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目的探讨牛黄熊脱氧胆酸(tauroursodeoxycholic acid,TUDCA)对TNF-α刺激3T3-L1细胞脂肪分解及其可能的机制。方法以3T3-L1前脂肪细胞经1-甲基-3-异丁基-黄嘌呤、地塞米松、胰岛素诱导分化成3T3-L1脂肪细胞,设:①对照组;②TNF-α(50 ng/ml)组;③TUDCA(1 mmol/L)+TNF-α(50 ng/ml)组;④TUDCA组(1 mmol/L)。通过油红O染色观察脂滴形态变化并测定培养基中甘油含量作为评价脂肪分解的指标,同时用Western blot检测脂滴包被蛋白perilipinA蛋白表达。结果 TNF-α可以显著刺激3T3-L1细胞脂肪分解,包括功能学显示甘油释放量显著增加[(2.784±0.104)mmol/L,P<0.01],形态学显示脂滴碎裂;TUDCA可以明显抑制TNF-α刺激的3T3-L1细胞脂肪分解,包括阻止脂滴形态的改变,逆转甘油的释放[(1.718±0.085)mmol/L,P<0.01]。通过Western blot检测结果显示TNF-α可以下调perilipin A蛋白表达[(0.227±0.004),P<0.01],而TUDCA可以阻断TNF-α对perilipin A蛋白表达的下调作用[(0.705±0.066),P<0.01]。结论 TUDCA可能通过阻止perilipin A蛋白下调而抑制TNF-α诱发脂肪分解,通过减少游离脂肪酸FFA,从而对胰岛素抵抗、糖尿病等具有一定的改善作用。
Objective To investigate the effects of TUDCA on lipolysis of 3T3-L1 cells stimulated by TNF-α and its possible mechanism. Methods 3T3-L1 preadipocytes were induced to differentiate into 3T3-L1 adipocytes by 1-methyl-3-isobutyl-xanthine, dexamethasone and insulin. ①Thirty- ml); ③TUDCA (1 mmol / L) + TNF-α (50 ng / ml); ④TUDCA group (1 mmol / L). The lipid droplet morphology was observed by oil red O staining and the glycerol content in the medium was determined as an indicator of lipolysis. At the same time, the protein expression of perilipin A was detected by Western blot. Results TNF-α significantly stimulated the lipolysis of 3T3-L1 cells, including a significant increase in glycerol release ([(2.784 ± 0.104) mmol / L, P <0.01] Lipolysis of TNF-α-stimulated 3T3-L1 cells, including preventing lipid droplet morphological changes and reversing glycerol release [(1.718 ± 0.085) mmol / L, P <0.01]. The results of Western blot showed that TNF-α down-regulated the expression of perilipin A protein [(0.227 ± 0.004), P <0.01], while TUDCA blocked the down-regulation of perilipin A expression by TNF-α [(0.705 ± 0.066) P <0.01]. Conclusion TUDCA may inhibit TNF-α-induced lipolysis by preventing the down-regulation of perilipin A protein, and may improve insulin resistance and diabetes by reducing free fatty acid (FFA).