Han L P, Sun B, Xie Y, Chen L M
Tianjin Metabolic Diseases Hospital & Tianjin Institute of Endocrinology, Tianjin Medical University, Key Laboratory of Hormones and Development (Ministry of Health), Tianjin Key Laboratory of Metabolic Diseases, Tianjin 300070, China.
Zhonghua Yi Xue Za Zhi. 2018 Aug 28;98(32):2591-2596. doi: 10.3760/cma.j.issn.0376-2491.2018.32.012.
To investigate the effect and mechanism of celastrol on free fatty acids (FFAs)-induced HepG2 cells. Cultured human HepG2 cells were transfected with toll-like receptor 4 (TLR4) siRNA, and the interference efficiencies were examined by real-time PCR. HepG2 cells were treated with FFAs and celastrol, and the untreated cells were used as a normal control (NC). Deposition of lipids in the HepG2 cells were visualized by Oil Red O staining. The protein expression of TLR4 and downstream inflammatory mediators [myeloid differentiation factor 88 (MyD88), nuclear factor (NF)-κBp65, interleukin (IL)-1β and tumor necrosis factor α (TNF-α)] in the HepG2 cells were determined by Western blotting. The significance of the data obtained was evaluated using analysis of variance (ANOVA). Red lipid droplets were extensively deposited in HepG2 cells after 0.5 mmol/L FFAs induction and significantly decreased in the celastrol-treated group. The protein expression of TLR4 and downstream inflammatory mediators (MyD88, NF-κBp65, IL-1β and TNF-α) in the FFAs-induced HepG2 cells increased significantly compared with those of the NC group (all <0.05), and were suppressed in TLR4 siRNA-treated and celastrol-treated group (TLR4: 0.69±0.14, 1.63±0.12 vs 2.46±0.23; MyD88: 1.21±0.12, 1.35±0.18 vs 1.62±0.19; NF-κBp65: 1.69±0.14, 1.54±0.36 vs 2.19±0.47; IL-1β: 1.51±0.16, 1.45±0.38 vs 1.82±0.27; TNF-α: 1.60±0.14, 1.41±0.29 vs 1.88±0.19) (all <0.01). Co-treatment with TLR4 siRNA and celastrol further reduced the expression of inflammation mediators compared with those of the TLR4 siRNA-treated group (MyD88: 1.09±0.23 vs 1.21±0.12; NF-κBp65: 1.24±0.20 vs 1.69±0.14; IL-1β: 1.28±0.31 vs 1.51±0.16; TNF-α: 1.10±0.29 vs 1.60±0.14) (all <0.01). Celastrol exerts its protective effect partly via inhibiting the TLR4-mediated signaling pathways in the steatotic HepG2 cells.
探讨雷公藤红素对游离脂肪酸（FFAs）诱导的HepG2细胞的作用及机制。将培养的人HepG2细胞用Toll样受体4（TLR4）小干扰RNA（siRNA）转染，通过实时聚合酶链反应（PCR）检测干扰效率。用FFAs和雷公藤红素处理HepG2细胞，未处理的细胞作为正常对照（NC）。通过油红O染色观察HepG2细胞中脂质的沉积情况。采用蛋白质免疫印迹法检测HepG2细胞中TLR4及下游炎症介质[髓样分化因子88（MyD88）、核因子（NF）-κBp65、白细胞介素（IL）-1β和肿瘤坏死因子α（TNF-α）]的蛋白表达。采用方差分析（ANOVA）评估所得数据的显著性。0.5 mmol/L FFAs诱导后，HepG2细胞中出现大量红色脂滴沉积，而雷公藤红素处理组脂滴明显减少。与NC组相比，FFAs诱导的HepG2细胞中TLR4及下游炎症介质（MyD88、NF-κBp65、IL-1β和TNF-α）的蛋白表达显著增加（均P<0.05），而在TLR4 siRNA处理组和雷公藤红素处理组中受到抑制（TLR4：0.69±0.14、1.63±0.12比2.46±0.23；MyD88：1.21±0.12、1.35±0.18比1.62±0.19；NF-κBp65：1.69±0.14、1.54±0.36比2.19±0.47；IL-1β：1.51±0.16、1.45±0.38比1.82±0.27；TNF-α：1.60±0.14、1.41±0.29比1.88±0.19）（均P<0.01）。与TLR4 siRNA处理组相比，TLR4 siRNA和雷公藤红素联合处理进一步降低了炎症介质的表达（MyD88：1.09±0.23比1.21±0.12；NF-κBp65：1.24±0.20比1.69±0.14；IL-1β：1.28±0.31比1.51±0.16；TNF-α：1.10±0.29比1.60±0.14）（均P<0.01）。雷公藤红素部分通过抑制脂肪变性的HepG2细胞中TLR4介导的信号通路发挥保护作用。
