摘要
目的探讨连翘苷对无机粉尘诱导的人肺上皮细胞(A549细胞)炎症反应的抑制作用,探讨其通过调控NF-κB P65/TLR4/IL-1β信号通路的具体机制。方法使用人肺上皮细胞A549进行体外实验,将A549细胞分为对照组、粉尘暴露组、粉尘+连翘苷低剂量组、粉尘+连翘苷高剂量组。采用ELISA检测细胞中的炎症因子水平;通过CCK-8法评估细胞活性;DCFH-DA法检测细胞内活性氧(ROS)水平;TUNEL染色法检测细胞凋亡情况;Western Blot检测NF-κB P65和TLR4蛋白的表达。结果ELISA检测结果显示,相较对照组,粉尘暴露组的白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)、白细胞介素-1α(IL-1α)显著升高(P<0.05),而连翘苷处理组IL-1β、IL-6和IL-1α水平显著降低(P<0.05),其中高剂量组效果最佳。CCK8检测结果显示,相较对照组,细胞存活率在粉尘暴露组显著下降(P<0.05),连翘苷处理组的细胞存活率较粉尘暴露组升高(P<0.05),其中高剂量组的细胞存活率恢复最为显著(P<0.05)。TUNEL染色法检测结果显示,相较对照组,凋亡细胞数量在粉尘暴露组显著增加(P<0.05),连翘苷处理组凋亡细胞数量显著减少(P<0.05),其中高剂量组的凋亡细胞数量减少最为明显。DCFH-DA法检测结果显示,相较对照组,ROS水平在粉尘暴露组显著升高(P<0.05),在连翘苷处理组中,ROS水平显著降低(P<0.05),其中高剂量组的ROS水平降低最为显著。Western Blot结果显示,相较对照组,粉尘暴露组的NF-κB P65、TLR4、TRAF6蛋白表达水平显著上升(P<0.05),IκBα蛋白水平显著降低(P<0.05),在连翘苷处理组中,NF-κB P65、TLR4、TRAF6蛋白表达水平显著降低(P<0.05),IκBα蛋白水平显著升高(P<0.05),其中高剂量组的蛋白表达水平恢复最为明显(P<0.05)。结论连翘苷能够通过抑制NF-κB P65/TLR4/IL-1β通路改善无机粉尘诱导的肺上皮细胞炎症反应,表现出潜在的抗炎作用,可能为治疗粉尘相关肺部疾病提供新的药物选择。
Objective To explore the inhibitory effect of forsythiarin on the inflammatory response of human lung epithelial cells(A549 cells)induced by inorganic dust,and to explore its specific mechanism by regulating the NF-κB P65/TLR4/IL-1βsignaling pathway.Methods Human lung epithelial cells A549 were used for in vitro experiments,and A549 cells were divided into the control group,the dust exposure group,the dust+forsythin low-dose group,and the dust+forsythin high-dose group.ELISA was used to detect the levels of inflammatory factors in cells.Cell viability was assessed by CCK-8 method,intracellular reactive oxygen species(ROS)levels were detected by DCFH-DA method,cell apoptosis was detected by TUNEL staining,and the expression of NF-κB P65 and TLR4 was detected by Western blot.Results The ELISA test results showed that compared with the control group,interleukin-1β(IL-1β),interleukin-6(IL-6),and interleukin-1α(IL-1α)were significantly increased in the dust exposure group(P<0.05),while the levels of IL-1β,IL-6 and IL-1αin the forsythiarin-treated group were significantly reduced(P<0.05),among which the high-dose group had the best effect.The CCK8 test results showed that compared with the control group,the cell survival rate in the dust exposure group decreased significantly(P<0.05).The cell survival rate in the forsythin-treated group was higher than that in the dust exposure group(P<0.05).Among them,the cell survival rate recovery was the most significant in the high-dose group(P<0.05).The results of TUNEL staining showed that compared with the control group,the number of apoptotic cells in the dust exposure group was significantly increased(P<0.05),and the number of apoptotic cells in the forsythin-treated group was significantly reduced(P<0.05).Among them,the number of apoptotic cells in the high-dose group was the most obvious.The DCFH-DA method test results showed that compared with the control group,ROS levels were significantly increased in the dust exposure group(P<0.05),and in the forsythin-treated group,ROS levels were significantly reduced(P<0.05).Among them,the ROS levels were reduced most significantly in the high-dose group.Western Blot results showed that compared with the control group,the protein expression levels of NF-κB P65,TLR4,and TRAF6 in the dust exposure group were significantly increased(P<0.05),and the IκBαprotein level was significantly decreased(P<0.05).In the forsythin-treated group,medium,the protein expression of NF-κB P65,TLR4,and TRAF6 were significantly reduced(P<0.05),and the protein expression of IκBαwas significantly increased(P<0.05).Among them,the recovery of protein expression levels in the high-dose group was the most obvious(P<0.05).Conclusion Forsythin can improve the inflammatory response of lung epithelial cells induced by inorganic dust by inhibiting the NF-κB P65/TLR4/IL-1βpathway,showing potential anti-inflammatory effect and may provide new drugs for the treatment of dust-related lung diseases.
作者
赵娜
张玉龙
姚小青
ZHAO Na;ZHANG Yulong;YAO Xiaoqing(Department of Respiratory and Critical Care Medicine,Baoji Hospital of Traditional Chinese Medicine,Baoji,Shaanxi 721000,China)
出处
《临床肺科杂志》
2025年第4期559-565,共7页
Journal of Clinical Pulmonary Medicine
作者简介
通信作者:姚小青,E-mail:yxq631995929@163.com。
