Wang Hui, Yan Xiaopei, Xu Li
Pulmonary and Critical Care Medicine, Affiliated Suzhou Hospital of Nanjing Medical University/Suzhou Municipal Hospital, Suzhou 215000, China.
Pulmonary and Critical Care Medicine, Affiliated Suzhou Hospital of Nanjing Medical University/Suzhou Municipal Hospital, Suzhou 215000, China. *Corresponding author, E-mail:
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2024 May;40(5):411-418.
Objective To explore the effects of aloperine (Alo) on cigarette smoke-induced injury in human bronchial epithelial cells and its potential mechanism. Methods After human bronchial epithelial 16HBE cells were co-treated by 100 mL/L cigarette smoke extract (CSE) and various concentrations (50,100 and 200 μmol/L) of Alo, cell viability was assessed using CCK-8 assay. Lactate dehydrogenase (LDH) activity was measured with a related kit. Cell apoptosis was evaluated using the terminal-deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay (TUNEL) and Western blot analysis. The levels of inflammatory factors were detected by ELISA. Oxidative stress levels were assessed using 2'7'-dichlorofluorescin diacetate (DCFH-DA) staining. The expression of Toll-like receptor 4 (TLR4)/nuclear factor-kappaB (NF-κB)/NLR family pyrin domain containing 3 (NLRP3) signaling-associated proteins was measured by Western blot analysis. After cells were co-treated with 100 mL/L CSE and 200 μmol/L Alo, the aforementioned assays were applied to evaluate the effects of TLR4 overexpression on the TLR4/NF-κB/NLRP3 signaling, LDH activity, apoptosis, inflammatory response and oxidative stress in cells. Results CSE exposure might inhibit 16HBE cell viability, increase LDH activity, apoptosis, inflammatory response and oxidative stress levels and activate TLR4/NF-κB/NLRP3 signaling. Treatment with Alo promoted cell viability, decreased LDH activity, cell apoptosis, inflammation and oxidative stress levels, and inactivated TLR4/NF-κB/NLRP3 signaling. Furthermore, TLR4 overexpression might reverse the protective role of Alo treatment in CSE-induced injury in 16HBE cells. Conclusion Alo may ameliorate CSE-induced injury in human bronchial epithelial cells via inhibiting TLR4/NF-κB/NLRP3 signaling.
目的 探讨氧化苦参碱（Alo）对香烟烟雾诱导的人支气管上皮细胞损伤的影响及其潜在机制。方法 将人支气管上皮16HBE细胞与100 mL/L香烟烟雾提取物（CSE）及不同浓度（50、100和200 μmol/L）的Alo共同处理后，采用CCK-8法检测细胞活力。用相关试剂盒检测乳酸脱氢酶（LDH）活性。采用末端脱氧核苷酸转移酶介导的dUTP生物素缺口末端标记法（TUNEL）和蛋白质免疫印迹分析评估细胞凋亡。通过酶联免疫吸附测定（ELISA）检测炎症因子水平。用二氯荧光素二乙酸酯（DCFH-DA）染色评估氧化应激水平。通过蛋白质免疫印迹分析检测Toll样受体4（TLR4）/核因子-κB（NF-κB）/含NLR家族pyrin结构域3（NLRP3）信号相关蛋白的表达。细胞与100 mL/L CSE和200 μmol/L Alo共同处理后，应用上述检测方法评估TLR4过表达对细胞中TLR4/NF-κB/NLRP3信号、LDH活性、细胞凋亡、炎症反应和氧化应激的影响。结果 CSE暴露可能抑制16HBE细胞活力，增加LDH活性、细胞凋亡、炎症反应和氧化应激水平，并激活TLR4/NF-κB/NLRP3信号。Alo处理可促进细胞活力，降低LDH活性、细胞凋亡、炎症和氧化应激水平，并使TLR4/NF-κB/NLRP3信号失活。此外，TLR4过表达可能会逆转Alo处理对CSE诱导的16HBE细胞损伤的保护作用。结论 Alo可能通过抑制TLR4/NF-κB/NLRP3信号改善CSE诱导的人支气管上皮细胞损伤。
