CAJ | 학술논문

目的观察染料木素(GEN)对油酸(oleic acid)诱导的HEⅡ4肝细胞脂质沉积细胞模型的作用,以及对腺苷酸活化蛋白激酶(AMPK)磷酸化的影响.方法采用MTT法检测不同浓度的GEN对细胞的毒性；以0.2 mmol/L油酸诱导HEⅡ4肝细胞内脂质大量沉积,给予GEN干预后以GPO·DAOS法检测细胞内甘油三酯(TG)含量,以DCTM法检测细胞内总蛋白含量,用蛋白含量校正细胞内TG含量(TG/蛋白)来评估细胞内脂质沉积情况；Western blotting法检测肝细胞内AMPK及P-AMPK (Thr172)蛋白表达水平,以P-AMPK (Thr172)/AMPK表示AMPK的磷酸化水平.结果 GEN可减少肝细胞内脂质沉积(分别为139.64±18.60、192.20±34.17,P＜0.05),并且提高细胞内AMPK的磷酸化水平(分别为1.93±0.36、0.78±0.27,P＜0.05),而GEN降低细胞内脂质沉积作用可被AMPK阻滞剂CompoundC所逆转.结论 GEN可改善油酸诱导的HEⅡ4肝细胞内脂质沉积,其作用机制与增加AMPK磷酸化水平有关.
목적 관찰염료목소(GEN)대유산(oleic acid)유도적HEⅡ4간세포지질침적세포모형적작용,이급대선감산활화단백격매(AMPK)린산화적영향.방법 채용MTT법검측불동농도적GEN대세포적독성；이0.2 mmol/L유산유도HEⅡ4간세포내지질대량침적,급여GEN간예후이GPO·DAOS법검측세포내감유삼지(TG)함량,이DCTM법검측세포내총단백함량,용단백함량교정세포내TG함량(TG/단백)래평고세포내지질침적정황；Western blotting법검측간세포내AMPK급P-AMPK (Thr172)단백표체수평,이P-AMPK (Thr172)/AMPK표시AMPK적린산화수평.결과 GEN가감소간세포내지질침적(분별위139.64±18.60、192.20±34.17,P＜0.05),병차제고세포내AMPK적린산화수평(분별위1.93±0.36、0.78±0.27,P＜0.05),이GEN강저세포내지질침적작용가피AMPK조체제CompoundC소역전.결론 GEN가개선유산유도적HEⅡ4간세포내지질침적,기작용궤제여증가AMPK린산화수평유관.
Objective To observe the effects of genistein(GEN)on oleic acid(OA)induced lipid accumulationin in H4 ⅡE cells and to discuss the possible mechanism of GEN in the pointof AMPK.Methods H4ⅡE cells were cultured in vitro.The control group(NOR),OA treatment group(MOD group)and GEN treatment group were established according to the experimental requirements.The effects of GEN on the proliferation of H4 Ⅱ E cells were measured with MTT assay.The intraeellular TG mass was quantified spectrophotometrically using TG test kit.Cell protein was determined by DCTM Protein Assay kit.The intracellular TG concentration which was used to evaluate lipid accumulation was corrected using protein content as an internal standard.Western blotting was applied to determine the expression of AMPK and P-AMPK (Thr172).Accordingly the phosphorylation levels of AMPK was by means of P-AMPK(Thr172)/AMPK.Results OA treatment can induce lipid accumulation in H4 Ⅱ E cells while GEN treatment can decrease the intracellular TG concentration through up-regulate the phosphorylation levels of AMPK,though the effect was blocked by Compound C that is the inhibitor of AMPK.Conclusion GEN has anti-accumulation of lipid effect in H4ⅡE cell.The mechanism of GEN protective effect is partially due to AMPK.