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目的:通过培养3T3-L1前脂肪细胞,并诱导其分化至成熟,研究游离脂肪酸对脂肪细胞糖代谢的影响。方法:培养诱导3T3-L1脂肪细胞,用油红O染色鉴定并比较其形态结构的变化。LPS、EPA、SA、PA干预成熟脂肪细胞,收集不同时间的培养基,葡萄糖氧化酶法算出各组脂肪细胞的葡萄糖消耗量。用Western blot检测不同时间各组干预后细胞AMPK、GLUT4蛋白含量。结果:油红O染色鉴定成熟脂肪细胞胞浆中的脂滴染成红色,并出现“戒环”样结构;诱导分化第8天,90%以上细胞均分化成熟。含LPS、EPA、SA、PA的培养基作用于成熟脂肪细胞,随着时间的延长,显著抑制脂肪细胞对葡萄糖的吸收(P<0.05),同时,脂肪细胞AMPK、GLUT4蛋白含量在减少(P<0.05)。结论:游离脂肪酸可以诱导胰岛素抵抗的分子机制可能是通过胰岛素信号通路激活蛋白激酶(AMPK),进而影响GLUT4的蛋白表达,使脂肪细胞的葡萄糖吸收率减低,影响脂肪细胞的糖代谢。
OBJECTIVE: To study the effects of free fatty acids on glucose metabolism in adipocytes by culturing 3T3-L1 preadipocytes and inducing them to differentiate to mature. Methods: The induced 3T3-L1 adipocytes were identified by oil red O staining and their morphological changes were compared. LPS, EPA, SA, PA intervention of mature adipocytes, collect media at different times, glucose oxidase method to calculate the glucose consumption of each group of fat cells. Western blot was used to detect the content of AMPK and GLUT4 protein in each group at different time. Results: Oil red O staining identified lipid droplets in the mature adipocyte cytoplasm stained red, and the “ring” like structure; induced differentiation on the 8th day, more than 90% of cells were mature. The medium containing LPS, EPA, SA and PA had the effect on the mature adipocytes, and inhibited the glucose uptake by adipocytes (P <0.05) with the prolongation of time. At the same time, the content of AMPK and GLUT4 decreased <0.05). CONCLUSION: The molecular mechanism by which free fatty acid induces insulin resistance may be through the activation of protein kinase (AMPK) by insulin signaling pathway, which in turn influences the protein expression of GLUT4 and reduces the glucose uptake of adipocytes and affects the glucose metabolism of adipocytes.