龙胆环烯醚萜类成分对NAFLD细胞模型脂质积累及炎症的改善作用及机制

Improvement effects and mechanism of gentian iridoids on lipid accumulation and inflammation in NAFLD cell model

目的研究龙胆环烯醚萜类成分(gentian iridoids,GI)对非酒精性脂肪性肝病(NAFLD)细胞模型脂质积累及炎症的改善作用及机制。方法以游离脂肪酸(0.5 mmol/L)诱导的人肝癌HepG2细胞为NAFLD细胞模型。考察不同浓度(0.125、0.25、0.5、1、2、4 mg/mL)GI对HepG2细胞活力的影响。将HepG2细胞分为对照组、模型组和GI低、中、高浓度组(0.25、0.5、1 mg/mL),除对照组外,其余组均加入0.5 mmol/L游离脂肪酸进行造模。培养24 h后,考察HepG2细胞内脂滴形成的情况,检测细胞内三酰甘油(TG)含量,检测细胞上清液中天冬氨酸转氨酶(AST)、丙氨酸转氨酶(ALT)和炎症指标[白细胞介素1β(IL-1β)、IL-6、肿瘤坏死因子α(TNF-α)]的水平;检测细胞中固醇调节元件结合蛋白1c(SREBP-1c)、脂肪酸合成酶(FAS)mRNA和核因子κB(NF-κB)信号通路相关蛋白[NF-κB抑制蛋白α(IκBα)、NF-κB、磷酸化NF-κB(p-NF-κB)]的表达。结果HepG2细胞经不同浓度GI作用后,细胞活力均显著升高(P<0.01)。与模型组比较,GI各浓度组细胞内红色脂滴明显减少,细胞核萎缩不明显、体积大小正常;细胞内TG含量(低浓度组除外),细胞上清液中AST、ALT、IL-1β、IL-6(低剂量组除外)、TNF-α水平,细胞中SREBP-1c、FAS mRNA表达水平以及NF-κB蛋白磷酸化水平均显著降低(P<0.05或P<0.01),IκBα蛋白表达水平均显著升高(P<0.01)。结论GI可减少NAFLD细胞模型的脂质积累,减轻炎症反应,其作用机制可能与抑制SREBP-1c/FAS及NF-κB信号通路有关。

OBJECTIVE To study the improvement effects and mechanism of gentian iridoids(GI)on lipid accumulation and inflammation in non-alcoholic fatty liver disease(NAFLD)model.METHODS Human hepatoma HepG2 cells induced by free fatty acids(0.5 mmol/L)were selected as NAFLD cell models.The effects of GI at different concentrations(0.125,0.25,0.5,1,2,4mg/mL)on HepG2 cell viability were investigated.HepG2 cells were divided into control group,model group and low,medium and high concentration groups of GI(0.25,0.5 and 1 mg/mL).Except for control group,0.5 mmol/L free fatty acid was added in other groups for modeling.After 24 h of culture,the formation of lipid droplets was observed in HepG2 cells;the content of triglyceride(TG)was detected.The levels of aspartate aminotransferase(AST),alanine aminotransferase(ALT)and inflammation indexes [interleukin-1 β(IL-1 β),IL-6,tumor necrosis factor α(TNF-α)] in cell supernatant were determined.The mRNA expressions of sterol-regulatory element binding protein(SREBP-1c) and fatty acid synthase(FAS) as well as the expression of nuclear factor-κB(NF-κB)related proteins [NF-κB inhibitor protein α(IκBα),NF-κB,phosphorylated NF-κB(p-NF-κB)] were also detected.RESULTS After treated with different concentrations of GI,the cell viability of HepG2 cells was improved significantly(P<0.01).Compared with model group,the number of red lipid droplets in the cells of each concentration group of GI was significantly reduced,the nuclear atrophy was not obvious,and the size was normal;the content of TG(except for low concentration group),and the levels of ALT,AST,IL-1β,IL-6(except for low concentration group) and TNF-α in cell supernatant were all decreased significantly;m RNA expression of SREBP-1c and FAS and the phosphorylation level of NF-κB were also decreased significantly(P<0.05 or P<0.01);the protein expression of IκBα was increased significantly(P<0.01).CONCLUSIONS GI can reduce the lipid accumulation and inflammatory response of NAFLD cell model induced by free fatty acids,and its mechanism may be related to the inhibition of SREBP-1 c/FAS and NF-κB signaling pathway.