染料木黄酮抑制3T3-L1细胞成脂分化机制

目的：研究染料木黄酮是否通过雌激素受体介导影响3T3-L1前脂肪细胞成脂分化，并探讨其可能的机制。方法：以不同浓度染料木黄酮处理3T3-L1细胞，四甲基偶氮唑蓝（methyl thiazolyl tetrazolium，MTT）法测定细胞存活率；在3T3-L1前脂肪细胞成脂分化过程中分别加入不同浓度染料木黄酮、染料木黄酮和ICI182780（一种雌激素受体抑制剂）混合物及不同浓度雌二醇，油红染色观察分化结果，测定细胞甘油三酯（triglyceride，TG）含量；染料木黄酮、染料木黄酮和ICI182780混合物及不同浓度雌二醇处理诱导成熟后的脂肪细胞，测定培养液甘油含量；分别用染料木黄酮（30 μmol/L）、染料木黄酮（30 μmol/L）和ICI182780混合物干预细胞成脂分化，Western blotting法检测细胞外信号调节激酶（extracellular signal-regulated kinase，ERK）、p-ERK、腺苷酸活化蛋白激酶（adenosine monophosphate activated protein kinase，AMPK）、p-AMPK、激素敏感性脂肪酶（hormone sensitive lipase，HSL）、脂肪酸合成酶（fatty acid synthetase，FAS）蛋白表达量。结果：3T3-L1细胞存活率随染料木黄酮浓度的升高而降低，50～200 μmol/L染料木黄酮能抑制细胞生长（P＜0.01）；染料木黄酮能负向调节成脂分化后细胞胞浆内TG含量，在0～50 μmol/L浓度范围内呈剂量依赖关系，高剂量雌二醇（100 nmol/L）能下调TG含量；染料木黄酮能促进成熟脂肪细胞脂解，提高细胞培养液甘油浓度；染料木黄酮（30 μmol/L）能上调p-AMPK、HSL蛋白表达，下调FAS蛋白表达（P＜0.01），对ERK、p-ERK、AMPK表达量无明显影响（P＞0.05）。ICI182780（1 μmol/L）能部分阻断染料木黄酮（30 μmol/L）对p-AMPK的调节作用（P＜0.05）。结论：染料木黄酮能抑制3T3-L1细胞成脂分化，其机制可能是染料木黄酮一方面下调FAS蛋白表达，抑制脂肪合成，另一方面激活AMPK-HSL途径促进脂肪分解；染料木黄酮还可能通过非雌激素受体途径抑制3T3-L1细胞成脂分化。

Anti-adipogenic Effect of Genistein in 3T3-L1 Cells and Its Mechanism

Objective: To investigate the anti-adipogenic effect and underlying mechanism of genistein in 3T3-L1 cells
and its estrogen receptor (ER) mediated pathway. Methods: After treated with various concentrations of genistein, the
survival rate of 3T3-L1 cells was assessed by MTT assay. In addition, after treated with genistein, genistein-ICI182780,
or estrogen, differentiation degree and lipid accumulation of the cells were assessed by oil red staining and a TG assay kit,
and adipocyte lipolysis was assessed by a glycerin assay kit. After intervention by genistein or genistein-ICI182780, the
expression of extracellular signal-regulated kinase (ERK), phospho-extracellular signal-regulated kinase (p-ERK), adenosine
monophosphate activated protein kinase (AMPK), phospho-adenosine monophosphate activated protein kinase (p-AMPK),
hormone sensitive lipase (HSL), and fatty acid synthetase (FAS) were analyzed by Western blotting. Results: Genistein and
high-dose estrogen inhibited lipid accumulation of 3T3-L1 and increased lipolysis in adipocytes. The presence of ICI182780
partly decreased both functions of genistein. Furthermore, genistein increased the expression of p-AMPK and HSL, and
decreased the expression of FAS. The presence of ICI182780 could slightly suppress the increase of p-AMPK caused by
genistein intervention. Conclusion: Genistein can inhibit the adipogenesis and differentiation of 3T3-L1 cells via a non-ER
pathway through a mechanism which may be relate to p-AMPK, HSL and FAS.