山楂多酚缓解苯并芘诱导的呼吸道上皮细胞炎性损伤活性

Hawthorn Polyphenols Relieve Benzo(a)pyrene-Induced Inflammatory Injury in Respiratory Epithelial Cells

目的:基于苯并芘(benzo(a)pyrene,BaP)暴露的人支气管上皮(16HBE)细胞模型评价山楂活性多酚(hawthorn bioactive polyphenols,HBPs)细胞保护活性及潜在机制。方法:基于BaP暴露的16HBE细胞模型采用活性追踪方法分离HBPs;噻唑蓝(methyl thiazolyl tetrazolium,MTT)法检测HBPs细胞毒性,CCK-8法检测HBPs对BaP诱导的16HBE细胞损伤的细胞保护活性;酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)法检测化合物1(HBP-1)对BaP诱导的16HBE细胞炎症因子(肿瘤坏死因子α(tumor necrosis factor alpha,TNF-α)、白细胞介素-1β(interleukin-1β,IL-1β)、IL-18、IL-10、IL-6)及细胞内活性氧(reactive oxygen species,ROS)的影响;流式细胞术检测HBP-1对BaP诱导的16HBE细胞凋亡的影响;Western blotting检测HBP-1对BaP诱导的16HBE细胞芳香烃受体(aryl hydrocarbon receptor,AhR)及核因子-κB(nuclear factor kappa-B,NF-κB)信号通路蛋白表达的影响。结果:从山楂中共分离出10个活性酚类化合物,其中,HBP-1对BaP暴露导致的16HBE细胞炎性损伤的细胞保护活性最佳。HBP-1能够显著抑制BaP诱导的16HBE细胞炎症因子(IL-1β、IL-18及TNF-α)过量分泌及细胞内ROS水平升高;并抑制BaP诱导的16HBE细胞凋亡;Western blotting结果显示,HBP-1能够抑制BaP诱导的16HBE细胞AhR及NF-κB信号通路的激活。结论:HBP-1可通过抑制AhR及NF-κB信号通路的激活,抑制BaP诱导的16HBE细胞氧化应激及炎症因子过量分泌,从而发挥其细胞保护活性。

Objective:The objective of this study is to evaluate the cytoprotective activity and potential mechanism of hawthorn bioactive polyphenols(HBPs)using human bronchial epithelial(16HBE)cells exposed to benzo(a)pyrene(BaP).Methods:HBPs were isolated by activity-guided separation,and their cytotoxicity was assessed by the methyl thiazolyl tetrazolium(MTT)method.The cytoprotective activity of HBPs against BaP-induced injury in 16HBE cells was determined by the cell counting kit-8(CCK-8)method.Enzyme-linked immunosorbent assay(ELISA)was performed to detect the effect of HBP-1 on BaP-induced inflammatory factors such as tumor necrosis factor alpha(TNF-α),interleukin-1β(IL-1β),IL-18,IL-10,IL-6 and reactive oxygen species(ROS)in 16HBE cells.Flow cytometry was utilized to explore the effect of HBP-1 on BaP-induced apoptosis in 16HBE cells.Western blotting was conducted to examine the effect of HBP-1 on protein expression related to the aryl hydrocarbon receptor(AhR)and nuclear factor kappa-B(NF-κB)signaling pathways in BaP-induced 16HBE cells.Results:A total of 10 bioactive phenolic compounds were isolated from hawthorn.Among them,HBP-1 exhibited the most significantly cytoprotective activity against BaP-induced inflammatory injury in 16HBE cells.HBP-1 significantly inhibited the oversecretion of IL-1β,IL-18 and TNF-αand the increase in ROS level and BaP-induced apoptosis in 16HBE cells.Western blotting results indicated that HBP-1 could inhibit the activation of the AhR and NF-κB signaling pathways in 16HBE cells induced by BaP.Conclusion:HBP-1 is able to inhibit BaP-induced oxidative stress and excessive production of inflammatory cytokines in 16HBE cells by suppressing the activation of the AhR and NF-κB signaling pathways,highlighting the cytoprotective effect of HBP-1 against BaP-induced damage.