翻译水平上均显着增加。有趣的是，FOXO3a 的敲低增强了 CSE 介导的细胞因子/趋化因子（IL-6、IL-8 和 MCP-1）、ARP 和 FOXO1 转录因子表达的增加。此外，FOXO1 敲低挽救了 CSE 介导的 A549 细胞中 ARPs 的上调。此外，使用 ROS 抑制剂 N-乙酰基-L-半胱氨酸 (NAC)，我们观察到几种 ARPs 的 mRNA 表达消失和炎性细胞因子/趋化因子（IL-6、IL-8、MCP-1 和 CCL-5）的产生) 在 CSE 攻击的细胞中表明 ROS 在调节 CSE 诱导的自噬中的重要作用。CSE 攻击细胞中的BECLIN1、ATG5、ATG12、ATG16和LC3。这些发现表明 FOXO1 在 CSE 暴露期间调节这些基因的表达中的作用。总体而言，我们的研究结果为 FOXO3a 依赖性 FOXO1 介导的 CSE 攻击细胞自噬调节提供了证据。