METHODS: Human fibroblast cells WI-38 were treated with lipopolysaccharide (LPS). The viability of WI-38 cells was measured via Cell counting kit-8. Reverse transcription-quantitative polymerase chain reaction was used to evaluate the levels of interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α). Western blot analysis revealed the protein levels of IL-1β, IL-6, TNF-α, Bax, and cleaved-caspase 3. Flow cytometry was applied for exploring the apoptosis of WI-38 cells. The concentrations of IL-1β, IL-6, and TNF-α were assessed via enzyme-linked-immunosorbent serologic assay.
RESULTS: DHK modulated the viability of WI-38 cells in infantile pneumonia. Furthermore, we identified that DHK treatment inversely changed LPS induction-mediated elevation on the levels of inflammation biomarkers. Besides, DHK counteracted LPS-induced production of reactive oxygen species (ROS) in WI-38 cells. DHK also decreased LPS-induced elevation of WI-38 cells apoptosis and mediated the levels of apoptosis-associated indexes. Moreover, modulating sirtuin-1 (SIRT1) protein level was lowered by the induction of LPS, and was reversed by DHK treatment. In addition, DHK counteracted LPS induction-mediated elevation of p-p65 and phosphorylated inhibitor of nuclear factor kappa-B kinase subunit alpha (p-IκBα) protein levels.
CONCLUSIONS: DHK alleviated LPS-induced WI-38 cells inflammation injury in infantile pneumonia through SIRT1/NF-κB pathway. The results shed light on the implications of DHK on the prevention and treatment of infantile pneumonia.
方法:用脂多糖(LPS)处理人成纤维细胞WI-38。通过细胞计数试剂盒-8测量WI-38细胞的活力。逆转录-定量聚合酶链反应用于评估白细胞介素(IL)-1β的水平,IL-6和肿瘤坏死因子-α(TNF-α)。蛋白质印迹分析显示IL-1β的蛋白质水平,IL-6,TNF-α,Bax,和切割的半胱天冬酶3.流式细胞术用于探索WI-38细胞的凋亡。IL-1β的浓度,通过酶联免疫吸附血清学测定评估IL-6和TNF-α。
结果:DHK调节WI-38细胞在小儿肺炎中的活力。此外,我们发现DHK治疗对LPS诱导介导的炎症生物标志物水平的升高有相反的改变.此外,DHK抵消了LPS诱导的WI-38细胞中活性氧(ROS)的产生。DHK还降低了LPS诱导的WI-38细胞凋亡的升高,并介导了凋亡相关指标的水平。此外,调节沉默调节蛋白-1(SIRT1)蛋白水平通过LPS的诱导降低,并被DHK治疗逆转。此外,DHK可抵抗LPS诱导的p-p65升高和核因子κB激酶亚基α(p-IκBα)蛋白水平的磷酸化抑制剂。
结论:DHK通过SIRT1/NF-κB通路减轻LPS诱导的小儿肺炎WI-38细胞炎症损伤。结果揭示了DHK对预防和治疗婴儿肺炎的意义。