Department of Respiratory Intensive Care Unit (ICU), The First Affiliated Hospital of Zhengzhou University, No. 1 JianShe East Road, Zhengzhou, 450000, Henan Province, China.
Inflammation. 2021 Oct;44(5):1969-1981. doi: 10.1007/s10753-021-01474-3. Epub 2021 May 20.
Sepsis-induced lung injury was the most common cause of death in patients. This study aimed to investigate whether PD-L1 regulates the inflammation in LPS-induced lung epithelial cells and vascular endothelial cells by interacting with the HIF-1α signaling pathway. Sepsis-induced lung injury mice were constructed by cecal ligation and puncture (CLP) procedure, and lipopolysaccharide (LPS)-induced lung epithelial cells and vascular endothelial cells simulate the sepsis-induced lung injury model in vitro. Hematoxylin-eosin (HE) staining detected the morphological changes of the lung tissues, and immunohistochemistry (IHC) detected the PD-L1 expression in lung tissues. Bicinchoninic acid (BCA) assay determined the protein concentration in bronchial alveolar lavage fluid (BALF). The number of PD-1 (+) cells in blood was detected by flow cytometry. The apoptosis in lung tissues and LPS-induced cells was analyzed by TUNEL assay. The inflammatory factor levels and HIF-1α in lung tissues and LPS-induced cells were analyzed by ELISA. The transfection effects of KD-PDL1 or KD-HIF1A in lung epithelial cells and vascular endothelial cells were confirmed by qRT-PCR analysis. The protein expression related to the PD-L1- and HIF-1α-related pathway was determined by Western blot analysis. As a result, LMT-28, as an IL-6 inhibitor, alleviated lung injury and suppressed the apoptosis and inflammation in lung tissues in BALF and the number of PD-1 (+) cells in blood. Sepsis-induced lung injury activated the PD-L1- and HIF-1α-related pathway, while LMT-28 could not completely inhibit the pathway. In addition, downregulation of PD-L1 or downregulation of HIF-1α suppressed the apoptosis and alleviated the inflammation in LPS-induced lung epithelial cells and vascular endothelial cells. Downregulation of PD-L1 had significant effects on lung epithelial cells but had greater effects on vascular endothelial cells. Downregulation of HIF-1α could decrease PD-L1 expression, and downregulation of PD-L1 could also suppress the protein expression of HIF-1α and related pathways. In conclusion, downregulation of PD-L1 alleviated the inflammation in LPS-induced lung epithelial cells and vascular endothelial cells by suppressing the HIF-1α signaling pathway.
脓毒症诱导的肺损伤是患者死亡的最常见原因。本研究旨在探讨 PD-L1 是否通过与 HIF-1α 信号通路相互作用来调节 LPS 诱导的肺上皮细胞和血管内皮细胞中的炎症。通过盲肠结扎和穿孔 (CLP) 程序构建脓毒症诱导的肺损伤小鼠，并在体外模拟 LPS 诱导的肺上皮细胞和血管内皮细胞的脓毒症诱导的肺损伤模型。苏木精-伊红 (HE) 染色检测肺组织的形态变化，免疫组织化学 (IHC) 检测肺组织中 PD-L1 的表达。双缩脲 (BCA) 法测定支气管肺泡灌洗液 (BALF) 中的蛋白浓度。通过流式细胞术检测血液中 PD-1(+)细胞的数量。TUNEL 检测肺组织和 LPS 诱导细胞的凋亡。ELISA 检测肺组织和 LPS 诱导细胞中炎症因子水平和 HIF-1α。通过 qRT-PCR 分析肺上皮细胞和血管内皮细胞中 KD-PDL1 或 KD-HIF1A 的转染效果。通过 Western blot 分析确定与 PD-L1 和 HIF-1α 相关途径相关的蛋白表达。结果表明，IL-6 抑制剂 LMT-28 减轻了肺损伤，抑制了 BALF 中肺组织的凋亡和炎症以及血液中 PD-1(+)细胞的数量。脓毒症诱导的肺损伤激活了 PD-L1 和 HIF-1α 相关途径，而 LMT-28 不能完全抑制该途径。此外，下调 PD-L1 或下调 HIF-1α 可抑制 LPS 诱导的肺上皮细胞和血管内皮细胞的凋亡并减轻炎症。下调 PD-L1 对肺上皮细胞有显著影响，但对血管内皮细胞的影响更大。下调 HIF-1α 可降低 PD-L1 的表达，下调 PD-L1 也可抑制 HIF-1α 及其相关途径的蛋白表达。综上所述，下调 PD-L1 通过抑制 HIF-1α 信号通路缓解 LPS 诱导的肺上皮细胞和血管内皮细胞的炎症。
